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DNA replication
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During DNA replication in eukaryotes, would a given gene tend to always be replicated in the same direction?
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https://biology.stackexchange.com/questions/115932/during-dna-replication-in-eukaryotes-would-a-given-gene-tend-to-always-be-repli
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<p>DNA replication in humans starts from ~ 50,000 origin of replications<a href="https://www.nature.com/articles/nrm2976" rel="nofollow noreferrer"> [Méchali 2010]</a>. From each origin, replication then proceeds in both directions.</p>
<p>Taking the replisome running from left to right; the 5'–3' strand will be the "lagging" strand and the 3'–5' strand will be the "leading" strand.</p>
<p>Taking the replisome running from right to left; the 5'–3' strand will be the "leading" strand and the 3'–5' strand will be the "lagging" strand.</p>
<p>Now, imagine gene X which is just at the right of a constitutive origin of replication. It sounds to me that this gene will almost always be replicated by a replisome running from left to right (as the origin of replication is at its left), i.e. with the 5'–3' strand as lagging and 3'–5' strand as leading.</p>
<p>Is this the case? I cannot find papers exploring this idea but I may not be using the expected jargon/expression in my search. ChatGPT (of course) agrees with me but I do not really trust this guy...</p>
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<p>Yes.</p>
<p>Here is one somewhat randomly selected <a href="https://www.nature.com/articles/ncomms10208" rel="nofollow noreferrer">paper</a> describing the replication dynamics of genes.</p>
<p>The replication behavior additionally appears to be to some extent controlled by transcription (possibly not as much vice versa?), in that replication and transcription tend to move through genes in the same orientation:</p>
<blockquote>
<p>Initiation was routinely (70–90%) detected between active genes separated by >20 kb, but less often within smaller intergenes; a preference for divergent over tandem over convergent gene pairs was observed, particularly in small intergenes (Fig. 5g). This preferential initiation upstream of active genes contributed to a significant co-orientation of replication with transcription (Fig. 5h), extending previous findings in skew N-domains19. These data show that many dispersive initiation zones consist of large DNA segments precisely circumscribed by active genes. They are in agreement with early studies of a few model loci1,33,34,37 and with the recent key observation that transcription shapes a dramatic redistribution of the MCM2–7 complex, a core component of the replicative helicase marking potential origin sites, to exclude MCM2–7 from transcribed genes before S-phase entry in D. melanogaster38. However, this is probably not the single mechanism for specifying initiation zones.</p>
</blockquote>
<p>This provides one potential mechanism for the variable strength of different origins of replication.</p>
<p>I strongly recommend looking at their figure 5 and reading associated text.</p>
| 234
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DNA replication
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Can replication occur if DNA is methylated?
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https://biology.stackexchange.com/questions/10169/can-replication-occur-if-dna-is-methylated
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<p>Can a methylated strand of DNA be replicated without removing methylation? Does it make any difference if the strand is methylated or not (during replication)?</p>
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<p>Absolutely. It's a pretty cool process, actually. Most (<a href="http://www.sciencemag.org/content/341/6146/1237905" rel="noreferrer">well...</a>) DNA methylation occurs in the context of what are called <strong>CpG</strong>; that is, a <strong>C</strong> (Cytosine) followed by a <strong>G</strong> (Guanine). Because C and G are the Watson-Crick pair for each other, the sequence on the opposite strand will <strong>also</strong> be CG. Usually, both Cs are methylated, which turns out to be rather critical for maintenance of methylation.</p>
<p>DNA replication occurs through a semi-conservative mechanism, which means each old, original strand is copied and paired with a new strand. The new strand has no methylation on it, however; it is at this point that the enzyme DNA methyltransferase (DNMT1, specifically) comes into play. DNMT1 finds the CpGs methylated on one strand ("hemimethylated") and methylates the other strand, providing complete inheritance. <a href="http://www.fbmc.fcen.uba.ar/materias/genmol/bibliografia-1/review%20DNA%20methylation%20in%20mammals.pdf" rel="noreferrer">Here</a> is a nice summary, and <a href="http://www.igh.cnrs.fr/equip/mechali/publications/RevieworiginsNRCMB2010.pdf" rel="noreferrer">this</a> is a review that (briefly) summarizes the evidence that CpGs may play a role in irigins of replication (the jury's still out).</p>
| 235
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DNA replication
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What data would Meselson and Stahl have expected if DNA replication was dispersive rather than semiconservative?
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https://biology.stackexchange.com/questions/97796/what-data-would-meselson-and-stahl-have-expected-if-dna-replication-was-dispersi
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<blockquote>
<p>What data would Meselson and Stahl have expected if DNA replication was conservative rather than semiconservative?</p>
</blockquote>
<p>Answer:</p>
<p>In the first generation, there would be two bands, one of light density and one of heavy density. In the second generation there would still be two bands, one of light density and one of heavy density.</p>
<p>Question 1:</p>
<p>In the second generation, are the heavy-density bands getting closer to the end of the test tube, and the light-density bands closer to the top of the tube, than the first generation? (due to the larger number of DNA molecules)</p>
<blockquote>
<p>What data would Meselson and Stahl have expected if DNA replication was dispersive rather than semiconservative?</p>
</blockquote>
<p>Answer:</p>
<p>In the first generation, there would be one band, one of intermediate density. In the second generation there would still be one band, one of intermediate density.</p>
<p>Question 2:</p>
<p>Are the bands closer to the top of the test tube in the second generation than the first generation?
(Because DNA molecules would become lighter)
If this model was approved, could we see this change in the test tube? (Between first and second generation)</p>
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<p>A few baseline ideas should be understood. 1. The assumption is made that Meselson and Stahl's cell line at the start of the experiment contains DNA composed entirely of the <span class="math-container">$N_{15}$</span> isotope. 2. DNA with a greater proportion of <span class="math-container">$N_{15}$</span> isotope is denser than DNA with lesser proportion of <span class="math-container">$N_{15}$</span> isotope. 3. Higher density DNA sinks lower in the test tube than lighter density DNA. (Same density DNA sinks to the same level.)</p>
<p>Q1: Let's follow 100 parent (generation 0) cells. If DNA replication was conservative, after they first replication (generation 1), we would expect 200 DNA strands. 100 strands come from the "parent" cells and are entirely composed of <span class="math-container">$N_{15}$</span> isotope (the conserved template strand). The other 100 strands come from "daughter" cells and are entirely composed of <span class="math-container">$N_{14}$</span> isotope (the newly synthesized strand). After the second replication then, we would expect 400 DNA strands. 200 strands come from the "generation 1" cells and share their DNA composition: 100 strands of <span class="math-container">$N_{15}$</span> isotope and 100 strands of <span class="math-container">$N_{14}$</span> isotope. The other 200 strands come from "daughter" cells and are entirely composed of <span class="math-container">$N_{14}$</span> isotope. Thus, in both generations, there are only DNA strands of 100% <span class="math-container">$N_{15}$</span> isotope and 100% <span class="math-container">$N_{14}$</span> isotope. The only difference is that the second generation has proportionally more DNA <strong>STRANDS</strong> composed entirely <span class="math-container">$N_{14}$</span> isotope than the first. Because the proportion of <span class="math-container">$N_{15}$</span> isotope and <span class="math-container">$N_{14}$</span> isotope in an <strong>INDIVIDUAL</strong> strand determines the strand's density, and because the strands in both generations comprise of 100% <span class="math-container">$N_{14}$</span> isotope strands and 100% <span class="math-container">$N_{15}$</span> isotope strands, the bands visible in the two test tubes are at the same level. They do not move closer to the ends of tube.</p>
<p>Q2: Yes this is correct. The first generation's DNA would all be ~50% <span class="math-container">$N_{15}$</span> isotope/~50% <span class="math-container">$N_{14}$</span> isotope strands. The second generation's DNA would all be ~25% <span class="math-container">$N_{15}$</span> isotope/~75% <span class="math-container">$N_{14}$</span> isotope strands. Strands of proportionally less <span class="math-container">$N_{15}$</span> isotope are lighter and thus closer to the top of the test tube.</p>
| 236
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DNA replication
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DNA replication : binding
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https://biology.stackexchange.com/questions/44836/dna-replication-binding
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<p>Regarding an activator, does the C amp cap complex, bind to the coding strand, the template strand, or both?</p>
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<p>Both. You should note that activation in this case involves recruitment of RNA polymerase to drive transcription, and that this process is irrelevant to DNA replication. B in the figure shows amino acid contacts (from CAP) with specific nucleotides in the DNA duplex.<a href="https://i.sstatic.net/QcZBw.jpg" rel="nofollow noreferrer"><img src="https://i.sstatic.net/QcZBw.jpg" alt="enter image description here"></a></p>
<p>DNA binding by CAP: structure of the CAP-DNA complex.</p>
<p>(A) Structure of CAP in complex with its consensus DNA site (PDB 1RUN) [14], showing primary- and secondary-kink sites. CAP is in cyan; DNA and cAMP bound to CAP are in red. The crystallization DNA fragment contained a single-phosphate gap between positions 9 and 10 of each DNA half-site (Fig 1b).</p>
<p>(B) Summary of CAP-DNA interactions. Shaded boxes indicate positions at which CAP exhibits strong sequence preferences [11,15,16,17]. The black circle, black rectangles, and black diamonds indicate, respectively, the two fold-symmetry axis, the primary-kink sites; and the secondary-kink sites. The black vertical lines indicate the positions of single-phosphate gaps present in the crystallization DNA fragment. The cyan ovals and cyan circles indicate, respectively, amino acid-base contacts and amino acid-phosphate contacts.</p>
<p>Figure from:
Catherine L. Lawson, et al. 2004. Catabolite activator protein (CAP): DNA binding and transcription activation. Curr. Opin. Struct. Biol. 14:10.</p>
| 237
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DNA replication
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Error frequency of DNA replication without proof reading activity of DNA polymerase?
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https://biology.stackexchange.com/questions/77249/error-frequency-of-dna-replication-without-proof-reading-activity-of-dna-polymer
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<p>Different domains of DNA polymerase contain different activity, like <code>5'->3'</code> polymerisation and <code>3'->5'</code> proof reading activity (a general case), and these domains can be exploited separately to make them perform single activity at a time. For example if we are conducting detection of some sort of mutation on a strand of DNA, we need to suppress the activity of DNA polymerase proof reading, ie, stopping it to correct the error it makes.</p>
<p>Since the proof reading activity increases the fidelity of replication to some 99.9%, I want to know what would be the efficiency if there is no proof reading activity.</p>
<p>To simplify my question-
If DNA repair system commits 1 mistakes out of 10^8 base pairs it added in nucleotide chain, how many errors would occur if we remove it's proof reading mechanism?</p>
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<p>Klenow Fragment could be used to have a polymerase which lacks the exonuclease activity: not just the <code>5' -> 3'</code> but also the <code>3' -> 5'</code> .</p>
<p>Lacking the exonucleases activity, the only kind of control is given by the biochemical property of the enzyme: remember that the DNA polymerase has an active site similar to a right hand, which make possible to have correct couple between A,G,C,T. Then, according to this <a href="http://www.jbc.org/content/272/11/7345.full.pdf" rel="nofollow noreferrer">study</a>, the error rate is something around <code>1e-4</code>. </p>
| 238
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DNA replication
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Repair wrong DNA pair after ending replication?
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https://biology.stackexchange.com/questions/28015/repair-wrong-dna-pair-after-ending-replication
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<p>If there are wrong DNA pair, for example A=G, is there mechanism that could repair such things - <strong>after</strong> the replication was <strong>finished</strong>?</p>
<p>Or it happens only during replication?</p>
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<p>DNA mismatches and indels can be repaired after replication through an aptly named mechanism called <a href="http://en.m.wikipedia.org/wiki/DNA_mismatch_repair" rel="nofollow noreferrer">DNA mismatch repair</a>.</p>
<p>I'll briefly describe the mechanism in <em>Escherichia coli</em>, but it's similar in other organisms. A protein called MutS scans DNA for lesions caused by mismatched nucleotides incorporated during replication that escaped DNA polymerase proofreading. These are detected by the kink they produce in the DNA backbone. MutS recruits MutL and MutH; the former activates the latter, which is an endonuclease. MutH cuts the DNA backbone near the lesion in the newly synthesized strand (the one containing the misincorporated nucleotide). This strand can be identified because it has not yet been methylated, while the parent strand contains the original methylation. After cleavage, UvrD helicase unwinds the strand and an exonuclease digests it. The gap around the former lesion can then be filled by DNA polymerase III and DNA ligase. </p>
<p><img src="https://i.sstatic.net/Z1lHQ.jpg" alt="enter image description here"></p>
<p>[ <a href="http://commons.m.wikimedia.org/wiki/File:DNA_mismatch_repair.png" rel="nofollow noreferrer">image adapted from</a> ]</p>
<p>Here's an interesting review in a Nature publication that discusses it in more depth, as well as the mechanism in humans:
<a href="http://www.nature.com/cr/journal/v18/n1/full/cr2007115a.html" rel="nofollow noreferrer">Guo-Min L. 2008. Mechanisms and functions of DNA mismatch repair. Cell Res 18:85-98</a></p>
| 239
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DNA replication
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Are there other mechanisms for mutation besides imperfect DNA replication?
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https://biology.stackexchange.com/questions/2331/are-there-other-mechanisms-for-mutation-besides-imperfect-dna-replication
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<p>I was reading <a href="http://www.askamathematician.com/2012/05/q-is-quantum-randomness-ever-large-enough-to-be-noticed/">http://www.askamathematician.com/2012/05/q-is-quantum-randomness-ever-large-enough-to-be-noticed/</a> and saw:</p>
<blockquote>
<p>[...] the evolution of entire species can be changed by a single
mistake in the replication of a strand of DNA (this is one mechanism
for mutation).</p>
</blockquote>
<p>... implying other mechanisms for mutation exist. Do they? </p>
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<p>Absolutely. Ionizing (X-rays, neutrons, electrons, heavy ions) and non-ionizing radiation (UV), chemicals, etc. are able to induce DNA Damage, which is then imperfectly repaired. So it's not an issue of imperfect replication, but also of imperfect damage repair.</p>
| 240
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DNA replication
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Why doesn't telomerase activity cause DNA to get longer each time a cell undergoes DNA replication?
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https://biology.stackexchange.com/questions/39402/why-doesnt-telomerase-activity-cause-dna-to-get-longer-each-time-a-cell-undergo
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<p>Telomerase extends the ends of the lagging strands in order for all of DNA to be be copied. Doesn't this also mean that DNA gets progressively longer each time it undergoes replication? Why is this not the case?</p>
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<p>I will assume that you are referring to humans, though much of the research to elucidate telomerase function was performed in yeast.</p>
<p>The first reason is that only a small subset of somatic cells express telomerase. Most somatic cells are terminally differentiated and mitotically inactive, so they are not called upon to replicate their DNA and divide. And when they do, their telomeres get shorter due to the 3' end replication problem.</p>
<p>The end replication problem arises due to the biochemical limitations of DNA Polymerase. Pol requires a primer, 5' to 3', in order to bind to the template strand and begin to catalyze the elongation reaction. So when DNA Polymerase gets to the 3' end of the template strand, it is unable to catalyze the reaction because it doesn't have a free 3' hydroxyl. When the end RNA primer that was used to catalyze the Okazaki fragment just before the 3' end is removed, there is no place left on the template strand to prime, so there is no 3' hydroxyl to begin the elongation reaction and you are left with a 3' overhang of single-stranded DNA. </p>
<p>On the 5' end of the Template Strand, even though the polymerase is able to catalyze the reaction all the way to the end, the end of the chromosome is processed and nucleotides are removed to make a single-stranded 3' overhang on this end. The overhangs are necessary to do strand invasion to form the T-Loops of the telomeres. So every time a chromosome in a cell that has inactive telomerase replicates, it gets shorter.</p>
<p>For the cell types that have telomerase actively express, such as lymphocytes, partially-differentiated stem cells, and the precursor cells to sperm, there is a negative feedback loop based on the length of the telomeres, which in turn regulates the activity of telomerase. Telomerase maintains the ends of the chromosomes, especially in these mitotically active cells, but it will only elongate them to a certain point.</p>
<blockquote>
<p>It has long been known that telomere length is regulated by a negative feedback loop (reviewed in Smogorzewska and de Lange, 2004). Long telomeres contain more negative regulators that limit further telomere elongation by telomerase. Because of this, the enzyme preferentially acts on the shortest telomeres in the cell (Hemann et al., 2001; Zhu et al., 1998).
<strong>- Loayza, et. al.; Cell; Volume 117, Issue 3 30 April 2004</strong></p>
</blockquote>
| 241
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DNA replication
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Why AZT is selective towards HIV and doesn't impair human DNA replication?
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https://biology.stackexchange.com/questions/30053/why-azt-is-selective-towards-hiv-and-doesnt-impair-human-dna-replication
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<p>I've found <a href="http://www.pnas.org/content/71/12/4980.full.pdf" rel="nofollow noreferrer">this article</a>, which is a very old one (from the time when nucleoside analogs where researched as a possible way to prevent replication of virus genetic material, before the HIV epidemics).
In the last page, the authors suggest that cellular repair enzymes could remove azidothymidine (AZT, zidovudine) from cellular DNA if it's incorporated, but viruses' polymerases lack this ability.
As it's an old article, I would like to know if this hypothesis was confirmed. As both azidothymidine and thymidine can form normal base pairings, how could human enzymes detect an azidothymidine was incorporated?</p>
<p>If someone is not familiar with AZT mechanism: basically, it's an analog of thymidine which has an azide group instead of 3'-OH</p>
<p><img src="https://i.sstatic.net/dKoGi.jpg" alt="(picture)"></p>
<p>It can be phosphorylated by cell kinases and incorporated to the growing DNA chain; however, this will stop the polymerization because it depends on free 3'-OH. The natural question which follows is: why wouldn't AZT also block the replication of human cells' DNA?</p>
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<p>Retroviruses depend on being able to convert their RNA genome into a DNA copy, and have a reverse transcriptase enzyme to do that. This unique activity is not found in human cells, allowing for potential antiviral therapy if a drug can be used to inhibit the reverse transcriptase while not affecting the human enzymes. </p>
<p>AZT is one such drug, by mimicking the nucleotide thymidine, it can it can act as a substrate for nucleotide kinases and become 5' phosphorylated. AZT is phosphorylated by host cell enzymes, specifically thymidine kinase, this means that the levels of AZT phosphates are similar between HIV infected and non-infected cells. Once the 5' triphosphate is formed, the drug is a substrate for the viral reverse transcriptase. If an AZT is incorporated into the DNA backbone, it terminates the DNA because the 3' OH is not available for further polymerization.</p>
<p>However, at <a href="http://www.ncbi.nlm.nih.gov/pubmed/2430286">high enough doses</a> AZT can be toxic to healthy cells. One mechanism for toxicity is the depletion of thymidine, because AZT competes with thymidine for the kinase enzymes, but is phosphorylated at a slower rate. At high doses, >1mM, AZT can act as a substrate for human DNA polymerases. Thankfully, AZT is about 100 fold more selective for HIV reverse transcriptase than human DNA polymerase.</p>
<p>The mitochondrial DNA polymerase seems to be a little more sensitive to AZT than other human DNA polymerases. This can lead to toxicity in the heart, skeletal muscles, hepatocytes, or other cells that use a lot of mitochondria. This toxicity is generally reversible if doses are reduced.</p>
<p>For more information, read about other nucleotide analogs, such as <a href="http://en.wikipedia.org/wiki/Aciclovir">acyclovir</a>, <a href="http://en.wikipedia.org/wiki/Ganciclovir">ganciclovir</a>, and <a href="http://en.wikipedia.org/wiki/Reverse-transcriptase_inhibitor">others</a> which work through similar mechanisms.</p>
| 242
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DNA replication
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How does high-fidelity of DNA replication depend on the formation of hydrogen bonds?
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https://biology.stackexchange.com/questions/23243/how-does-high-fidelity-of-dna-replication-depend-on-the-formation-of-hydrogen-bo
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<p>Replication has an error rate of less than 1 in 100 million. DNA polymerase forms H-bond with the H-bond acceptor atoms in the minor groove. <-- enhance fidelity here?</p>
<p>Binding of the triphosphate group to the active site of DNA polymerase triggers a conformational change. Changing a conserved Tyr residue increases the error rate by 40 fold.</p>
<p>I don't quite understand the above two statements. Can anyone explain in detail to me? Thanks!</p>
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<p>DNA polymerase must catalyse the addition of 4 different nucleotides to the growing strand. This means that it cannot directly determine which base to incorporate at a specific point (how would it 'know' which base to incorporate and how it would it change its specificity for different bases). This means that the specificity for which base pair to incorporate is dependent on the template DNA strand.</p>
<p>Correct Watson-Crick base pairing (that is, hydrogen bonding) between the template strand and the nucleotide to be incorporated triggers the closing of the finger domain of DNAP around the primer-template junction and positions the latter in the optimal position for catalysis (with the $\ce{\alpha-PO_4}$ of the incoming nucleotide near the $\ce{3'-OH}$ of the primer for a nucleophilic attack catalysed by two $\ce{Mg^{2+}}$ ions). This is where the conserved tyrosine residue you mentioned comes into play. An incorrectly paired nucleotide will not trigger this conformational change and will not be positioned optimally, thus catalysis is less likely.</p>
<p>Furthermore, the DNA polymerase makes contacts with the minor groove of the primer-template junction through hydrogen bonds. This interaction is not base-specific (all Watson-Crick base pairs have he same pattern of hydrogen-bond acceptors in the minor groove) but only occurs when the correct nucleotide is incorporated, thus stabilising the complex. </p>
<p>Finally, discrimination between ribonucleotides and deoxyribonucleotides is done by steric exclusion of the $\ce{2'-OH}$ by amino acid residues in the binding pocket.</p>
<p>These factors can be thought of as kinetic proofreading as they simply slow down the reaction rate and provide time for an incorrectly paired nucleotide to dissociate. However, they can still be incorporated and many DNA polymerases have a $\ce{3'->5'}$ proofreading exonuclease that can remove incorrectly paired nucleotides. This proofreading is again mediated by interactions between the DNAP and the primer-template junction (ie hydrogen bonding with the minor groove). A weakened interaction due to an incorrectly paired base reduces the affinity between DNA and the catalytic site and increases the affinity between DNA and the proofreading site (because it has a preference for cleaving ssDNA from the 3' end).</p>
| 243
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DNA replication
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Are there any particular chemicals that initiate either DNA replication or Transcription ?
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https://biology.stackexchange.com/questions/77958/are-there-any-particular-chemicals-that-initiate-either-dna-replication-or-trans
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<p>When does the nucleus of a cell "know" when to bind DNA nucleotides ( for Replication ) or RNA nucleotides ( for Transcription ). From what i read, they're both structurally different and free nucleotides of both are present in the nucleoplasm. So when one's free nucleotides bind ( mRNA nucleotides for transcription for example ) why doesn't the other ( DNA nucleotides in this case )bind, or what keeps the other from doing so? </p>
<p>Are their any chemical prerequisites of some sort that initiate the processes?</p>
<p>TIA</p>
|
<p>Enzymes, nature’s catalysts, proteins that speed up chemical reactions. In this case, creating a linear polymer out of deoxribonucleotide monomers, to yield a polynucleotide (<em>DNA</em>). This reaction also requires a DNA template strand. Enzymes contain an active site that brings all of the substrates into alignment, and then key amino acid side chain residues that are located in the active site enhance the intended organic chemical reaction, often by stabilizing a transition state in one or more of the substrates.</p>
<p>You are correct that ribonucleotides and deoxyribonucleotides share many structural similarities, but enzyme active sites can indeed distinguish between the two molecules. Even if the wrong substrate diffuses into the active site it will usually not be able to participate in the reaction.</p>
| 244
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DNA replication
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Why there is replication of DNA before meiosis?
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https://biology.stackexchange.com/questions/65693/why-there-is-replication-of-dna-before-meiosis
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<p>It seems to me that, even without replication of DNA before meiosis, the homologous pairs can still do crossover, and then be pulled to opposite poles, directly forming 2 haploid gametes.</p>
| 245
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DNA replication
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Why is DNA replication not 100% accurate
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https://biology.stackexchange.com/questions/100165/why-is-dna-replication-not-100-accurate
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<p>I've been reading about DNA mismatch repair (MMR) and how this process improves DNA fidelity. However, I was wondering, what is stopping MMR from correcting all mistakes in the daughter DNA with 100% fidelity? Why is the error rate still around 1 in 10^9 base pairs? Is it because the MMR proteins aren't present in cells in a high enough concentration? What would you have to change about this process to achieve 100% fidelity?</p>
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<p>Nothing is 100% precise - any measurement or process allow for some error, the only difference is how often such errors occur (i.e., the probability of an error). These wildely range in biology - e.g., it is about 1 per <span class="math-container">$10^4$</span> in HIV replication, but only 1 per <span class="math-container">$10^9$</span> for human DNA (due to the repair mechanisms).</p>
<p>Note that <span class="math-container">$1$</span> in <span class="math-container">$10^9$</span> for human DNA means pretty much that there is about 1 error per each copying of the genome (which has the size of about <span class="math-container">$3\times10^9$</span>). One could therefore still pose a question of <em>why the error rate is so high?</em> E.g., Human body consists of about <span class="math-container">$15\times10^{12}$</span> cells. Thus, if the error rate were about 1 per <span class="math-container">$10^{22}$</span>(<span class="math-container">$\approx 3\times10^9$</span> by <span class="math-container">$15\times10^{12}$</span>), we could have human organisms consisting of the cells with identical DNA.</p>
<p>The answer to this is that errors are not necessarily bad: <em>the copying errors are the source of the mutations driving the evolution!</em> Note also that many of the errors have no effect at all on the well-being of the organism. Thus, it is fair to say that the existing error rates were <em>selected</em> by the evolution for assuring the appropriate rate of the evolutionary change/adaptation, without causing immediate harm to the organism. (One could even speculate that 1 error per genome copy is the appropriate rate across the organisms, but for a moment I cannot support this assertion by references).</p>
| 246
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DNA replication
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Why are telomeres needed to allow DNA replication at the ends of linear chromosomes?
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https://biology.stackexchange.com/questions/108144/why-are-telomeres-needed-to-allow-dna-replication-at-the-ends-of-linear-chromoso
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<p>I’m reading <em>Molecular Biology of the Cell</em> by Alberts et. Al and at one point the authors mention the following:</p>
<blockquote>
<p>We saw earlier that synthesis of the lagging strand at a replication
fork must occur discontinuously through a backstitching mechanism that
produces short DNA fragments. This mechanism encounters a special
problem when the replication fork reaches an end of a linear
chromosome. <strong>The final RNA primer synthesized on the lagging-strand
template cannot be replaced by DNA because there is no 3ʹ-OH end
available for the repair polymerase</strong>. Without a mechanism to deal with
this problem, DNA would be lost from the ends of all chromosomes each
time a cell divides [emphasis mine].</p>
</blockquote>
<p>They use this explanation as a motivation for why telomeres are needed. But I don’t quite understand it. Below is a diagram that they show earlier. It seems to me like the process shown can just as easily be done at the ends of chromosomes. In particular, if we imagine that the left-most primer has no DNA following it, it seems to me like polymerase can just do what it normally does to replicate the DNA until the end is reached. Where’s the problem?
<a href="https://i.sstatic.net/J5dAv.jpg" rel="nofollow noreferrer"><img src="https://i.sstatic.net/J5dAv.jpg" alt="enter image description here" /></a></p>
| 247
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DNA replication
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When does DNA replicate actually?
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https://biology.stackexchange.com/questions/81451/when-does-dna-replicate-actually
|
<p>I read on Wikipedia that when the cell enters prophase during mitosis , the DNA has already been duplicated , that is the DNA is replicated in the chromatin form , but here I see the picture which shows the already condensed DNA ( now chromosome ) divide into sister chromatids. When does DNA divide actually ?<br><img src="https://i.sstatic.net/o5Mll.png" alt="enter image description here"><img src="https://i.sstatic.net/vvuBq.png" alt="enter image description here"><br>link to wikipedia -<a href="https://en.wikipedia.org/wiki/Prophase" rel="nofollow noreferrer">https://en.wikipedia.org/wiki/Prophase</a></p>
|
<p>According to <a href="https://www.ncbi.nlm.nih.gov/pubmed/15838518" rel="nofollow noreferrer">This paper</a> and most biology textbooks, S-phase or synthesis phase is literally defined by the beginning and ending of DNA replication. The first picture does not accurately represent the nature of DNA replication and is instead a model to help people track DNA copies during stages of cell division. DNA is replicated as chromatin and condenses to "double" chromosome form. You can see some micrographs of DNA condensation in <a href="http://downloads.hindawi.com/journals/tswj/2006/814090.pdf" rel="nofollow noreferrer">this paper</a>. You'll see that they don't condense as single chromatids, but they are already replicated. </p>
| 248
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DNA replication
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What errors can occur during DNA replication?
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https://biology.stackexchange.com/questions/15800/what-errors-can-occur-during-dna-replication
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<p>When there is an error in copying DNA (a mutation), what exactly goes wrong?</p>
<p>If G goes with C and A goes with T, I don't see how that part can mess up.</p>
<p>Is the idea that when the double helix is split, an A gets ruined and replaced with a G by mistake, which then pairs with a C in one of the copies? So something that was <em>supposed</em> to be AT is now GC.</p>
|
<p>There are many way in which DNA can be damaged. As already pointed out in the comment by @skymninge, the <a href="http://en.wikipedia.org/wiki/DNA_repair" rel="nofollow noreferrer">Wikipedia page on DNA repair</a>, as well as the <a href="http://en.wikipedia.org/wiki/DNA_error" rel="nofollow noreferrer">mutation page</a> detail some of the things that can go wrong. </p>
<p>You say:</p>
<blockquote>
<p>If G goes with C and A goes with T, I don't see how that part can mess up.</p>
</blockquote>
<p>This, however, would imply that the four bases are completely different so that mispairing cannot exist. This is (may I add luckily?) not true.<br>
Indeed the chemical structure of the bases is very similar, and changes between one base and another (transversions and transitions) are common.</p>
<p><img src="https://upload.wikimedia.org/wikipedia/commons/thumb/3/35/Transitions-transversions-v3.png/480px-Transitions-transversions-v3.png" alt="Transversions and transitions"></p>
<p>These can result for instance from exposure to external agents such as <a href="http://en.wikipedia.org/wiki/Ionizing_radiation" rel="nofollow noreferrer">ionizing radiation</a> and <a href="http://en.wikipedia.org/wiki/Alkylating_agent#Alkylating_agents" rel="nofollow noreferrer">alkylating agents</a>, or from exposure to endogenous products such as <a href="http://en.wikipedia.org/wiki/Reactive_oxygen_species" rel="nofollow noreferrer">reactive oxygen species</a>.</p>
<p>Furthermore, the enzyme synthetizing DNA, called <a href="http://en.wikipedia.org/wiki/DNA_polymerase" rel="nofollow noreferrer">DNA polymerase</a>, can insert the wrong base, although most of these errors can be corrected by its "proof-reading mechanisms". However, it is estimated that</p>
<blockquote>
<p>the frequency at which human DNA undergoes lasting, uncorrected errors range from $1 * 10^{-4}$ to $1 * 10^{-6}$ mutations per gamete for a given gene. </p>
</blockquote>
<p>Source: <a href="http://www.nature.com/scitable/topicpage/dna-damage-repair-mechanisms-for-maintaining-dna-344" rel="nofollow noreferrer">Nature Scitable</a></p>
<p>So, in summary, the system is not perfect, and mutation can be introduced even endogenously. This may seem bad, but, at least for relatively small rates of mutation, is not, as it allows change in the population, which is the <a href="http://www.nature.com/scitable/knowledge/library/mutations-are-the-raw-materials-of-evolution-17395346" rel="nofollow noreferrer">basis for evolution</a>.</p>
| 249
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DNA replication
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When does histone synthesis occur in relation to DNA replication?
|
https://biology.stackexchange.com/questions/57533/when-does-histone-synthesis-occur-in-relation-to-dna-replication
|
<p>Do histones have to be synthesized before DNA is replicated to allow the DNA to coil around histones? </p>
|
<p>Yes, they have to. But that is just half of the story.</p>
<p>The (canonical) histones which are used in DNA replication are synthesized at the beginning of the S phase, and subsequently transported into the nucleus. Studies have shown that newly synthesized DNA is immediately packed into nucleosomes. Thus, it is necessary that these structures are available prior to (or at least just in time with) the replication.</p>
<p>However, there are different models on how the histones are inserted into new DNA. One model assumes that old and new nucleosomes are both incorporated into newly synthesized DNA after the replication fork. Others assume a semi-conservative approach where the old histones are disassembled into their subunits and then mixed together with the newly synthesized ones. For example, there is a model that proposes that the parental H2A/H2B and the H3/H4 dimers disassociate, while another assumes a disassociation of the H3/H4 dimers as well.</p>
<p><strong>Figure:</strong> Nucleosome synthesis models for DNA replication
<a href="https://i.sstatic.net/8pExg.png" rel="nofollow noreferrer"><img src="https://i.sstatic.net/8pExg.png" alt="enter image description here"></a></p>
<p><strong>References</strong></p>
<p><sup>a</sup> Angélique Galvani and Christophe Thiriet (2013). Replicating – DNA in the Refractory Chromatin Environment, The Mechanisms of DNA Replication, Dr. David Stuart (Ed.), InTech, DOI: 10.5772/52656. Available from: <a href="https://www.intechopen.com/books/the-mechanisms-of-dna-replication/replicating-dna-in-the-refractory-chromatin-environment" rel="nofollow noreferrer">https://www.intechopen.com/books/the-mechanisms-of-dna-replication/replicating-dna-in-the-refractory-chromatin-environment</a></p>
| 250
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DNA replication
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Why replication collapse but not stall leads to DNA break?
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https://biology.stackexchange.com/questions/71650/why-replication-collapse-but-not-stall-leads-to-dna-break
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<p>I have been looking into the concept of replication dynamics and was wondering why collapsing but not stalling leads to a DNA break. </p>
|
<p><em>Stalling precedes collapsing. And collapsing precedes breaking.</em> Hence, both stalling and collapsing precede, and lead to, breaking; but not every stalling leads to collapsing, and not every collapsing leads to breaking.</p>
| 251
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DNA replication
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Telomere shortening during replication
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https://biology.stackexchange.com/questions/13425/telomere-shortening-during-replication
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<p>It is widely know that each cell cycle during DNA replication some fraction of the telomeres is lost, and this phenomenon is called the end replication problem. Well this is due to the fact that the DNA polymerase only adds nucleotides in 5´--> 3´ direction, thus the synthesis of one of the two DNA strands will need some RNA primers for its polymerization (and DNA polymerase will replicate DNA in a "jumping" pattern). Then, in the very end of the synthesis of this strand (called lagging strand) one last fraction will not be copied from the template as even if the last primer is set in the very end of the chromosome, once this RNA is degraded a void space is going o remain invisible for DNA polymerase. </p>
<p>Well this is all well known, but my question is whether during each DNA replication it is ONLY the telomere of the chromatid whose origin was in the lagging strand which will be shortened? (Thus chromosome shortening is happening in a truly asymmetrical and stochastic pattern)</p>
|
<p>In replication, both the chromosomal halves (which are simultaneously threaded through the replication complex) have a lagging and a leading strand. A part of the segment will be replicated as leading and a part as lagging.<br>
<img src="https://i.sstatic.net/vdD7H.jpg" alt="enter image description here"></p>
| 252
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DNA replication
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How long would it take for DNA bases on a strand to become random if replication errors were not fixed?
|
https://biology.stackexchange.com/questions/116347/how-long-would-it-take-for-dna-bases-on-a-strand-to-become-random-if-replication
|
<p>How long would it take for the sequence of DNA bases on a chromosome to become random if replication errors were not repaired?</p>
<p>I ask this from an evolution point of view. When life on Earth began if DNA replicated but error checks were not in place, then presumably after a certain time there would be a random base arrangement on a strand of DNA and no useful proteins would be made.</p>
|
<p>Even without fixing errors, any errors that prevent the organism from reproducing will not be transmitted. So, important coding regions of DNA will never become random.</p>
<p>Either the mutation rate will be too high for survival and the organism will go extinct and not produce any DNA, or selection will keep the DNA non-random.</p>
| 253
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DNA replication
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Can a dividing cell that skipped DNA replication become cancerous?
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https://biology.stackexchange.com/questions/79603/can-a-dividing-cell-that-skipped-dna-replication-become-cancerous
|
<p>Let's assume that a cell fails to replicate its DNA during the <a href="https://en.wikipedia.org/wiki/S_phase" rel="nofollow noreferrer">S Phase</a> of the cell cycle. Let's also assume that the appropriate <a href="https://en.wikipedia.org/wiki/Cyclin-dependent_kinase" rel="nofollow noreferrer">CDKs</a> are inactive (perhaps due to mutation or lack of <a href="https://en.wikipedia.org/wiki/Cyclin" rel="nofollow noreferrer">cyclin</a> proteins etc.) and the <a href="https://en.wikipedia.org/wiki/G2-M_DNA_damage_checkpoint" rel="nofollow noreferrer">G2-M</a> checkpoint fails.</p>
<p>2 questions:</p>
<ol>
<li><p>Will the <a href="https://en.wikipedia.org/wiki/Spindle_checkpoint" rel="nofollow noreferrer">Spindle Checkpoint</a> fail due to not having duplicated DNA, or can mitosis complete and form <a href="https://en.wikipedia.org/wiki/Aneuploidy" rel="nofollow noreferrer">aneuploid</a> daughter cells?</p>
</li>
<li><p>More to the point, <strong>if mitosis <em>can</em> complete after a cell fails to replicate its DNA (irrespective of the answer to question 1), can the resulting cell (if viable) lead to a <a href="https://en.wikipedia.org/wiki/Neoplasm" rel="nofollow noreferrer">neoplasm</a> or cancer?</strong></p>
</li>
</ol>
<p>My thought is <em>no</em> because the odds of having subsequent generations of cells experience similar losses of DNA via mitosis of un-replicated cells would eventually lead to mostly non-viable next-generation cells and the "uncontrolled" division would cease.</p>
<ul>
<li><p>Though, admittedly, this assumption is dependent on the assumption that the original cell has a heritable issue that "disables" the appropriate checkpoints (vs some random fluke event) allowing the uncontrolled cell line to continue duplicating with un-replicated genomes.</p>
</li>
<li><p>I'm also not sure how ultrafine anaphase bridges (see <a href="http://www.pnas.org/content/113/39/E5757" rel="nofollow noreferrer">Moreno <em>et al.</em> 2016</a>) can be informative for this question.</p>
</li>
</ul>
<p>I found a paper (<a href="https://www.nature.com/articles/1208618" rel="nofollow noreferrer">Deshpande <em>et al.</em> 2005</a>) that discusses cyclins and cdks in development and cancer, but their paper focuses on the transition from G1 to the S phase and stops short of discussing mitosis of un-replicated cells</p>
<hr />
<p><em>Note: this might sound like a homework question, but <em>I'M</em> the one writing the assignment! In fact, it's an exam, and I'm trying to cover my bases for the answers I provide for a different cancer-related question to ensure there are not multiple correct answers.</em></p>
|
<p><strong>Question 1</strong></p>
<blockquote>
<p>Will the Spindle Checkpoint fail due to not having duplicated DNA, or
can mitosis complete and form aneuploid daughter cells?</p>
</blockquote>
<p>Yes it is possible. Meiosis 2 essentially does not involve DNA replication.</p>
<p>Perhaps you are only talking about mitotic divisions. I have found two reports that say that cells can replicate without DNA replication.</p>
<ol>
<li><a href="https://doi.org/10.1038/s41586-022-04641-0" rel="nofollow noreferrer">Chan et al. (2022)</a> show that zebrafish skin (superficial epithelial) cells can continue to "split" without DNA replication. This process apparently helps in rapid skin expansion.</li>
</ol>
<blockquote>
<p>Using time-lapse imaging, we found that many SECs readily divide on
the animal body surface; during a specific developmental window, a
single SEC can produce a maximum of four progeny cells over its
lifetime on the surface of the animal. Remarkably, EdU assays, DNA
staining and hydroxyurea treatment showed that these terminally
differentiated skin cells continue splitting despite an absence of DNA
replication, causing up to 50% of SECs to exhibit reduced genome size.</p>
</blockquote>
<ol start="2">
<li>A preprint by <a href="https://doi.org/10.1101/2020.07.08.193607" rel="nofollow noreferrer">Ganier et al. (2020; still unpublished)</a> reports a study on a triploid mouse embryonic fibroblast cell line (<a href="https://en.wikipedia.org/wiki/3T3_cells" rel="nofollow noreferrer">3T3</a>). They show that some daughter cells can receive 1N copy.</li>
</ol>
<blockquote>
<p>The fraction of unlicensed cells (24%) that divided produced two
daughter cells, despite the complete absence of DNA replication. If
the unlicensed genomes were equally distributed in the two daughter
cells, these cells should have 1C DNA content. In agreement, flow
cytometry analyses always highlighted a distinct population of
ΔDboxGeminin-expressing cells with 1C DNA content that appeared after
the first cell cycle (Figures 1A and 4A)</p>
</blockquote>
<p>They say that these cells can also re-enter mitosis.</p>
<blockquote>
<p>We removed DOX 24 hours after induction to stop ΔDboxGeminin induction
(T1, Figure 4F; control cells shown in Supplementary Figure 9) and
transfected these cells with a CDT1-encoding plasmid (or empty vector;
control) to counteract the effect of the remaining ΔDboxGeminin. 24h
after CDT1 transfection (T2), 15.3% of 1C cells were BrdU-positive
(Figure 4F, BrdU 24h, right panels) compared with 4.9% of 1C cells
transfected with empty vector.</p>
</blockquote>
<p><strong>Question 2</strong></p>
<blockquote>
<p>if mitosis can complete after a cell fails to replicate its DNA
(irrespective of the answer to question 1), can the resulting cell (if
viable) lead to a neoplasm or cancer?</p>
</blockquote>
<p>If the "asynthetic fission" occurs naturally in zebrafish, it may not be predominantly neoplastic. I think that a reduced genome would make the cells unable to re-replicate.</p>
<p>However, the 1N 3T3 cells can re-enter mitosis and they may undergo some cancerous transformation. This cell line is non-cancerous but is laboratory-generated/propagated. So we don't know about what happens <em>in vivo</em>. The authors did not check if the 1N cells that have re-entered mitosis, have cancer driver mutations in their genome.</p>
<p>Aneuploidy is indeed associated with cancers. Several cancer cells are aneuploid. Aneuploidy can result due to loss of stringency of cell cycle checkpoints (common in cancer cells). <a href="https://doi.org/10.1038/s41586-023-06266-3" rel="nofollow noreferrer">Shih et al. (2023)</a> show that aneuploidy can be a driver of tumorigenesis. Briefly, they show that somatic copy number alterations (SCNA) of entire chromosomal arms, can increase the fitness of cancer cells. These SCNAs include both expansions and deletions, and need not contain any oncogenes or tumor suppressor genes. For example,</p>
<blockquote>
<p>Chromosome arm 8p is frequently deleted across cancer types, but
canonical tumour suppressors have not been detected on 8p
(refs. 16,26). We observed less cell death by caspase activity
(P = 0.02) and flow cytometry (P = 0.004) in cells with engineered 8p
deletion (Extended Data Fig. 6c,d and Supplementary Fig. 2). Of the
three 8p del-pos peaks (Fig. 2d), the smallest peak, in 8p12,
contained two protein-coding genes, WRN and NRG1.</p>
</blockquote>
<p>This doesn't fully answer your question, I believe. So I end my answer with a few guesses.</p>
<p>It is quite likely that effect of aneuploidies on cancer are context dependent (<a href="https://doi.org/10.1038/s41576-019-0171-x" rel="nofollow noreferrer">Ben-David and Amon, 2019</a>). My gross expectation would be that loss of an entire chromosome should reduce cell fitness (loss of several genes). Gene essentiality would depend on cell type. It is possible that if essential genes are translocated from a chromosome to another, then the former can be dispensed. Cancer cells not only have aneuploidies but also frequent chromosomal rearrangements.</p>
<p>If one were to artificially stimulate cell division without replication, and that these cells don't have a perfect asymmetric division (2N, 0N), then aneuploidies can result: asymmetric segregation (xN, [2-x]N) followed by diploidization (<a href="https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10020200/" rel="nofollow noreferrer">common in haploid cell lines</a>). This can result in cancer.</p>
<p>However, it remains to be proven that an aneuploidy can indeed result in cancer.</p>
<hr>
<p><em>Perhaps this answer comes too late to be helpful for your assignment.</em></p>
| 254
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DNA replication
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Can DNA replicate without polymerase?
|
https://biology.stackexchange.com/questions/109269/can-dna-replicate-without-polymerase
|
<p>Would it be possible for short DNA molecules to replicate, for example, if it's heated to the point where the strands separate (as far as I know, that's what happens in PCR?) and freely floating bases could "connect" to their correspondent bases (A/T, C/G)?</p>
<p>I'm assuming that A/T and C/G bases strongly 'attract' each other naturally, perhaps that's where I'm wrong?</p>
|
<p>No, replication cannot happen in the absence of polymerase (on the timescales relevant to humans).</p>
<p>You are correct that in PCR, the first step is to separate DNA strands at 98 C. This heating dissociates the strands semi-permanently. So when you cool down the reaction, the strands do not re-associate very quickly.</p>
<p>When you cool down the reaction to 72 C, A/T and C/G can make temporary connections with their opposite bases. However, the free nucleotides cannot quickly form phosphodiester bonds with the growing chain of DNA in the absence of a polymerase. The nucleotide will most likely dissociate from the DNA strand before the correct bond forms. In other words, it is a kinetic problem.</p>
<p>However, at high temperatures, specialized conditions, and on long timescales, such as those present in earth's pre-biotic period, the answer may be different.</p>
| 255
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DNA replication
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Term for when part of DNA Strand is Reversed during Replication
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https://biology.stackexchange.com/questions/116252/term-for-when-part-of-dna-strand-is-reversed-during-replication
|
<p>I remember from my college genetics course that there is a type of mistake during DNA replicates that causes a section to be reversed. For example, if the original was sequence was 123456789, the resulting sequence would be 123487659. I was interesting in how this mistake would effect an organism's phenotype, but I came having trouble looking it up because I forget what it was called. Does anyone know?</p>
| 256
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DNA replication
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The relationship between the shape of the bacterial DNA and the blocking of replication machinery
|
https://biology.stackexchange.com/questions/53828/the-relationship-between-the-shape-of-the-bacterial-dna-and-the-blocking-of-repl
|
<p>I was reading a course about tolemers when I arrived to this phrase :</p>
<blockquote>
<p>[...] The ends of a linear DNA molecule cannot be replicated by the cellular replication machinery <strong>(which may be one reason why bacterial DNA molecules are circular)</strong>.</p>
</blockquote>
<p>I want to know what is the relationship between the circular shape of the bacterial DNA and the blocking of replication machinery ?</p>
|
<p>This has been covered elsewhere (I highly recommend <a href="https://www.boundless.com/biology/textbooks/boundless-biology-textbook/dna-structure-and-function-14/dna-replication-101/telomere-replication-438-11663/" rel="nofollow noreferrer">this page</a> ) but it basically has to do with the priming of the strands and the fact that all polymerases work from the 5' prime to the the 3'. I made a quick schematic to illustrate (the explanation is below):</p>
<p><a href="https://i.sstatic.net/8LFkm.png" rel="nofollow noreferrer"><img src="https://i.sstatic.net/8LFkm.png" alt="Schematic of DNA replication"></a></p>
<p>The priming is done with RNA during DNA replication in the cell, unlike PCR where you use DNA oligo's to prime. This RNA-DNA is called the Ozakazi fragments. After replication RNAse H will remove these primers leaving only DNA's and gaps. In the middle of the sequence the gaps can be filled in by the normal repair machinery but at the ends there is no 3' terminus to fill in the gaps. That is where the telomerase comes in and fill these gaps with a fixed sequence (also based on an RNA template but that is going into details).
If you have a circular chromosome there is always a 3' to complete the gap.</p>
| 257
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DNA replication
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Reason behind formation of positive supercoils during DNA replication/ transcription
|
https://biology.stackexchange.com/questions/56166/reason-behind-formation-of-positive-supercoils-during-dna-replication-transcrip
|
<p>When a twist is unwound without cutting the DNA strands or is removed by cutting the strand(s) and resealing, negative supercoils are introduced in the DNA.</p>
<p><a href="https://i.sstatic.net/o8fgc.png" rel="nofollow noreferrer"><img src="https://i.sstatic.net/o8fgc.png" alt="enter image description here"></a></p>
<p>From Cell and Molecular Biology -Karp</p>
<p>But strangely enough unwinding of DNA by helicase cause the DNA ahead of the primosome to form positive supercoils. Why is it so?</p>
<p><a href="https://i.sstatic.net/1cv9e.png" rel="nofollow noreferrer"><img src="https://i.sstatic.net/1cv9e.png" alt="enter image description here"></a></p>
<p>From: Molecular cell biology -Lodish</p>
<p><strong>My speculations:</strong> Underwinding cause the adjacent DNA to overwind and as the DNA overwinds the associated stress developed cause the DNA to form positive supercoils (which would have happened naturally had new twists been introduced in the DNA<sub> (Time:<a href="https://www.youtube.com/watch?v=HyP0cEbqKTc" rel="nofollow noreferrer">1:54</a>) </sub>).</p>
<p>I think removal of twist creates negative supercoils initially but when the unwinding causes too much overwinding of adjacent DNA, the strain developed cause the already negatively supercoiled DNA to form positive supercoils.</p>
|
<p>It's hard to explain in text, so here's a video:</p>
<p><a href="https://www.youtube.com/watch?v=J4YlcD59-yw" rel="nofollow noreferrer">https://www.youtube.com/watch?v=J4YlcD59-yw</a></p>
<p>Imagine the shoelaces are two DNA strands in a double helix. They are topologically constrained at each end. As the pen (helicase) moves through the helix, it creates overwound DNA in front of it and underwound DNA behind it.</p>
| 258
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DNA replication
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How can DNA replication result in hair pin structures?
|
https://biology.stackexchange.com/questions/92947/how-can-dna-replication-result-in-hair-pin-structures
|
<p>My professor said that one of the reasons SSB proteins are so important was to prevent the formation of hair pin structures, I can't see how or why DNA would form hairpin structures and there's not much about it on the internet so can anybody explain this hair pin thing and how SSB proteins prevent it from happening ?</p>
|
<p>DNA Hairpins are formed when two regions in same single stranded DNA are complementary in nucleotide sequence but in the opposite directions (as represented in image below). These two sets of nucleotide sequences base-pair with each other by forming hydrogen bonds between adenine-thymine and guanine-cytosine respectively to form hairpin loop. Same structures can be seen in case of RNA.</p>
<p><a href="https://i.sstatic.net/Mjvug.jpg" rel="nofollow noreferrer"><img src="https://i.sstatic.net/Mjvug.jpg" alt="enter image description here"></a></p>
<p>(Figure via: <a href="https://brilliant.org/problems/dna-zipper/" rel="nofollow noreferrer">https://brilliant.org/problems/dna-zipper/</a>)</p>
<p>Single stranded binding (SSB) proteins bind to single DNA nucleotide sequences and prevent the breakdown of newly synthesized DNA because of nucleases and it also removes the secondary structure of the DNA strands like hairpin loops so that other enzymes can bind to DNA strand and operate properly. As represented in figure below SSB bind to ssDNA through electrostatic interactions and prevent the bond formation within the nucleotides of single DNA strand, thus preventing formation of hairpin loops in DNA.</p>
<p><a href="https://i.sstatic.net/5eVaA.gif" rel="nofollow noreferrer"><img src="https://i.sstatic.net/5eVaA.gif" alt="enter image description here"></a></p>
<p>(Figure via: <a href="http://helicase.pbworks.com/w/page/17605582/Amanda-Kinney" rel="nofollow noreferrer">http://helicase.pbworks.com/w/page/17605582/Amanda-Kinney</a>)</p>
<p>(via: <a href="https://proteopedia.org/wiki/index.php/Single_stranded_binding_protein" rel="nofollow noreferrer">https://proteopedia.org/wiki/index.php/Single_stranded_binding_protein</a>)</p>
| 259
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DNA replication
|
Correlation of Meselson and Stahl with “multifork” replication in E.coli
|
https://biology.stackexchange.com/questions/114321/correlation-of-meselson-and-stahl-with-multifork-replication-in-e-coli
|
<p>Because of the limiting value of the rate of DNA replication, rapidly dividing <em>E.coli</em> use multiple replication forks [1][2]. Thus, DNA replication of one generation has already begun in the previous generation.</p>
<p>To me, this poses a problem in understanding the results of the Meselson and Stahl experiment[3]:</p>
<p><a href="https://i.sstatic.net/D7L33.png" rel="nofollow noreferrer"><img src="https://i.sstatic.net/D7L33.png" alt="Text book representation of Meselson and Stahl Experiment" /></a></p>
<p><strong>Source</strong> Biology, Textbook for Class XII National Council of Educational Research and Training (<a href="https://ncert.nic.in" rel="nofollow noreferrer">https://ncert.nic.in</a>), NCERT Publications (version: November 2021 Agrahayana 1943) <a href="https://ncert.nic.in/textbook/pdf/lebo105.pdf" rel="nofollow noreferrer">https://ncert.nic.in/textbook/pdf/lebo105.pdf</a></p>
<p>Shouldn’t the DNA from generation I have somewhat more of <sup>15</sup>N DNA, as its replication started when the <em>E.coli</em> was still in the <sup>15</sup>N medium? Is that the case or is there another mechanism?</p>
<ol>
<li><p><a href="https://doi.org/10.1016/0022-2836(68)90425-7" rel="nofollow noreferrer"><em>J. Mol. Biol.</em> (1968) <strong>31</strong>, 519–540</a></p>
</li>
<li><p><a href="https://doi.org/10.1038/sj.emboj.7601871" rel="nofollow noreferrer"><em>The EMBO Journal</em> (2007) <strong>26</strong>, 4514–4522</a></p>
</li>
<li><p><a href="https://doi.org/10.1073/pnas.44.7.671" rel="nofollow noreferrer"><em>Proc. Nat. Acad. Sci.</em> (1958) <strong>44</strong>, 671–682</a></p>
</li>
</ol>
|
<p>It is sometimes considered that <a href="https://www.ncbi.nlm.nih.gov/pmc/articles/PMC1307605/" rel="nofollow noreferrer">the result was an artifact for a reason similar to that which you mention</a>. However, it happened to be correct in its conceptual outcome, and was not rejected by subsequent better-controlled experimental tests. This is pretty standard for experimental science. The result would not have been so widely believed if it were not that everyone else's data agreed.</p>
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DNA replication
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How is each region of the DNA replicated only once?
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https://biology.stackexchange.com/questions/73365/how-is-each-region-of-the-dna-replicated-only-once
|
<p>In "Molecular Biology of THE CELL" 3rd Edition, 1994, by Alberts, Et al. (<em>Yes I know there is a newer edition</em>)</p>
<p>the question is posed on page 362</p>
<blockquote>
<p>How is each region of the DNA replicated only once?</p>
</blockquote>
<p>Two suggestions are offered:</p>
<ol>
<li>Inhibitor-addition model</li>
<li>Initiation-removal model</li>
</ol>
<p>Since the 3rd edition of the book is now 24 years old, has the answer been discovered and if so what is it?</p>
<p>I am reading the 3rd edition now because that is what I have and plan to buy the newer 6th edition if a 7th edition will not be out in the next few months. This is for self-study so no need for me to rush or have the latest and greatest info for the first pass of learning as I am only reading.</p>
|
<p>DNA Replication ensures it only occurs once by the use of licensing factors. These factors are released and bind to the <a href="https://en.wikipedia.org/wiki/Origin_of_replication?wprov=sfti1" rel="nofollow noreferrer">origins of replication</a> during one distinct phase. After the factors have been released and the first phase has ended, the actual replication initiates. </p>
<p>Information on the licensing factors:</p>
<p>The licensing factors used to create the pre-replication complex is the <a href="https://en.wikipedia.org/wiki/Minichromosome_maintenance?wprov=sfti1" rel="nofollow noreferrer">Minichromosome Maintenance (Mcm2-7)</a> protein, Cdc6, and Cdt1. The Cdc6 and Cdt1 are required to load the Mcm’s onto the DNA and therefore, are highly controlled during the cell cycle. The Mcm2-7 hexamer is the actual DNA helicase that is used during DNA replication. </p>
<p>The way that the re-replication of DNA is prevented is that when the DNA replication starts, the Mcm helicase moves away from the ORC and the newly replicated DNA, meaning it then cannot be reinitiated. Along with the physical moving of the helicase, Cdt1’s activity is suppressed by the protein <a href="https://en.wikipedia.org/wiki/Geminin?wprov=sfti1" rel="nofollow noreferrer">Geminin</a>, which prevents the licensing of the helicase during times of non-replication. </p>
<p>Image from the article on the licensing.
<a href="https://i.sstatic.net/N6mm1.jpg" rel="nofollow noreferrer"><img src="https://i.sstatic.net/N6mm1.jpg" alt="Image on licensing"></a></p>
<p>More information on this topic:
<a href="https://www.ncbi.nlm.nih.gov/pmc/articles/PMC2688777/#!po=21.8085" rel="nofollow noreferrer">https://www.ncbi.nlm.nih.gov/pmc/articles/PMC2688777/#!po=21.8085</a></p>
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DNA replication
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Why are RNA nucleotides added first, only to be replaced by DNA nucleotides during DNA replication?
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https://biology.stackexchange.com/questions/108339/why-are-rna-nucleotides-added-first-only-to-be-replaced-by-dna-nucleotides-duri
|
<p>I am not asking why RNA primer needs to be added, but rather why RNA nucleotides are added as a primer by Primase and replaced by DNA polymerase I? It seems very inefficient to first insert RNA nucleotides and then add DNA nucleotides when you could have evolved to just add DNA nucleotides directly, without having to add RNA nucleotides.</p>
<p>Perhaps, it is an evolutionary remnant? Proof that DNA arised RNA? What do you guys think?</p>
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DNA replication
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Nomenclature of substrates for DNA synthesis
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https://biology.stackexchange.com/questions/98906/nomenclature-of-substrates-for-dna-synthesis
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<p>I have read in my school textbooks that both deoxyribonucleoside triphosphate and deoxynucleotide triphosphate are used in DNA Replication as substrates.</p>
<p>However, it is unclear to me whether the terms refer to the same molecule as one uses the term <em>nucleotide</em> and the other uses <em>nucleoside</em>. What confuses me is that nucleoside is a compound with a purine or pyrimidine base and a sugar and a nucleotide is the same but with the inclusion of a phosphate group.</p>
<p>Is it possible that the “deoxynucleotide triphosphate“ is used in artificial DNA replication as in PCR, and “deoxyribonucleoside triphosphate“ is the natural substrate for the replication of DNA within cells?</p>
|
<p><strong>Summary</strong></p>
<p>The systematic chemical names of many important biological molecules are too long to write routinely in full, and in any case were preceded historically by abbreviated forms which generally stressed the distinguishing features of importance to those studying their metabolism. Thus, as the poster points out, a molecule such as dATP (image below) has a base-(deoxy)sugar-phosphate(s) structure, and can be termed a <em>deoxynucleotide</em>. Adding <em>triphosphate</em> provides the additional information that there is more than one phosphate. A base-sugar(deoxyribose) combination is, as the poster states, a <em>deoxynucleoside</em>, but by extending this name to <em>deoxyribonucleoside triphosphate</em>, one indicates the addition of phosphates, so that one is, by definition, describing a <em>deoxynucleotide</em>. The reference to ribonucleoside indicates that the sugar is ribose, a specification lacking from the other definition. However, in practice these two terms are used to describe the same entity, one which might a little more completely describe as 2′-deoxyribonucleoside triphosphate (to indicate the position of the reductive change). This answer attempts to explain with precision the aspects of the nomenclature that are likely to be important to the biochemist or molecular biologist.</p>
<p><strong>Disclaimer</strong></p>
<p>The original question might be considered not to represent a biological problem, and one might argue for closing it on that account. However because of the very nature of trivial nomenclature, it is difficult for those without experience of the usage in a field to find the answer. In answering it I have tried to broaden the scope to include the aspects of structure that I feel are most likely to be to biological students.</p>
<p><strong>Answer</strong></p>
<p>It is convenient to start with ribonucleoside triphosphates (rather a deoxy- ribonucleoside triphosphates), and I have chosen a specific example in the molecule that is generally known as ATP:</p>
<p><a href="https://i.sstatic.net/zPo1p.png" rel="nofollow noreferrer"><img src="https://i.sstatic.net/zPo1p.png" alt="ATP structure" /></a></p>
<p>Alternative names for this ribonucleoside triphosphate, in decreasing order of complexity, include:</p>
<ol>
<li>[[(2R,3S,4R,5R)-5-(6-aminopurin-9-yl)-3,4-dihydroxyoxolan-2-yl]methoxy-hydroxyphosphoryl] phosphono hydrogen phosphate</li>
<li>9-β-D-ribofuranosyl adenine 5′-triphosphate</li>
<li>adenosine 5′-triphosphate</li>
<li>adenosine triphosphate</li>
<li>ATP</li>
</ol>
<p>Of interest to us are the three components of the molecule: a purine base (adenine), a sugar (ribose), and three linked phosphates. This is not reflected in the chemical nomenclature (1), so let us focus on the, albeit rarely used, biochemical nomenclature (2).</p>
<ul>
<li>The <em>‘9’</em> specifies the position of the adenine ring that is linked to the sugar. This is not of particular interest in the context of the question. (The numbering of the ring is, however, relevant to describing differences between or modifications of the purine and pyrimidine bases<sup>1</sup>.)</li>
<li>The <em>D-ribofuranosyl</em> is a form of ribose (<em>see</em> below). The ‘D’ indicates that it is D-ribose enantiomer rather than the alternative, L-ribose. However, as all natural sugars are ‘D’ this is generally omitted (although discussions of <em>why</em> all natural sugars are ‘D’ abound<sup>2</sup>).</li>
<li><em>Ribofuranosyl</em> indicates that the ribose is in the five-membered hemiacetal ring form (rather than the straight chain or the six-membered pyranose ring), but this is always the case in nucleosides, and so is not of particular interest.</li>
<li>The <em>5′</em> specifies the position of the sugar ring to which the phosphates are attached, the prime (′) indicating that the numbering is for the sugar ring (not that of the base). This is important, as it is relevant to the linkage in the (asymmetric) phosphodiester bond made to the 3′-OH:</li>
</ul>
<p><a href="https://i.sstatic.net/3fgSJ.png" rel="nofollow noreferrer"><img src="https://i.sstatic.net/3fgSJ.png" alt="Phosphodiester bond in DNA" /></a></p>
<p>The image above is for DNA, but before addressing the nomenclature there, we would point out:</p>
<ul>
<li>The common α, β, γ designation of the three bond phospho-ester bonds. This is very relevant to the formation of polynucleotides (DNA and RNA) from their substrates, and in general to the utilization of the free energy the hydrolysis of these bonds for other processes<sup>3</sup>.</li>
</ul>
<p>Finally we come to a structure of interest to the poster, dATP:</p>
<p><a href="https://i.sstatic.net/eIECU.png" rel="nofollow noreferrer"><img src="https://i.sstatic.net/eIECU.png" alt="dATP structure" /></a></p>
<ul>
<li>In the designation, 2′-deoxyadenosine 5′-triphosphate, the <em>2′-deoxy</em> indicates that it is the 2 position in the ribose ring (′) that has had the oxygen removed (deoxy), i.e. OH is reduced to H. Although common knowledge, it is important that the student new to the subject understands this, as if it were the 3′ position that was reduced, it could not form the phosphodiester bonds of DNA. And it is relevant in many other contexts. For example the chain-termination method of DNA sequencing uses 2′,3′-dideoxy nucleotides to inhibit such bonds forming<sup>4</sup>, the single difference between ribose and deoxyribose allows us to understand the profound affect on the susceptibility of RNA to alkaline hydrolysis in nature<sup>5</sup>, and certain RNA molecules are hydroxy-methylated at this position.</li>
</ul>
<p><strong>Related information in SE Biology answers</strong></p>
<p><sup>1</sup> <a href="https://biology.stackexchange.com/questions/97619/does-n1-methyl-pseudouridine-occur-naturally-in-any-rna/97645#97645">Topical example related to base modification</a><br>
<sup>2</sup> <a href="https://www.ncbi.nlm.nih.gov/pmc/articles/PMC2857173/" rel="nofollow noreferrer">Speculation on the origin of biological homochirality — not SE Biology</a><br>
<sup>3</sup> <a href="https://biology.stackexchange.com/questions/95290/what-determines-whether-a-reaction-using-atp-produces-adp-or-amp/95293#95293">Why does ATP sometimes produce ADP and sometimes AMP?</a> <br>
<sup>4</sup> <a href="https://biology.stackexchange.com/questions/87678/understanding-the-strategy-of-sanger-dna-sequencing/87922#87922">Sanger sequencing by chain termination</a><br>
<sup>5</sup> <a href="https://biology.stackexchange.com/questions/24786/why-deoxyribose-for-dna-and-ribose-for-rna/45690#45690">Difference in stability of DNA and RNA</a></p>
| 263
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DNA replication
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Regulation of the replication of mtDNA at embryonic level
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https://biology.stackexchange.com/questions/13655/regulation-of-the-replication-of-mtdna-at-embryonic-level
|
<p>While reading an article on mitochondrial inheritance I came across <a href="http://dx.doi.org/10.1093/humupd/dmq002" rel="nofollow">this link</a>.</p>
<p>The results state that mitochondrial DNA replication is regulated in different cells of an embryo at different levels. How is this regulated?</p>
|
<p>This is a topic which seems to be not very clear yet. References say that this is obviously dependent on the tissue and the "gene dosage" seems to play a role, and that this is probably regulated by a yet unknown factor (see "<a href="http://www.ncbi.nlm.nih.gov/pubmed/12880201" rel="nofollow">Mitochondrial DNA replication: what we know.</a>" and "<a href="http://www.ncbi.nlm.nih.gov/pubmed/12525854" rel="nofollow">Replication and transcription of mammalian mitochondrial DNA.</a>").
Newer articles suggest that it depends also on the transcription of the mtDNA (see "<a href="http://www.ncbi.nlm.nih.gov/pubmed/22207204" rel="nofollow">The interface of transcription and DNA replication in the mitochondria.</a>" and "<a href="http://www.ncbi.nlm.nih.gov/pubmed/22465614" rel="nofollow">Mitochondrial transcription factor A regulates mitochondrial transcription initiation, DNA packaging, and genome copy number.</a>"). I will see, if there are other interesting publications, if there are problems with accessing these articles, let me know.</p>
| 264
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DNA replication
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How did genome replication first synchronise with cell division?
|
https://biology.stackexchange.com/questions/74402/how-did-genome-replication-first-synchronise-with-cell-division
|
<p>It is obvious that cell division in living organisms is now synchronised almost perfectly with DNA replication and, furthermore, the line of division has to intersect exactly the space between the two copies of all the DNA molecules in the cell. I know that as otherwise, most cells would be either empty or have a lot of excess DNA strands, which would lead to an eventual termination of replication due to scarcity of resources and energy required to trigger it.</p>
<p>Some theories (<a href="https://www.ncbi.nlm.nih.gov/books/NBK26876/" rel="nofollow noreferrer">https://www.ncbi.nlm.nih.gov/books/NBK26876/</a>) suggest the first protocell might have formed by a liposome simply gobbling up a self-replicating molecule. However, how would the molecule then 'know':</p>
<ol>
<li>when exactly it should replicate so that the replication coincides with the liposome diving and</li>
<li>where in the molecule it should be located at the time of division, so that its copy ends up in a different daughter cell to itself</li>
</ol>
<p>?</p>
| 265
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DNA replication
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Why isn't the DNA in bacteria always split up and replicating?
|
https://biology.stackexchange.com/questions/41241/why-isnt-the-dna-in-bacteria-always-split-up-and-replicating
|
<p>Isn't helicase always free floating in bacterial cells, and the DNA without a nuclear membrane and uncoiled and freefloating and so why doesn't the helicase keep breaking the double helix of DNA? Also, since other DNA synthesizing enzymes like polymerase and RNA primer and other replication enzymes also free floating, so why doesn't the DNA in bacterial cells keep replicating at all times?</p>
|
<p>Bacterial DNA replication is initiated at the oriC by DnaA in <em>E. coli</em>. Think about ways in which DnaA binding or activity can be regulated in a way that inhibits or permits DNA replication. In recently replicated bacterial DNA, the DNA is hemimethylated (parental strand has a methyl group, daughter strand doesn't): An inhibiting protein binds to hemimethylated DNA, that can block DnaA activity at the oriC. A DAM methylase adds methyl groups after a while to the newly synthesized DNA, in which case DnaA may bind. Due to the lag between replication and methylation, DNA replication can't be continuously initiated! This is one of many examples (for example, DnaA must bind ATP for it's mechanism of action), but the take home point is DNA replication must be <strong>initiated</strong> and if it can't be, then it doesn't take place!</p>
<p><a href="https://www-als.lbl.gov/index.php/research-areas/bioscience/223-the-initiation-of-bacterial-dna-replication.html" rel="nofollow">THE INITIATION OF BACTERIAL DNA REPLICATION</a></p>
<p><strong>Breaking that down:</strong> (note, this is in <em>E. coli</em>)</p>
<hr>
<p>DnaA is a protein with DNA-binding activity. When many DnaA bind at the oriC, it causes the DNA to bend open and loop. At this point, DnaB would have access to the DNA, and this is the helicase that unwinds everything for replication (you'll also see that DnaB must be loaded onto the DNA by DnaC, a helicase loader). So blunty enough, oriC is an origin of replication, everything starts here. There are repeats of AT rich regions denoted 13mer and 9mer in the oriC (number of bases in the region if you will). We know that this is beneficial because AT rich regions are easiest to pull apart due to the number of hydrogen bonds between A and T.</p>
<p>In the example I provided, the assumption is that the DNA of <em>E. coli</em> is methylated on adenines on both strands. When replication occurs, the new DNA doesn't have this methyl group on it's adenines like the parental strand. A protein that recognizes and binds so-called hemimethylated (half methylated) DNA, SeqA, binds where it can, including in the oriC. It blocks DnaA subsequently for a short time, and falls off the DNA on its own. The DAM methylase adds the methyl groups when it can, and once it does the SeqA can no longer bind, but the replication-initiation complex thus can. </p>
<p>There are <strong>other</strong> modulators of DNA replication, and the concentration of the DnaA itself is a big factor because you need many units of DnaA to bind to facilitate initiation. So then you have to consider is DnaA being expressed strongly enough? And this has it's own pathway outside the scope of this question.</p>
| 266
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DNA replication
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How do multiple replication forks function without 'colliding', and what is the benefit of this method?
|
https://biology.stackexchange.com/questions/5613/how-do-multiple-replication-forks-function-without-colliding-and-what-is-the
|
<p>I'm currently reading a little about DNA replication, and have come accross the following statement;</p>
<blockquote>
<p>Replication starts from a fixed point and is bi-directional ... In Eukaryotes, there are multiple replication forks, each progressing in a bi-directional fashion.</p>
</blockquote>
<p>If there is a single, long strand of DNA in a Eukaryotic cell, I see potential problems with this:</p>
<p>These forks involve opening up a section of double-stranded DNA, and each strand becoming a double strand in a newly synthesised piece of DNA. At some point, before any single fork has become two new double-stranded molecules, another fork could 'collide' with this, causing it to attempt to replicate the non-finished section. </p>
<p>Simply, how can one replication fork meet another without either exponentially increasing the number of strands being replicated?</p>
<p>Also, on a more general level, I would be quite interested to know the actual benefit of this, when, typically, only a single copy of the double-stranded molecule needs making.</p>
|
<p>To get to the exponentially increasing number of strands, the replication would have to be started on the already replicated strand, and not on the original strand. Replication in eukaryotes is tighly controlled and such an event is prevented by the regulation.</p>
<p>How can the cell prevent re-replication of the strand that is currently synthesized? By timing the steps of the replication in strict order and preventing any replication outside of that order. </p>
<p>Replication starts at defined origins of replication. In the G1 phase, pre replication complexes (pre-RCs) are assembled at the origins of replications. At these pre-RCs the replication can be initiated, but pre-RCs itself cannot be assembled in the later phases of the cell cycle. When the replication starts, the pre-RC is converted to a post-RC which can't initiate the replication anymore, preventing re-replication.</p>
<p>The benefit of multiple origins of replication is replication speed. Eukaryotic genomes can be much, much larger than those of simple bacteria. Replicating them using a single origin of replication would take a very long time.</p>
<p>Some further reading material:</p>
<ul>
<li><a href="http://www.ncbi.nlm.nih.gov/books/NBK21113/" rel="nofollow">Genomes, 2nd edition, chapter 13</a></li>
<li><a href="http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2292467/" rel="nofollow">Cell Cycle Regulation of DNA Replication</a></li>
</ul>
| 267
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DNA replication
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When using a primer to replicate DNA in a plasmid, does it replicate the whole plasmid?
|
https://biology.stackexchange.com/questions/72000/when-using-a-primer-to-replicate-dna-in-a-plasmid-does-it-replicate-the-whole-p
|
<p>I've been learning about PCR and plasmids. I understand that the reason for having both a forward and a reverse primer is to extract and amplify the specific piece of DNA between these two primers. </p>
<p>What I'm struggling to understand is that why after the first replication, a fully circular DNA strand isn't formed and hence, why isn't a fully circular DNA strand formed for each replication. </p>
<p>For the following diagram , let the strands be labelled strand 1, strand 2, strand 3, strand 4 from top to bottom. (Diagram from Lehninger Principle of Biochemistry, 5th Ed). </p>
<p><a href="https://i.sstatic.net/CkfaJ.png" rel="nofollow noreferrer"><img src="https://i.sstatic.net/CkfaJ.png" alt="2nd cycle of PCR"></a></p>
<p>For example, in the diagram here. When cloning a plasmid I assume strand 1 is fully circular, so strand 2 (which was transcribed from strand 1) should also be fully circular and in the diagrams it should show DNA to the left of the site of interest. </p>
<p>Does the transcription stop somewhere along the line, preventing it from copying the whole strand? Or does the strand never join up into a circle, so that the DNA that would appear to the left of strand 2 has been copied, but doesn't join up to the site of interest so wasn't included in the diagram. </p>
<p>Thanks for your help.</p>
|
<p>PCR produces linear DNA, not circular. You can replicate whole plasmids using PCR, but the DNA will still be linear, as if you cut the plasmid with a blunt-end restriction enzyme.</p>
<p>So why doesn't the whole plasmid get copied when using PCR? Let's assume the primers are designed to bind 1000 bases apart from each other. The first round of PCR will probably produce longer pieces of DNA, because the oligos can only bind to the plasmid and the Polymerase can keep going. But the polymerase will fall off at some point, either randomly or when the temperature is pushed up for the next round of PCR. But in round 2, there should be equal numbers of circular plasmid and linear copy. The proportion of linear copy will increase every round. As the linear copies increase, the oligos are more and more likely to bind to linear copies than to circular plasmid.</p>
<p>Binding to linear copies is why the PCR product size is controlled. Let's assume that a linear copy is 1500 bases, because the polymerase ran an extra 500 bases past the other oligo site on the plasmid. But when the reverse oligo binds that site on the linear copy, the polymerase will run 1000 bases and fall off the end of the linear copy. This new copy will have the forward and reverse oligo sites at the ends, so it's the right size. Any copies produced from this strand will also be the right size. As the reaction proceeds, the proportion of correct-sized strands increases, and create more correct-sized strands.</p>
| 268
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DNA replication
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Topology of closed circular DNA
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https://biology.stackexchange.com/questions/16992/topology-of-closed-circular-dna
|
<blockquote>
<p>Why are covalently closed circular plasmid DNAs naturally found in an underwound state ?</p>
</blockquote>
<p>Is it because this makes it easier for the DNA replication machinery to access and unwind DNA ? Or is it because underwound state is energetically more favourable than an overwound state ?</p>
|
<p>DNA is not always negatively supercoiled naturally. It is important to keep in mind that different regions of topologically constrained DNA can have different supercoiling values. For example, the action of unwinding DNA for transcription or replication introduces positive supercoils ahead of the polymerase and negative supercoils behind it.</p>
<p>Additionally, supercoiling is present in DNA normally. Supercoiling is the result of more or less twist in a DNA helix, which is most stable when it has ~10.5 bp/turn (for B-DNA). Supercoiling allows under- or overwound DNA to return to its most stable twist. Negative supercoiling is the result of underwound DNA. Underwound DNA is not thremodynamically favourable and will actually lower the melting point of DNA (the point where strand separation occurs). This can be compensated by introducing supercoils but it is also important for processes such transcription, which require ssDNA. Negative supercoils and ssDNA can be thought of as interchangeable. Negative supercoils are also used to package DNA around histones in eukaryotes and archaea. On the other hand, positive supercoiling is the result of overwound DNA (also not thermodynamically favourable) and will increase the melting point of DNA. Positive supercoiling is often found in thermophiles which live at higher temperatures and need to prevent their DNA from melting excessively. It has also been suggested that positive supercoiling can play a role in regulation of gene expression (by inhibiting promoter melting). Organisms like to keep an approximately constant amount of supercoiling in their DNA for the above reasons; this is called the superhelical density and is characteristic of different classes of organisms.</p>
| 269
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DNA replication
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why dna polymerase 3 requires a primer for replication
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https://biology.stackexchange.com/questions/40953/why-dna-polymerase-3-requires-a-primer-for-replication
|
<p>Why DNA polymerase 3 needs a primer to star replication.And whats happens when there is no AUG sequence on entire DNA.</p>
|
<p>You are confused among DNA replication, DNA transcrption, and RNA translation.</p>
<p>First, DNA replication happens during cell division, it create two exactly same daughter DNA.</p>
<p>Second, DNA transcription transcribes DNA sequence into RNA sequence, this RNA sequence may be used to synthesize protein, or RNA itself as signal, etc.</p>
<p>Third, RNA translation is the RNA sequence (codon) recognized by tRNA, and then synthesize protein sequence.</p>
<p>AUG is a codon in the RNA, the is recognized by tRNA, and then start translation. And please remember, DNA use A, G, C, T, but RNA use A, G, C, U, there is no <strong>U</strong> in DNA. </p>
<p>For DNA synthesis, why does it need primer? I have another answer for solving your question: <a href="https://biology.stackexchange.com/a/40954/17473">https://biology.stackexchange.com/a/40954/17473</a></p>
| 270
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DNA replication
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In DNA repair, how is it determined which strand contains the error?
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https://biology.stackexchange.com/questions/9120/in-dna-repair-how-is-it-determined-which-strand-contains-the-error
|
<p>DNA replication is more accurate than transcription (or RNA replication) because mechanisms exist for proof-reading and repair of DNA, but not for RNA.
Consider a segment of a DNA strand, AGTC. Its complement is GACT. Now suppose its complement is mutated to TACT — the DNA repair system will replace the wrong T by G. Why isn’t A in the original strand replaced by C? How does the system determine that the first strand is correct and its complement is incorrect? </p>
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<p>The reason the fact it isn't realistic is important. DNA repair machinery works by repairing common errors that occur due to common mutational pathways. The enzymes are specific for this, for example one particular enzyme targets mutations caused by UV and itself is activated by sunlight thus it's specificity makes it repair the correct chain. Also during replication, newly synthesised DNA lacks methylation. The repair enzymes thus know which chain is correct. </p>
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DNA replication
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Why is a solution of cesium chloride used in Meselson & Stahl's DNA replication experiment?
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https://biology.stackexchange.com/questions/96555/why-is-a-solution-of-cesium-chloride-used-in-meselson-stahls-dna-replication
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<p>Centrifugation involves separating particles of different sizes, masses, density and etc.</p>
<p>In the experiment, the DNA macromolecules are suspended in a solution of cesium chloride gradient and then centrifuged.</p>
<p>Why is cesium chloride needed? Considering the possible DNA produced in the experiment will be the same size but of different mass resulting in different density, DNA will be separated based on their density by virtue (without the need for cesium chloride) under the influence of a centrifugal force.</p>
<p>I noticed someone asked the same question on <a href="https://www.khanacademy.org/science/biology/dna-as-the-genetic-material/dna-replication/a/mode-of-dna-replication-meselson-stahl-experiment" rel="noreferrer">Khanacademy</a> but there was no answer for it. Other websites I found doesn't seem to offer a valid answer e.g. <a href="http://www.phschool.com/science/biology_place/biocoach/dnarep/cscl.html#:%7E:text=CsCl%20(cesium%20chloride)%20centrifugation%20is,separating%20DNA%20based%20on%20density.&text=As%20the%20CsCl%20gradient%20forms,a%20single%20band%20of%20DNA." rel="noreferrer">here</a> which says " the DNA comes to equilibrium in the gradient where its density equals the density of the surrounding CsCl" which I think doesn't really matter when considering the argument I proposed above in the third paragraph.</p>
<p>Here is a diagram of the experiment for reference. Thanks in advance.
<a href="https://i.sstatic.net/vSsKb.jpg" rel="noreferrer"><img src="https://i.sstatic.net/vSsKb.jpg" alt="enter image description here" /></a></p>
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<p>In saline solution all the DNA ends up at the very bottom.</p>
<p>Under centrifugation, the Caesium Chloride solution forms a density gradient, each DNA rises or sinks to the equivalent density.</p>
<p>The same procedure is used with Sucrose solutions for other separations.</p>
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DNA replication
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What regulates the timing of the motion of molecular machines during DNA Replication?
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https://biology.stackexchange.com/questions/20448/what-regulates-the-timing-of-the-motion-of-molecular-machines-during-dna-replica
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<p>This question is about <a href="https://www.youtube.com/watch?v=bee6PWUgPo8">this video</a> I found on Youtube. I just want to know what is the mechanism which regulates the timing of motion of the parts of these molecular machines.</p>
<p>I know that those big molecules moves using mechanical energy from ATP molecules and take advantage of the electric forces specially the hydrogen bonds, they also take advantage of the Brownian motion of water molecules around them.</p>
<p>From what is see, the green floating molecule triggers the whole process by attaching the purple one, then some electric equilibrium is disturbed so the {purple+green} molecule is attracted by the horizontal cyan molecule...etc.</p>
<p>But the problem is: does the green molecule attaches to this group in a periodic timing ? or in other words does the <a href="http://en.wikipedia.org/wiki/Okazaki_fragments">Okazaki fragments</a> have the same lenght evertytime ?</p>
<p>Thank you in advance !</p>
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<p>That's a pretty neat video, I'll just give you some background information first. It's an illustration of the "trombone model" of DNA replication. The darker blue molecule is helicase, it unwinds the DNA and facilitates translocation (this is an ATP dependent process). The dark purple molecules are DNA polymerase, they catalyze DNA strand synthesis (an NTP dependent process) using an ssDNA template and 3'-OH primer (the primer:template junction). The green circular molecules are sliding clamps, they increase the processivity of DNA polymerase. The light purple molecule is the sliding clamp loader, unsurprisingly it loads the sliding clamp onto the DNA (ATP dependent). The light blue molecule is a flexible linker that connects everything together. The other green molecule (the one that contacts helicase) is primase, it synthesizes RNA primers (which contains the 3'-OH initially used by DNA polymerase). </p>
<p>Certainly intermolecular forces play a significant role in this (and every) biochemical process, but I'm not really sure what you're talking about when you're describing electric forces and Brownian motion.</p>
<hr>
<p>Anyways, the answer to your question is no, Okazaki fragments do not have a fixed length. DNA synthesis, as mentioned above, occurs only by adding NTPs to a 3'-OH primer. Thus the length of Okazaki fragments is dependent on the spacing between primers on the lagging strand. This is dependent on <strong>when</strong> primase binds which in turn is dependent on primase concentration as well as its binding affinity with DNA and other replication fork proteins (especially helicase). High concentration and/or high affinity leads to a higher binding frequency and therefore shorter Okazaki fragments. The opposite is true for low concentration and/or low affinity. Note that most (or all?) primases have some degree of sequence specificity which means that specific DNA motifs will increase binding affinity. For example, <em>Escherichia coli</em> primase (DnaG) recognizes the trimer GTA.</p>
<p>This process also requires some degree of coordination between leading and lagging strand synthesis. Primer synthesis and DNA polymerase recycling on the lagging strand is much slower than the continuous synthesis on the leading strand. It has been suggested that primase acts as a "molecular brake" by halting the replication fork during primer synthesis to prevent the leading strand polymerase from rapidly outpacing those on the lagging strand (Lee JB, Hite RK, Hamdan SM, Xie XS, Richardson CC, van Oijen AM. 2006. DNA primase actas as a molecular brake in DNA replication. Nature. 439(7076):621-624.).</p>
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DNA replication
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DNA sequencing problem
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https://biology.stackexchange.com/questions/30340/dna-sequencing-problem
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<p>First off, let me start by outlining the problem:</p>
<p>Your laboratory has established a technique for examining DNA replication in a cellular extract. To the cellular protein extract, you add nucleotides, a small amount of radiolabeled 32P-dGTP to aid visualization of the synthesized DNA, and a 4000-base-pair linear double-stranded DNA molecule that contains an origin of replication in the middle. After allowing the reaction to proceed for 30 minutes at 30°C, you boil the mixture to denature the proteins and the DNA strands, separate the components on an acrylamide gel, and detect the radiolabeled DNA products. This complete reaction is shown in lane 2 on the gel in the Figure below.</p>
<p><img src="https://i.sstatic.net/q0jJs.png" alt="enter image description here"></p>
<p><strong>Problem A:</strong></p>
<p><em>You perform the assay by yourself for the first time with no one else is in lab. The tube labeled “nucleotide mixture” has only enough for a single reaction, but you find a tube with a label that reads “dATP, dTTP, dGTP, dCTP” and you use this in a second reaction. You find the first reaction looks like lane 2 and the second reaction looks like lane 3. Why was no DNA synthesized in the second reaction?</em></p>
<p>My answer:</p>
<p>Because deoxyribose triphosphates do not have a –OH group to attach. You essentially can’t build with them.</p>
<p><strong>Problem B:</strong></p>
<p><em>Your lab partner has recently isolated a mutant strain that can replicate its DNA normally at 30°C but exhibit no DNA replication at 40°C. He calls the mutant tsr1, for Temperature Sensitive Replication. The wild-type strain replicates efficiently at both temperatures. You believe that you can use the biochemical assay of cell extracts to identify which genes are defective in the mutant. You grow wild-type cells and tsr1 cells at 40°C, make the extracts, incubate the DNA replication reaction at 40°C, and detect the products on a gel. You observe the pattern shown in lanes 4 and 5. You are so excited by the results that you run to your lab partner and tell him that you know what enzyme is defective. He is delighted by your results but says that there are three different enzymes that can account for your results. What are they?</em></p>
<p>I am 100% lost on this one.</p>
<p>I understand from the figure that the wild-type at 40 degrees has 4000 nucleotides so it's not being cut off. The mutant, trs1 is being cut off one into two slices of 2000 and one of these slices has been cut off into slices of 500? Am I understanding this correctly?
I also don't understand why the 500 line is so thick compared to the others?</p>
<p>Anyhow I feel really confused on this. I've been learning about Sanger sequencing and it makes 100% sense with ddXTP added into the mix to view the length of each sequence and therefore be able to read the sequence, but I'm not even sure if that's what's happening here.</p>
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<p>Even though you - or the problem did not clarify, I assume in my answer that you work with eukaryote system, even though the principle of the replication is the same.</p>
<p>dNTPs do have an OH group on their 3rd carbon atom (for eg. check this <a href="http://en.wikipedia.org/wiki/Deoxycytidine_triphosphate" rel="nofollow">wiki-page</a> for dCTP) so I don't think that's the issue here. My guess is that the first tube contains the radio labeled GTP and the second does not, so even the DNA is there, you cannot see it - even though the question specifically asked why DNA was not synthesized, my opinion is that it's a tricky question to confuse you.</p>
<p>For the second problem we need to dig into the replication process a little bit:</p>
<p>In the replication fork the two strands are synthesized differently. The so called leading strand is made continuously, without interruptions, however the other strand - the so called lagging strand is synthesized by ligating small fragments - aka Okazaki fragments. These small fragments require their own RNA primers that need to be removed after synthesis is done.
Now since the origin of replication is in the middle and replication can proceed both ways, on a linear fragment such as yours you can get two 2000bp fragment from the two leading strands. Would it be a circular DNA, then you could still get 4kbp fragments since the replication could go around the DNA circle.
The smaller 500bp fragments that are quite abundant in the tsr1 lane, are the fragments that cannot be ligated. That being said I must admit that 500bp Okazaki-fragments would by atypical for eukaryotes (usual is around 100-200bp). So the enzymes responsible for this could be:</p>
<p>DNA Ligase - the one responsible for ligating (joining ) the lagging strand's fragments.
Polymerase delta (Pol δ): That is responsible for lagging strand synthesis and primer removal - in this case the primer removal function might be defective, this causes that the primers stay on the DNA and complete synthesis and ligation cannot occur.
The last protein although it is not quite an enzyme is the DNA clamp complex (PCNA). This complex stabilizes the polymerase to DNA, but when reaching a previous fragment the polymerase should dissociate. But if the clamp is dysfunctional than the polymerase gets stuck on the end of the fragment thus no primer removal and ligation can occur.</p>
<p>If need more info read this <a href="http://en.wikipedia.org/wiki/DNA_replication" rel="nofollow">wiki-page</a>.</p>
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DNA replication
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"Prime" structure of DNA Double Helix: Confusion
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https://biology.stackexchange.com/questions/42489/prime-structure-of-dna-double-helix-confusion
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<p>In <a href="https://www.youtube.com/watch?v=H_l0rnvPcTA" rel="nofollow noreferrer">this</a> video on DNA replication, the diagram shows the unwound DNA as still being anti-parallel, but the first diagram in <a href="https://biology.stackexchange.com/questions/31585/does-dna-polymerase-always-go-the-same-direction">this</a> post on Biology SE shows that the individual strands are 5'____5' and 3'_____3'. So my question is the following:</p>
<p>Does a single, unwound strand of DNA run as either 5'______3' or 3'_______5', or does it run either 5'______5' or 3'________3'?</p>
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<p>DNA strands always have one 3' end and one 5' end (since each nucleotide has one of each and a strand is formed by connecting the 3' side of one nucleotide to the 5' side of another nucleotide). In a double helix DNA molecule, the two strands run in opposite directions.</p>
<p>The Pearson Education diagram in the post you referenced has one pair of the strand ends mislabelled. The lefthand-most labels 5' above the 3' should have been 3' above 5'. </p>
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DNA replication
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Does DNA ligase have any role in replication on leading strand?
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https://biology.stackexchange.com/questions/105304/does-dna-ligase-have-any-role-in-replication-on-leading-strand
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<p>According to my notes, one RNA primer is required on the leading strand to start DNAP activity, and at the end, a repairing enzyme will remove the primer and replace it with the complementary DNA nucleotides.</p>
<p>After this stage, will DNA ligase join the newly added nucleotides, or is ligase not required? I've heard that ligase is not used at all on the leading strand, but that doesn't seem quite right.</p>
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<p>DNA ligase is indeed needed on the leading strand. Just not nearly as much as on the lagging strand.</p>
<p>The work of DNA ligase is to form phosphodiester bonds (bonds joining the phosphate of one nucleotide with the OH of another nucleotide), sealing the gaps between two nucleotide sequences. On the lagging strand, it's used to fill the gaps between Okazaki fragments. However, since Okazaki fragments do not form on the leading strand, its replication largely does not use ligase.</p>
<p>There are a couple notable exceptions:</p>
<ul>
<li>After the leading strand's RNA primer is replaced with DNA, ligase must join the new sequence to the rest of the strand with ligase.</li>
<li>If a DNA strand has multiple origins of replication, the leading strands of separate replicons must be joined with ligase.</li>
</ul>
| 276
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DNA replication
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What is the structure and function of chromosomes during interphase?
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https://biology.stackexchange.com/questions/1057/what-is-the-structure-and-function-of-chromosomes-during-interphase
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<p>Ok, it seems to be easy but I have probably ignored something by accident.</p>
<p>Interphase is the phase where things are growing and the preparation for cell division happens. Its stages G1, S and G2. DNA replication in S stage. So the DNA in some chromosomes must have the pieces of information about how to do the DNA replication. - I am not sure about thing.</p>
<p>I would say to the part: structure that chromosomes are diploid at the given stage. - It does not feel right anyway.</p>
<p><strong>What would you answer to the main question?</strong></p>
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<blockquote>
<p>So the DNA in some chromosomes must have the pieces of information about how to do the DNA replication. - I am not sure about thing.</p>
</blockquote>
<p>Genomes contain what is called the "origin of replication" - specific sequences in the DNA that tell DNA polymerase where to bind and to initiate replication. </p>
<p>As for your main question, I'm a little confused as to what you're asking. In a general sense, chromosomes function as carriers of genetic information. In eukaryotes, nuclear DNA is organized in the nucleus on linear chromosomes which carry most of the genetic information an organism needs to survive. In bacteria, the chromosome is a circular piece of DNA. In eukaryotes, the chromosome is also bound by histone proteins, which serves to regulate expression of certain genes and to help anchor the chromosomes to the inner nuclear membrane.</p>
| 277
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DNA replication
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Hydrogen bonding and the blocking thereof in nucleic acids during nuclear processes
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https://biology.stackexchange.com/questions/43504/hydrogen-bonding-and-the-blocking-thereof-in-nucleic-acids-during-nuclear-proces
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<p>In transcription, RNA polymerase unwinds the DNA double helix and begins attaching RNA nucleotides to the template strand. In its wake, the DNA double helix closes back—this is only natural, seeing as the DNA bases have the tendency to hydrogen bond. In DNA replication, this closing is prevented by single-strand binding proteins.</p>
<p>I have three questions regarding this:</p>
<ol>
<li>In transcription, the pre-mRNA strand peels off the template strand in the wake of RNA polymerase. Why? There are just as well hydrogen bonds between the pre-mRNA strand and the DNA template strand as there are between the two DNA strands.</li>
<li>Why doesn’t the pre-mRNA stick onto the DNA indefinitely and block the non-template DNA strand from reattaching to the template DNA strand?</li>
<li>In DNA replication, why is it imperative that the separated DNA be stabilized by single-strand binding proteins? Won’t the replicated strand block the template strands from closing together?</li>
</ol>
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DNA replication
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Is the cell cycle applicable to meiosis as well, or just mitosis?
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https://biology.stackexchange.com/questions/96939/is-the-cell-cycle-applicable-to-meiosis-as-well-or-just-mitosis
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<p>All the diagrams I can find, show the cell cycle as having G1 phase (growth 1), S phase (DNA replication), G2 (growth 2) before the Mitotic phase (mitosis + cytokinesis).</p>
<p>Is there an equivalent "cell cycle" for meiosis, since the chromosomes in parent cell in meiosis also having "double" the genetic material prior to cell division (presumably from DNA replication too)?</p>
<p>Is it simply the same cell cycle as mitosis but with a <strong>Meiotic</strong> phase instead of Mitotic?</p>
<p>If so, would appreciate if anyone had a diagram :) Thanks!</p>
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<p>The cell cycle is only <a href="https://en.wikipedia.org/wiki/Cell_cycle#Mitotic_phase_(chromosome_separation)" rel="nofollow noreferrer">associated with mitosis</a>. The cell cycle is the normal process of cell division with which cells can indefinitely increase their number by cyclically repeating the process. When a cell goes through the cycle, the result is two cells that are genetically identical.</p>
<p>Meiosis is a special type of cell division (which can occur only in <a href="https://en.wikipedia.org/wiki/Eukaryote" rel="nofollow noreferrer">eukaryotes</a>) that produces cells that are not genetically identical to the initiating cell. The number of chromosomes in each of the resulting cells is half the number that were in the initial cell. (These <a href="https://en.wikipedia.org/w/index.php?title=Haploid" rel="nofollow noreferrer"><em>haploid</em> cells</a> can later participate in <a href="https://en.wikipedia.org/wiki/Fertilisation" rel="nofollow noreferrer">fertilization</a>, producing a cell with the original number of chromosomes.) Many of the steps of meiosis are similar to the steps involved in mitosis, but overall the process is more complex. Since meiosis reduces the number of chromosomes, it cannot be repeated and so does not take part in a cell division cycle.</p>
| 279
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DNA replication
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In DNA replication, are there phosphodiester bonds in the primer ? between the RNA nucleotides before being replaced
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https://biology.stackexchange.com/questions/89332/in-dna-replication-are-there-phosphodiester-bonds-in-the-primer-between-the-r
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<p>When hydrogen bonds happen between the RNA nucleotide bases and the DNA bases , do phosphodiester bonds form between the RNA nucleotides in the primer ?
No source I read is clear about this, are their bonds between the RNA nucleotides ? </p>
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<p>A <a href="https://www.nature.com/scitable/definition/primer-305/" rel="nofollow noreferrer">primer</a> is by definition a short single strand of nucleic acid (i.e. a series of nucleotides linked together by phosphodiester bonds). See also <a href="https://en.wikipedia.org/wiki/Primer_(molecular_biology)]" rel="nofollow noreferrer">this wikipedia article</a>.</p>
| 280
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DNA replication
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Telomerase and End Replication in Eukaryote
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https://biology.stackexchange.com/questions/112036/telomerase-and-end-replication-in-eukaryote
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<p>Here is a picture of using telomerase in solving end replication problem (Courtesy: Molecular Biology of the Cell, Alberts, Garland Science Pub.)</p>
<p><a href="https://i.sstatic.net/lxXMh.png" rel="nofollow noreferrer"><img src="https://i.sstatic.net/lxXMh.png" alt="enter image description here" /></a></p>
<p>Now lets consider the <span class="math-container">$3'$</span> end here, we are extending this end, and then using DNA Pol we extend the <span class="math-container">$5'$</span> end, even though the <span class="math-container">$5'$</span> end would still be truncated w.r.t <span class="math-container">$3'$</span> end but the original strand would be maintained. Now we know that this Telomerase enzyme is active in stem cells but NOT in somatic cells. So far so good.</p>
<p>Now if I consider the above layout in more detail: Lets consider what happens after the above replication cycle:</p>
<p><a href="https://i.sstatic.net/wfot3.jpg" rel="nofollow noreferrer"><img src="https://i.sstatic.net/wfot3.jpg" alt="enter image description here" /></a></p>
<p>This is because in the 1st replication cycle, we have added more nucleotides to <span class="math-container">$3'$</span> end, as well as possible few more nucleotides to <span class="math-container">$5'$</span> end as well. Now this would create overhangs on each <span class="math-container">$3'$</span> end of my DNA. And I am confused on these overhangs. Either the replication machinery deletes these overhangs ( which I am not sure whether it happens), OR the second replication would start on these overhang DNAs. A possible issue with the second approach (i.e replication with overhangs intact) is that the length of each strand would keep on increasing with each replication cycle as we are keep adding new nucleotides to <span class="math-container">$3'$</span> end in every replication cycle!!</p>
<p>Can someone shed some light on how the second cycle would happen, and <strong>whether the overhangs would get deleted or it would remain</strong> (in this case strand length would keep on increasing)?</p>
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<p>The overhangs are needed for telomere function and maintaining chromosome stability, so they aren't deleted. This is an active area of research, so the amount of detail you need to know depends on your level of study. They undergo processing instead of being deleted, involving nucleases. Note that DNA polymerase is filling in most of the gap. It will continue doing this in subsequent replications, so the slight single-stranded region left over won't grow. I'm assuming you mean "single-strand length" by "strand length." If you mean overall telomere size, then it depends on if the cell is somatic or not, which you pointed out.</p>
<p>(The 3′ overhangs can also form structures called T-loops, where the single-stranded overhang "invades" the double-stranded telomeric DNA. This helps protect the chromosome ends from being recognized as damaged DNA, maintaining genome stability.)</p>
| 281
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DNA replication
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PCR and Semiconservative replication
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https://biology.stackexchange.com/questions/80338/pcr-and-semiconservative-replication
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<p>Why does PCR use heat as opposed to helicase like in semi-conservative replication in order to separate the double DNA strand?</p>
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<p>Simply because it is ways more practical and there is no need to use a helicase. Heating is fast and convenient and denaturation is reversible. Also all the DNA is denaturated, so afterwards, primers can bind to their target sites, setting up the start point of the polymerase.</p>
<p>If you want to do this with and helicase, you need to make sure that you have a heat stable polymerase (at least at 72°C), you need to have the right places on the DNA unpackaged and finally, you need two enzymes in the reaction (which is more expensive than just having one).</p>
| 282
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DNA replication
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Is variation a result of Evolution?
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https://biology.stackexchange.com/questions/86581/is-variation-a-result-of-evolution
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<p>We know that the DNA copying mechanism that replicates DNA during cellular division is not 100% accurate and the resultant errors are the source of variation in the members of a population.</p>
<p>At the same time, we are also aware of the benefits of variation - how it is useful in ensuring the survival of a species over time <strong>and leading to evolution</strong>.</p>
<hr>
<p>I would however like to know if 100% accurate DNA replication is possible (even if hypothetically) - because in my opinion, organisms can surely survive (atleast individually) without variation or evolution.</p>
<p>If it is so, <em>is it possible that <strong>organisms with 100% accurate DNA replication</strong> did exist, but eventually their populations died out (due to natural selection) and we were left only with organisms that showed variation</em>?</p>
<hr>
<p>Putting in simpler terms, it is possible that <strong><em>evolution itself is the cause of variation that now causes further evolution?</em></strong></p>
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<p>You're asking if cells arose with 100% replication accuracy and if lower accuracy was selected for under a feedback loop. Maybe there was a sweet spot for DNA replication accuracy in terms of efficiency, but it's highly unlikely that our ancestor cells had 100% fidelity in DNA replication because if we look at yeast for example, there are many genes dedicated to different types of post-replication repair including mismatch repair, base excision repair, and translesion synthesis. All of these can lead to mutations if they aren't working properly and some (translesion synthesis) inherently generate mutations. It doesn't seem consistent for a cell to develop all of these sophisticated post-replication repair systems to protect itself from DNA damage <em>and then</em> to give some more leeway to DNA replication because it was just <em>too</em> accurate.</p>
<p>Also there are papers <a href="https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3319127/" rel="nofollow noreferrer">like this</a> that talk about how <em>E. coli</em> have increase mutation rates under stress. But if your hypothesis was true, then <em>E. coli</em> would ideally have 100% replication fidelity under normal conditions and lower it when stressed, or they would just have a higher baseline mutation frequency all the time. But, neither of these is observed. What makes the most sense when all this information is put together is that cells try to have as high DNA replication fidelity as possible under ideal conditions. </p>
<p>As per your other question, maybe it's possible to have a 100% accurate DNA polymerase <a href="https://international.neb.com/tools-and-resources/feature-articles/polymerase-fidelity-what-is-it-and-what-does-it-mean-for-your-pcr" rel="nofollow noreferrer">NEB engineers high fidelity polymerases for lab use</a> maybe someday it can reach 0 errors. But there are other sources of mutations like the spontaneous deamination of cytosine that can generate mutations during replication, but that depends on the chemical properties of cytosine so it's not really fixable unless you use different nucleobases. </p>
<p>Reference:</p>
<p>Mutation as a Stress Response and the Regulation of Evolvability. (2012)</p>
<p>By: Rodrigo S. Galhardo, P. J. Hastings, and Susan M. Rosenberg</p>
| 283
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DNA replication
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Parallel DNA double-helices with Watson–Crick base-pairing: Why do they not occur?
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https://biology.stackexchange.com/questions/70568/parallel-dna-double-helices-with-watson-crick-base-pairing-why-do-they-not-occu
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<p>I know that parallel DNA helices exist and are governed by Hoogsten base pairing, but why can’t they be possible with Watson-Crick pairing? In the diagram below, if we were to flip one of the strands while keeping the other the same, it appears as though hydrogen bonding is still possible.</p>
<p>The only specific suggestions that I could find was because of the DNA replication process and the negative polarity of hydoxyl group on the phosphates. Moreover, after flipping one strand, the DNA nucleotides form enantiomers. Are these possible reasons, or are there others?</p>
<p><a href="https://i.sstatic.net/dtHqy.jpg" rel="nofollow noreferrer"><img src="https://i.sstatic.net/dtHqyl.jpg" alt="Anti-parallel and parallel DNA base-pairing"></a></p>
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<blockquote>
<p>“The only specific suggestions that I could find was because of the DNA replication process and…”</p>
</blockquote>
<p>No. The explanation can have nothing to do with DNA replication. If the structure does not exist, you can’t replicate it, if it does, Nature will evolve a mechanism. (The related SE question, mentioned by @Gilleain, asked whether it could still replicate if it were parallel, i.e. using the enzymes that have evolved for parallel DNA.)</p>
<blockquote>
<p>I know that parallel DNA helices exist…</p>
</blockquote>
<p>Let us clarify this first. Perhaps the most extensive parallel duplex DNA, the structure of which has been determined, is that described by <a href="https://academic.oup.com/nar/article/30/7/1500/2376009" rel="nofollow noreferrer">Parvathy et al.</a>. The two parallel strands of this are shown below:</p>
<p><a href="https://i.sstatic.net/B6cV7.png" rel="nofollow noreferrer"><img src="https://i.sstatic.net/B6cV7.png" alt="Parallel DNA sequence"></a></p>
<p>The following points should be noted:</p>
<ol>
<li>This parallel DNA (and the shorter examples that preceded it) is not a pure stretch of complementary base pairs. </li>
<li>It is stabilized by what the authors refer to as “CC+ clamps” at either end. One is left to conclude that without these the duplex would not form.</li>
<li>All the complementary base-pairs are of the type AT (actually reverse Watson–Crick base pairs). Presumably GC base-pairs would have destablized the structure.</li>
</ol>
<p>(You can inspect this structure in three-dimensions <a href="https://www.rcsb.org/3d-view/1JUU" rel="nofollow noreferrer">here</a>. Choose ‘Licorice’ style and colour ‘by chain’ and notice the non-planarity of the three base ‘pairs’ at each end.) </p>
<p>So although the question refers specifically to parallel DNA helices with Watson–Crick base pairs, it should be recognized that extended parallel DNA helices composed of any kind of complementary AT and GC base pairs are not found, and the question applies equally well to them.</p>
<blockquote>
<p>“In the diagram if we were to flip one of the strands while keeping the other same, hydrogen bonding is still possible”</p>
</blockquote>
<p>The diagram in the question is <em>two-dimensional</em>; DNA is <em>three-dimensional</em>. It is only by considering the three-dimensional structure of DNA can you approach this question. </p>
<p>So how would one do that? One must consider the free energy of alternative structures in the relevant millieu to determine which will occur (i.e. be more thermodynamically stable). </p>
<ol>
<li><p>This will tell you whether single DNA strands with parallel sequences will form a double-stranded (ds) structure or not.</p></li>
<li><p>This will tell you whether ds-parallel DNA is more or less energetically stable than a ds-antiparallel DNA. Hence, even if both can form (which I doubt, without some special circumstances*) the lower thermodynamic free energy of the anti-parallel dsDNA would give organisms adopting it an evolutionary advantage. </p></li>
</ol>
<p><strong>And the answer to the question?</strong> </p>
<p>It seems unlikely that one single factor is responsible or it would have been pointed out in elementary text books such as <a href="https://www.ncbi.nlm.nih.gov/books/NBK22386/#A636" rel="nofollow noreferrer">Berg <em>et al.</em></a>.</p>
<p>To answer would require a complete theoretical analysis of the structure or structures. First one would have to build a model of a proposed parallel structure that could accommodate Watson–Crick base pairs. This in itself is a problem because there are likely to be many alternative structures. Perhaps there are computer programs that can find the structure with the lowest energy. This would be calculated in the classic manner, calculating the positive contribution of hydrogen bonding (which depends on distance and angle), ionic interaction etc.* against the negative contribution of charge and steric repulsions.</p>
<p>*Etc? The two-dimensional diagram fails to consider the contribution of base <a href="https://en.wikipedia.org/wiki/Stacking_(chemistry)" rel="nofollow noreferrer">stacking</a> (how could it?), which contributes to a considerable extent to the stability of nucleic acid helices, as the original cover design of Stryer’s <em>Biochemistry</em> is a constant reminder:</p>
<p><a href="https://i.sstatic.net/IzEeh.jpg" rel="nofollow noreferrer"><img src="https://i.sstatic.net/IzEeh.jpg" alt="Cover of Stryer"></a></p>
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DNA replication
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Is PCR a DNA cloning technique?
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https://biology.stackexchange.com/questions/57626/is-pcr-a-dna-cloning-technique
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<p>According to <a href="https://www.ncbi.nlm.nih.gov/books/NBK21129/#A5997" rel="nofollow noreferrer">Genomes</a></p>
<p>PCR is </p>
<blockquote>
<p>A technique that results in exponential amplification of a selected region of a DNA molecule [in test tube].</p>
</blockquote>
<p>DNA cloning is</p>
<blockquote>
<p>Insertion of a fragment of DNA into a cloning vector, and subsequent propagation of the recombinant DNA molecule <strong>in a host organism</strong>.</p>
</blockquote>
<p>While, <a href="https://books.google.co.in/books?id=jK6UBQAAQBAJ&printsec=frontcover&dq=Molecular%20Biology%20of%20the%20Cell&hl=en&sa=X&redir_esc=y#v=onepage&q&f=false" rel="nofollow noreferrer">Molecular Biology of the Cell</a>, Sixth Edition says in a summary:</p>
<blockquote>
<p><strong>DNA cloning</strong> (through the use of either cloning vectors or the <strong>polymerase chain reaction</strong>) in which portion of a genome (often an individual gene) is purified away from the remainder of the genome by repeatedly copying to generate many billions of identical molecules.</p>
</blockquote>
<p>Here DNA cloning has been used to mean DNA replication in general.</p>
<p><strong>Question:</strong> Which author's view is correct, or at least provides the most accepted definitions?</p>
|
<h3>Short answer</h3>
<p>The Oxford English Dictionary is quite clear on this. For the verb <em>clone</em> there are two meanings:</p>
<p><strong>Biology</strong> To propagate (an organism or cell) as a clone.</p>
<p><strong>Molecular Biology</strong> To make copies of (a DNA sequence or gene).</p>
<p>The latter definition clearly encompasses PCR.</p>
<h3>History lesson</h3>
<p>Most of the information below is taken from the Oxford English Dictionary.</p>
<p>The term <em>clone</em> has a long history, first as a noun and more recently as a verb. The evolution of the meaning and use of the word suggests that we shouldn't be too precious about this.</p>
<blockquote>
<p>When I use a word,” Humpty Dumpty said, in rather a scornful tone, “it means just what I choose it to mean—neither more nor less.” “The question is,” said Alice, “whether you can make words mean so many different things.” “The question is,” said Humpty Dumpty, “which is to be master—that’s all.</p>
</blockquote>
<p>The word is based on a Greek word for twig and was originally used as a noun in the form <em>clon</em>.</p>
<blockquote>
<p><em>1903 H. J. Webber in Science 16 Oct. 502/2</em> Clons..are groups of plants that are propagated by the use of any form of vegetative parts.</p>
</blockquote>
<p>It soon gained the final <em>e</em>.</p>
<blockquote>
<p><em>1905 C. L. Pollard in Science 21 July 88/1</em> I therefore suggest clone (plural clones) as the correct form of the word.</p>
</blockquote>
<p>By 1930 it had become a verb...</p>
<blockquote>
<p>1930 <em>Jrnl. Ecol. 18 357</em> This is probably a record for the number of ‘individuals’ obtained at one time by cloning a herbaceous plant</p>
</blockquote>
<p>...and, presumably because bacteria were considered to be plants at that time, it was used in microbiology too.</p>
<blockquote>
<p><em>1929 Bibliographia Genetica 5 234</em> In Bacillus coli communis...a biotype was also found having lower motility than the remainder of the clone from which it came.</p>
</blockquote>
<p>By the early 1960s it was extended to animals, probably as a direct result of Gurdon's experiments with <em>Xenopus</em></p>
<blockquote>
<p><em>1963 J. B. S. Haldane in G. Wolstenholme Man & his Future 352</em> Perhaps the first step will be the production of a clone from a single fertilized egg, as in Brave New World.</p>
</blockquote>
<p>And finally the meaning was extended to encompass DNA molecules, thus <em>molecular cloning</em>.</p>
<blockquote>
<p><em>1974 Proc. National Acad. Sci. U.S.A. 71 3459/1</em> ColE1 has been shown to serve as an effective molecular vehicle for cloning and amplifying specific regions of unrelated DNA.</p>
</blockquote>
<p>(Extra points if you have read this far AND you know what ColE1 is.)</p>
<p>This final quotation seems to me to show that the confusion over what molecular biologists mean by cloning arose very early. If cloning means “to make copies of” then why is the word <em>amplifying</em> there? From the outset I think that molecular cloning carried the sense of purifying (by cutting and ligating). Or that's what I thought when I did it for the first time in 1980.</p>
<p><strong>Update</strong></p>
<p>Despite my reliance on dictionary definitions above, and prompted by Tyto alba's discovery of a contradictory definition in a different OUP publication (see comment) here's what I <strong>really</strong> think:</p>
<p>Anyone who has ever obtained a bacterial colony containing a plasmid with the inserted fragment that they wanted announced "I have cloned the gene!" and meant that they had made a recombinant plasmid AND got it into cells. On the other hand, if they started with a PCR amplification then ran a gel to check before ligating and saw a fragment of the expected size they did not say "I have cloned the gene!" And finally, the successful cloner, when streaking out their transformant colony, will rarely have referred to this step as cloning.</p>
| 285
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DNA replication
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How is it possible for phosphate to form two ester bonds in DNA replication?
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https://biology.stackexchange.com/questions/82364/how-is-it-possible-for-phosphate-to-form-two-ester-bonds-in-dna-replication
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<p>I understand that in phosphodiester bond formation, two hydroxl groups on the phosphate molecule bind to the 3' and 5' OH groups on two independent pentose sugars. This is a condensation reaction, so two molecules of water are released.</p>
<p>I am just confused about the hydroxl groups on the phosphate. From what I know, the phosphate molecule looks like this.</p>
<p><a href="https://i.sstatic.net/7DWK1.png" rel="nofollow noreferrer"><img src="https://i.sstatic.net/7DWK1.png" alt="enter image description here"></a></p>
<p>How is phosphate able to do a condensation reaction with two pentose sugars if it has no hydroxide groups?</p>
<p>Thanks a lot</p>
| 286
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DNA replication
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Don't understand how multiple replication bubbles work
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https://biology.stackexchange.com/questions/68717/dont-understand-how-multiple-replication-bubbles-work
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<p>I'm not exactly sure how multiple replication bubbles work, assuming were working with a linear, eukaryotic chromosome.</p>
<p>This is a diagram for reference:</p>
<p><a href="https://i.sstatic.net/LMG6h.png" rel="nofollow noreferrer"><img src="https://i.sstatic.net/LMG6h.png" alt="enter image description here"></a></p>
<p>It appears that the DNA is being synthesized "towards" the centre. What I mean is that in one replication bubble, the top strand is being replicated from left to right, whereas for the other replication bubble, the top strand is being replicated from right to left.</p>
<p>I don't understand why this is. Aren't the directions reversed? Isn't synthesis always supposed to be from 5' to 3'?</p>
<p>Thanks!</p>
|
<p>You mistake replication direction with polymerase synthesis direction. Indeed, the polymerase synthesizes new strands 5' -> 3' but if the replication of each strand was continuous, there would be no such structure as a replication bubble whatsoever. Take a good look at the drawings:
<a href="https://i.sstatic.net/S48Is.png" rel="nofollow noreferrer"><img src="https://i.sstatic.net/S48Is.png" alt="DNA replication"></a></p>
<p><a href="https://i.sstatic.net/BJMt8.png" rel="nofollow noreferrer"><img src="https://i.sstatic.net/BJMt8.png" alt="Replication bubbles"></a></p>
<p>The thing is the direction of replication is consistent with the direction of leading strand synthesis, so the bubbles will "spread" outwards. Lagging strand is synthesized in the form of fragments 5' -> 3'. If you look closely fragments that are closest to the center of the replication bubble are synthesized first, then polymerase detaches and works on the next fragment which is actually closer to the 3' end of the lagging strand.
Here's an article on Wikipedia about those fragments: <a href="https://en.wikipedia.org/wiki/Okazaki_fragments" rel="nofollow noreferrer">https://en.wikipedia.org/wiki/Okazaki_fragments</a></p>
<p>Hope this makes things clear.</p>
<p>Cheers and happy learning,</p>
<p>Krzysztof</p>
| 287
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DNA replication
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What is the purpose of two cell divisions in meiosis?
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https://biology.stackexchange.com/questions/55832/what-is-the-purpose-of-two-cell-divisions-in-meiosis
|
<p>At the moment, my thoughts are that the two cell divisions are necessary for recombination to occur, although I am not sure. I cannot really see why technically, the chromosome from each parent cannot just recombine with the other chromsome when each is a single DNA strand and not in the form of two sister chromatids joined at the centromere. Something tells me that there are several reasons why there are two cell divisions. Perhaps the S phase of interphase triiggers some further stages of cell division such as cytokinesis, and the splitting of the cell will not occur unless some DNA replication has taken place? However in meiosis 2 there is no further DNA replication, so this can't quite be right...</p>
|
<p>For developing a 2N cell, we need a N cell from each parent. In any division(meiosos or meitosis), chromosomes are doubled at first. In firs meiosis a 2N cell in divided into two N cells and as you know these N chromosomes are doubled( 2N chromatids). In second meiosis a N cell is divided into two N cells but this time chromosomes are not doubled(in fact N chromatid cells)
1.Firs division in meiosis is needed because we need N cells to combine, if they are 2N the result of recombination is a 4N cell.
2.Maybe your question is: why a 2N cell is not directly divided into two N cells without doubling? the reason is same in mitosis and meiosos, it is because of a stage of division as you know it is called anaphase in which a kind of protein, connects centromer to centriol, this stage is needed because we need one chromatid of each chromosome in each cells so if this part is not done there is no guarantee to have all N chromosomes in both cells and in fact a kind of distribution is made.
( I hope that I could understand you, well.)</p>
| 288
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DNA replication
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Following DNA replication during S-phase of the cell-cycle, are all genomic regions subjected to the same stringent level of DNA-Repair?
|
https://biology.stackexchange.com/questions/35141/following-dna-replication-during-s-phase-of-the-cell-cycle-are-all-genomic-regi
|
<p>To my (limited) understanding, there are 2 main ways that mutations can occur in DNA: Environmental (UV, etc) and mistakes during cell division.</p>
<p>I was wondering if there is a mechanism that can give priority to certain genes to be accurately duplicated. Some sort of trigger that says "<em>double-check this specific gene</em> before continuing with the duplication".</p>
<p>And if there <em>is</em> such a mechanism, then I wonder if there is some sort of dependency system for genes that control groups of other genes. So that if a certain gene "activates" the double-check trigger, it would automatically add that trigger to the group of genes which are affected by it.</p>
<p>Thanks.</p>
|
<p>So far, the known mechanism of DNA repair is to recognize mismatches or damaged nucleotides by enzymes surrounding DNA rather than by scanning along DNA. Therefore double check could not happen under such conditions. </p>
| 289
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DNA replication
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Transcription takes place from the 5’ to the 3’ end of the m-RNA. Why?
|
https://biology.stackexchange.com/questions/14093/transcription-takes-place-from-the-5-to-the-3-end-of-the-m-rna-why
|
<p>Only one side of the DNA ladder is copied (the sense side). The sense side starts with a 3’ end. This means the corresponding mRNA will have to assemble starting from the 5’ end. This is my initial thought, but can someone expand on it? Also, is this explained by why replication is performed in the 5' to 3' direction as suggested by this thread: <a href="https://biology.stackexchange.com/questions/477/why-is-dna-replication-performed-in-the-5-to-3-direction">Why is DNA replication performed in the 5' to 3' direction?</a></p>
<p><img src="https://i.sstatic.net/9yepb.png" alt="enter image description here"></p>
| 290
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DNA replication
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Apparant inconsistency in DNA topology theory in formation of origin of replication
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https://biology.stackexchange.com/questions/108048/apparant-inconsistency-in-dna-topology-theory-in-formation-of-origin-of-replicat
|
<p>I'm studying an introductory course in genetics and came across something I don't fully understand. I obviously used Google to find where I'm thinking wrong, but I still can't understand it.</p>
<p>To catalyse strand separation, a negative supercoil is introduced due to the binding of DnaA-proteins, so the strands aren't being broken to form the supercoil. This means that the linking number must stay the same. Due to the negative supercoil the Writhe is decreased by 1, and thus the Twist must increase by 1 (because Lk stays the same).</p>
<p>A higher twist means that the DNA strand is overwound. Meanwhile my Textbook says this negative supercoiling implies underwounding.</p>
<p>Where am I wrong in my thinking?</p>
| 291
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DNA replication
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Do cells not grow during mitotic S phase (synthesis phase of interphase of the cell cycle)?
|
https://biology.stackexchange.com/questions/108960/do-cells-not-grow-during-mitotic-s-phase-synthesis-phase-of-interphase-of-the-c
|
<p>Both these links mention cell growth during G1 and G2 phase, but not during synthesis phase (only DNA replication is mentioned). Is replication all that happens and is there no cell growth during S phase?</p>
<p><a href="https://www.sciencedirect.com/topics/biochemistry-genetics-and-molecular-biology/interphase" rel="nofollow noreferrer">https://www.sciencedirect.com/topics/biochemistry-genetics-and-molecular-biology/interphase</a></p>
<p><a href="https://bio.libretexts.org/Bookshelves/Introductory_and_General_Biology/Book%3A_General_Biology_(Boundless)/10%3A_Cell_Reproduction/10.02%3A_The_Cell_Cycle/10.2A%3A_Interphase" rel="nofollow noreferrer">https://bio.libretexts.org/Bookshelves/Introductory_and_General_Biology/Book%3A_General_Biology_(Boundless)/10%3A_Cell_Reproduction/10.02%3A_The_Cell_Cycle/10.2A%3A_Interphase</a></p>
|
<p>By growth I'm assuming you mean something like an increase in cell mass/volume.</p>
<p>This can likely vary depending on the organism and cell type (biology is riddled with exceptions), but in budding yeast (<em>Saccharomyces cerevisiae</em>) cell mass increases during mitotic S phase<sup>1</sup>.</p>
<p>The following is a quote from the introduction to reference 1:</p>
<blockquote>
<p>Typical pie chart representation of the cell cycle (Figure 1A) stresses the discontinuous events that have to take place only once per cell cycle (i.e., S and M phases), but fails to show that proliferating somatic cells are continuously increasing in their mass throughout the cell cycle (Figure 1B). As pointed out as early as 1971 by Mitchinson<sup>2</sup>, the “continuous events of the growth cycle” (i.e., increase in cell mass) and the “discontinuous events of the DNA division cycle” (i.e., DNA replication, mitosis, and cell division) need to be tightly coordinated in order to maintain cell size homeostasis.<p>
<a href="https://i.sstatic.net/TdXlW.png" rel="nofollow noreferrer"><img src="https://i.sstatic.net/TdXlW.png" alt="enter image description here" /></a></p>
<h3>Main Events That Occur during the Yeast Cell Cycle<p></h3>
<p>(A) General representation of the cell cycle showing the discontinuous events that have to take place only once per cell cycle, namely the S phase and the M phase, spaced with G1 and G2 phases that allow increase of the cell size before DNA replication and cell division, respectively.<p>
(B) During the dynamics of the cell cycle, RNA and proteins increase exponentially, while the DNA content show a typical doubling amount until the cell divides to generate a newborn daughter. From G1 to M phases, the cell increases continuously in mass.</p>
</blockquote>
<h3>References:</h3>
<ol>
<li><a href="https://www.ncbi.nlm.nih.gov/pmc/articles/PMC1851985/" rel="nofollow noreferrer">Barberis, M., Klipp, E., Vanoni, M., & Alberghina, L. (2007). Cell size at S phase initiation: an emergent property of the G1/S network. PLoS computational biology, 3(4), e64.</a></li>
<li><a href="https://agris.fao.org/agris-search/search.do?recordID=US201300474975" rel="nofollow noreferrer">Mitchison, J. M. (1971). Biology of the cell cycle.</a></li>
</ol>
| 292
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DNA replication
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Where does (retro)virus replication take place?
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https://biology.stackexchange.com/questions/109386/where-does-retrovirus-replication-take-place
|
<p>When a virus replicates, it has to create several copies of its genome to the "daughter viruses"? Where in the cell does this replication of the viral genome take place? And how?</p>
<p>In my book, they use HIV-viruses as an example, which I've understood is a retrovirus. The first steps they describe are attachment, entry and uncoating, which I understand. Then, the next step is integration, which they say start with that the RNA translates to DNA and the new DNA strand works as a template to create a dsDNA. dsDNA is then integrated in the host cells genome, and then after that the transcription and translation of virus protein can start. What I don't understand is when the replication of RNA strands take place? After the synthesis of protein, there is the assembly, where the protein and RNA build up new viruses. Where does the RNA come from? Does the replication take place before the integration? Or when the DNA is integrated in the host cell genome, at the same time as the protein synthesis?</p>
<p>For other types of virus, they say that the genome replicates by a "rolling circle mechanism"? Where does that take place? In the cytoplasm? In those cases, isn't the viral genome integrated in the host cells genome at all? They just use the host cells polymerase, ribosomes etc.? Is retro viruses the only viruses that integrates in the genome?</p>
<p><strong>This became kind of long, but to summarize my questions:</strong></p>
<ol>
<li>Retroviruses: Where and how does the replication of the RNA take place? Before or during integration of the host cell genome?</li>
<li>Where does the replication of genome of other viruses (that use the rolling circle mechanism) take place? In the cytoplasm?</li>
</ol>
|
<h2>First Question:</h2>
<p>Integration is essential for retroviruses, otherwise cDNA isn‘t transcribed efficiently into RNA (viral genome). Eucaryotic transcription requires multiple factors that are only present in the nucleus and work best when associated to genomic DNA (host). Retroviruses also lack an RNA-dependent RNA polymerase, which would replicate viral genome in the cytosol.</p>
<p><a href="https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3385939/" rel="nofollow noreferrer">Craige & Bushman 2012:</a></p>
<blockquote>
<p>Integration of a DNA copy of the viral genome into a host cell chromosome is an essential step in the retroviral replication cycle (Varmus et al. 1989; Coffin et al. 1997).</p>
</blockquote>
<p>Textbook like reference <a href="https://www.nature.com/scitable/content/replication-of-retroviruses-14465196/" rel="nofollow noreferrer">Ref 1</a>, <a href="https://www.niaid.nih.gov/diseases-conditions/hiv-replication-cycle" rel="nofollow noreferrer">Ref 2</a></p>
<blockquote>
<p>The viral DNA is transported across the nucleus, where the HIV protein integrase integrates the HIV DNA into the host’s DNA. The host’s normal transcription machinery transcribes HIV DNA into multiple copies of new HIV RNA. <strong>Some of this RNA becomes the genome</strong> of a new virus, while the cell uses other copies of the RNA to make new HIV proteins.</p>
</blockquote>
<p>For more details see <a href="https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7761857/" rel="nofollow noreferrer">Dutilleul et al. 2020</a></p>
<blockquote>
<p><strong>Once randomly integrated</strong> into the host cellular genome, HIV-1 gene expression is mainly regulated at the transcriptional level by <strong>hijacking the cellular RNA polymerase II</strong> (RNAPII) machinery. HIV-1 transcription initiates at the U3/R junction in the 5′-long terminal repeat (5′LTR) and is regulated by cellular transcription factors […]</p>
</blockquote>
<p>So Replication happens after insertion of cDNA (complementary DNA, product of reverse transcription and second strand synthesis) into the host genome in the nucleus. Then transcription factors can bind the promoter and initiate elongation.</p>
<h2>Second Question:</h2>
<p>For DNA- viruses, some require integration into host genome, too. Interestingly, non-integrated viral DNA can populate the nucleus as dormant (circular) episomes (see <em>viral latency</em>, <a href="https://www.ncbi.nlm.nih.gov/pmc/articles/PMC6980450" rel="nofollow noreferrer">De Leo et al. 2020</a>). Non-integrating DNA viruses often bring a DNA-dependent DNA polymerase and can replicate in the cytosol.</p>
<p>(If interested, see this review on viral polymerases that enable replication of viral genomes in the cytosol <a href="https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4711277/" rel="nofollow noreferrer">Choi 2016</a>)</p>
<h3>About Rolling Circle Replication:</h3>
<p>Rolling circle replication is mainly found in procaryotic viruses (<strong>Phages</strong>). For eucaryotic viruses, rolling circle is pretty rare. The Geminivirus however, is one example of a non integrating DNA virus that replicates by rolling circle within the <strong>nuclei</strong> of plants. It seems that it relies heavily on nuclear factors for efficient DNA-Replication <a href="https://www.sciencedirect.com/science/article/abs/pii/S0022283699929169" rel="nofollow noreferrer">Castellano et al 1999</a></p>
| 293
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DNA replication
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What is a DNA clamp exactly?
|
https://biology.stackexchange.com/questions/67489/what-is-a-dna-clamp-exactly
|
<p>I used to think that a DNA clamp is a protein. But today I noticed it doesn't appear in <a href="https://upload.wikimedia.org/wikipedia/commons/8/8f/DNA_replication_en.svg" rel="nofollow noreferrer">this</a> picture. Then I went to it's Wikipedia page, where it was written: </p>
<blockquote>
<p>A DNA clamp, also known as a sliding clamp, is a protein fold that
serves as a processivity-promoting factor in DNA replication. As a
critical component of the DNA polymerase III holoenzyme, the clamp
protein binds DNA polymerase and prevents this enzyme from
dissociating from the template DNA strand. The clamp-polymerase
protein–protein interactions are stronger and more specific than the
direct interactions between the polymerase and the template DNA strand</p>
</blockquote>
<p>Which I find a little confusing. For now it seems to me like DNA clamp is a subunit of DNA Polymerase and it doesn't have any function by itself and it is not an actual protein but it binds to Polymerase and becomes functional. Can somebody please clarify this matter to me?! </p>
|
<p>Your partly right both ways. In a sense, the DNA clamp is a protein, in another sense, it's only part of a protein. What it actually is is what we call a protein sub-unit, which <em>is</em> a protein, but which binds with other protein sub-units to form complex proteins.</p>
<p>In order to understand this, you have to remember what a protein is. A protein is, in essence, a chain of amino acids. Those can be short chains, long chains, medium chains: but they're all proteins. Sometimes, a few short proteins (sub-units) will bind together to form one long protein (protein complex). This is what happens in the case of a DNA clamp.</p>
<p>Basically, the DNA clamp is (an) independent protein unit(s) which has (have) the ability to encircle the DNA strand and travel along it. They bind to the polymerase, which is why you don't see them on your diagram. Their function is, very simply, to make it possible for the polymerase to stay closely attached to the DNA stand.</p>
<hr>
<p>Sources:</p>
<ul>
<li><a href="https://www.ncbi.nlm.nih.gov/pmc/articles/PMC2691718/" rel="nofollow noreferrer">DNA Repair (Amst). 2009 May 1; 8(5): 570–578</a></li>
</ul>
| 294
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DNA replication
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ATP required for cell processes
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https://biology.stackexchange.com/questions/34783/atp-required-for-cell-processes
|
<p>I haven't been able to find anything that tells me how much ATP is needed for DNA replication, transcription, and translation in humans, just papers that mention ATP used in those processes.</p>
<p>I need to know how much ATP is needed for these processes because once my cell from scratch is alive I feed each cell 50 nanograms of glucose which yields 5 trillion ATP. If I use these numbers plus the numbers for other metabolic processes I get roughly the amount of ATP needed per cell before mitosis. If I know this then I will know if I need to feed my cells more glucose or if 50 nanograms is enough.</p>
<p>So how much ATP is needed for DNA replication, transcription, and translation? I want it in terms of per nucleotide and per amino acid so that I can multiply that by the amount of DNA total, amount of DNA in any given gene, and amount of amino acids in any given protein.</p>
|
<p>For DNA replication and transcription you need NTPs. In a dsDNA purine content will be same as pyrimidine content. I am considering that all nucleotides are synthesized <em>de novo</em> which would consume more ATP than getting nucleotides from the salvage pathway.
<hr>
<img src="https://i.sstatic.net/JUC4E.png" alt="enter image description here"></p>
<blockquote>
<p>Fig 1: Pyrimidine synthesis. Taken from <a href="http://en.wikipedia.org/wiki/Nucleotide#Synthesis" rel="nofollow noreferrer">Wikipedia.</a></p>
</blockquote>
<hr>
<p> <img src="https://i.sstatic.net/tSIGZ.png" alt="enter image description here"></p>
<blockquote>
<p>Fig 2: Purine synthesis. Taken from <a href="http://en.wikipedia.org/wiki/Nucleotide#Synthesis" rel="nofollow noreferrer">Wikipedia.</a></p>
</blockquote>
<hr>
<p>CTP synthesis requires one extra ATP:</p>
<pre>UTP + Glutamine + ATP + H<sub>2</sub>O → CTP + ADP + P<sub>i</sub></pre>
<p>UTP synthesis requires 2 (Step 1; Fig 1) + 1 (Step 5; Fig 1: PRPP requires 1 ATP to form) + <br> 2 (UMP → UTP) = 5 ATPs</p>
<p>CTP requires 6 ATPs</p>
<p>GTP requires 8 ATPs (Fig 2) (ATP synthesis would also require 8 ATPs but for simplicity let us assume that ATP is already present).</p>
<p>Conversion of dNTP from NTP requires thioredoxin which in turn requires NADP but let us forget that.</p>
<p>Assuming a 50% GC content one base pair of DNA would require in an average 10 ATPs (assume a stretch of 4nt — ACTG, this would require 1+6+5+8 ATPs; i.e. 5 ATPs per nt; 10 ATP per bp)</p>
<p>For RNA the ATP consumption would be 5 ATPs per nt.</p>
<p>For protein synthesis, for every codon, the elongation factor requires <a href="http://en.wikipedia.org/wiki/Translation_%28biology%29" rel="nofollow noreferrer">one molecule of GTP</a> and <a href="http://en.wikipedia.org/wiki/Aminoacyl_tRNA_synthetase" rel="nofollow noreferrer">aminoacyl-tRNA synthetase</a> requires 1 molecule of ATP. So the net consumption per amino acid would be 9 ATPs.</p>
<p>I am ignoring the initiation reaction that also requires ATP to phosphorylate the initiation factor. Also, I assume that amino acids are obtained from diet and are not synthesized by the organism. </p>
| 295
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DNA replication
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How to calculate information content of a DNA sequence
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https://biology.stackexchange.com/questions/45289/how-to-calculate-information-content-of-a-dna-sequence
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<p>How does one calculate the information content of DNA sequence like ATCGGCT where mutation rate of G's is 10% and the most common mutation product binds with C and A with equal frequency.</p>
<p>I know that the individual information of a sequence is the dot product of the sequence and the weights matrix for each base. Essentially I = sum (base1xweight + base2xweight + ...)</p>
<p>What I can't figure out is how to determine the the weight of each base so as to incorporate the 10% probability of mutation of G's.. Can anyone help me figure that out</p>
<p>EDITED since wording may be confusing:
Original problem says:
" Spontaneous deamination of exocyclic amine is dG (deoxyguanosine) can occur occasionally in DNA strand. Given probability of this occurring during the time it takes DNA polymerase to copy DNA is 20% per dG in a particular DNA. Given that most common deamination product pairs with dC and dA with equal frequency calculate information content during DNA replication in the given sequence. "
As an example I choose ATCGGCT and changed it to 10% deamination of dG.</p>
<p>PROPOSED SOLUTION:
For sequence ATCGGCT,
I am only consider the initial strand bases A,T,C are 2 bits a base, given by I=-1/ln(2)ln(n_before/n_after) ,where n_before = 1, and n_after = 4
In the case of mutated G (Gm) ,
I say, I = -1/ln(2)ln(1/3)= 1.58 bits , n_after=3 since A,C are equivalent to Gm</p>
<p>Perfect copy would give us 7 bases x 2 bits each = 14 bits content of strand.
Since there are 2 G's, with 10% change of mutating --> 0.02 of G in strand will mutate. </p>
<p>Thus I content during replication,
Information content I(ideal) - I(inc. mutation) = 14-0.03 = 13.97 bits during time of replication</p>
| 296
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DNA replication
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In the lysogenic cycle, does the provirus split from the cell's main genetic material for replication?
|
https://biology.stackexchange.com/questions/15613/in-the-lysogenic-cycle-does-the-provirus-split-from-the-cells-main-genetic-mat
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<p>In a diagram of the lysogenic cycle sent by my instructor in a video, it shows the provirus splitting from the cell's main DNA when the dormancy period ends and the viral DNA is "activated". Is this how it happens, or was he just trying to illustrate the activation?</p>
<p>I guess a better way of phrasing this would be, is the viral DNA replicated as part of the cell's normal DNA, or is it replicated separately?</p>
<p>Thanks!</p>
<p>evamvid</p>
|
<p>When the virus is integrated into the hosts genome (and becomes a provirus) it is replicated with the cell genome, since it is now part of it. When the provirus gets activated (this happens by changes in the host's environmental conditions or health), it will get transcribed.
This is followed by the translation of the viral proteins which then leads to a cell which exclusively produces viruses until its resources are exhausted and the cell gets destroyed.</p>
| 297
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cell signaling pathways
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Question about cell signaling pathways (RTK, Jak-Stat, SMAD, etc)
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https://biology.stackexchange.com/questions/7393/question-about-cell-signaling-pathways-rtk-jak-stat-smad-etc
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<p>I am in an embryology course right now and we've just started covering cell-cell communication in development. We were talking about the roles of the various cadherins and their discoveries but we got to cell signaling pathways and in reading my textbook I'm finding myself confused by these pathways-- some of those that are mentioned include Notch, Wnt, FGF, TGF-beta, RTK, Jak-Stat, SMAD, integrins, PTHrP, Hedgehog, Discoidin domain receptors 1&2, and the unfolded protein response (UPR). </p>
<p>I know the RTK pathway and I know the SMAD pathway, but the others just seem to be specific ligands that are using these pathways -- is this correct?</p>
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<p>@Alexandria Jak/Stat are two families of proteins which mediate signals through phosphotyrosines. </p>
<p>JAK is a tyrosine kinase which binds to cell receptors and STAT is dimerized by JAK action. JAK specificity seems to be your question. A specific JAK protein (e.g. JAK1 or JAK2..) may mediate for <a href="http://www.sciencedirect.com/science/article/pii/S1074552111000883" rel="nofollow noreferrer">different receptors in different cells</a>. </p>
<p>There may be hetero- or homo-dimers of JAK, there may be more than one receptor in a cell using the same JAK enzyme, which may result in cross-talk between the two signals. I think its a pathway dynamic that can vary quite a bit from one cell to another, depending on the receptor and cytokine environment the cell is seeing. JAK action can be modulated by any number of proteins which respond to other pathway signals or the cell state. The diagram below from a <a href="http://www.sciencedirect.com/science/article/pii/S016372580800140X" rel="nofollow noreferrer">review of heart muscle cell JAK action</a> is not unusual nor is it complete. </p>
<p><img src="https://i.sstatic.net/A2wP1.jpg" alt="enter image description here"></p>
| 298
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cell signaling pathways
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Contact Inhibition of Cell Division: Signaling Pathway
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https://biology.stackexchange.com/questions/30707/contact-inhibition-of-cell-division-signaling-pathway
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<p>The following article refers to contact inhibition of cell division in epithelial cells, specifically MDCK cells: <a href="http://www.pnas.org/content/109/3/739.full" rel="noreferrer">Collective and single cell behavior in epithelial contact inhibition</a>.</p>
<p>In their review of the literature, there are a number of possible signalling pathways implicated in contact inhibition of cell division. These pathways are outlined in the following quotes:</p>
<blockquote>
<p>It is widely accepted that contact inhibition requires establishment of E-cadherin-mediated cell-cell contacts and subsequent maturation of the adherens junctions (AJs) that link E-cadherin and F-actin in a synapse-like complex involving numerous other proteins</p>
<p>One possible pathway involves β-catenin, a mediator of Wnt signalling, that, in addition to its function as a transcriptional cofactor, is associated with the AJs at the cell surface</p>
<p>A contact inhibition role has been reported for NF2/Merlin, a tumor suppressor gene that encodes a membrane-cytoskeletal scaffolding protein, which most likely acts via the Hippo kinase pathway, controlling nuclear localization of the transcriptional activator YAP—itself a known regulator of cell proliferation.</p>
<p>Contact inhibition is known to involve the MAPK pathway, which, in turn, promotes cell cycle entry by regulating the expression of cyclinD1. Also implicated are Nectins—a family of cell adhesion molecules that are involved, together with integrins and other proteins, in the regulation of cell motility and proliferation. <strong>Yet, this accumulated knowledge falls far short of a comprehensive picture of contact inhibition.</strong></p>
</blockquote>
<p>In the article, the findings were as follows:</p>
<blockquote>
<p>Our findings show that contact between cells is not sufficient for inhibition of mitosis in MDCK cells. Instead, inhibition of cell proliferation is a consequence of mechanical constraint that causes successive cell divisions to reduce cell area.</p>
</blockquote>
<p>Where, in the discussion, they note the following:</p>
<blockquote>
<p>Our measurements also suggest that inhibition of cell division is a distinct single cell state rather than a global state induced by cell-cell signalling across the layer, as illustrated in Fig. 4E. In fact, confluent MDCK cell cultures with an average cell density corresponding to the morphological transition are often sufficiently heterogeneous in local cell density that highly motile cells and completely arrested cells coexist in the same colony. Thus contact inhibition is a local phenomenon...</p>
</blockquote>
<p>Published later, the article <a href="http://www.pnas.org/content/111/15/5586.full.pdf+html" rel="noreferrer">Spatial constraints control cell proliferation in tissues</a> showed that reducing the amount of space in a tissue prevented entry into S-phase. Actually stretching the tissue quickly reactivated the cell cycle, and compression leads quickly to arrest. More so, they found that cells had no memory of past constraint, and were able to suggest a model of growth in relation to the spatial constraint. </p>
<p>I'm editing my question because the way I posted it initially, to me, was unanswerable. Being said, I suspect that at this point cell volume and extracellular contacts are synergistic. This is probably going to need two separate questions, but:</p>
<p>1) How does the mitotic entry machinery roughly respond to cell volume? I'm currently reading into Cdc25/Wee1 regulation in addition to some stuff about <a href="http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3736675/" rel="noreferrer">internal fluidics</a> that are influenced by water, ion channels, etc. but I still don't fully understand.</p>
<p>2) What's a relatively straightforward downstream pathway (core elements), starting at the adherens junctions, that illustrates how cell-cell signalling gets the STOP signal to the mitotic entry machinery (based on what we know)? This might require some much more in-depth knowledge about the field, however, but articles with some good figures are appreciated, too.</p>
|
<blockquote>
<p>How does the mitotic entry machinery roughly respond to cell volume?</p>
</blockquote>
<p>This is a broad question but luckily there is one article that addresses this precise question in detail. However, it is very likely that more discoveries are made on this topic in future.</p>
<p><a href="https://doi.org/10.1016/j.cell.2019.01.018" rel="nofollow noreferrer">Neurohr et al. (2019)</a> have extensively studied the effect of cell volume on mitotic re-entry and cell cycle progression. They used <em>Saccharamyces cerevesiae</em> with temperature sensitive alleles of CDC28 to arrest the cells in G1 phase. At this stage high glucose concentration (2%) led to increase in cell volume compared to that of starved (0.1% glucose) or protein synthesis inhibited (2% glucose + cycloheximide) cells. Using this model they find that cytoplasmic dilution causes impairment of transcriptional regulation during cell cycle progression. This is because the <strong>transcription factors do not scale with cell size</strong>. </p>
<p>Read the paper for more details. </p>
<hr>
<blockquote>
<p>What's a relatively straightforward downstream pathway (core
elements), starting at the adherens junctions, that illustrates how
cell-cell signalling gets the STOP signal to the mitotic entry
machinery?</p>
</blockquote>
<p>This is again a broad question if you expect all details. However, google scholar search on "contact inhibition pathway" suggests that the main downstream pathway is the <a href="https://en.wikipedia.org/wiki/Hippo_signaling_pathway" rel="nofollow noreferrer">Hippo signalling pathway</a>. </p>
<p>Top hits on this topic:</p>
<ul>
<li>Zeng, Qi, and Wanjin Hong. "<em>The emerging role of the hippo pathway in cell contact inhibition, organ size control, and cancer development in mammals.</em>" <a href="https://doi.org/10.1016/j.ccr.2008.02.011" rel="nofollow noreferrer">Cancer cell 13.3 (2008): 188-192.</a></li>
<li>Kim, Nam-Gyun, et al. "<em>E-cadherin mediates contact inhibition of proliferation through Hippo signaling-pathway components.</em>" <a href="https://doi.org/10.1073/pnas.1103345108" rel="nofollow noreferrer">PNAS 108.29 (2011): 11930-11935.</a></li>
<li>Gumbiner, Barry M., and Nam-Gyun Kim. "<em>The Hippo-YAP signaling pathway and contact inhibition of growth.</em>" <a href="https://dx.doi.org/10.1242/jcs.140103" rel="nofollow noreferrer">J Cell Sci 127.4 (2014): 709-717</a>.</li>
</ul>
| 299
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cell signaling pathways
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What is the purpose of Prostaglandin F2-alpha and the Prostaglandin F receptor in the melatonin cell signaling pathway?
|
https://biology.stackexchange.com/questions/72843/what-is-the-purpose-of-prostaglandin-f2-alpha-and-the-prostaglandin-f-receptor-i
|
<p>I've been doing a lot of research recently on the melatonin cell signaling pathway for an extra credit project at school. I've included an image in this post, which is a diagram of the MT1 pathway. It seems to be a fairly standard GPCR sequence. I can identify all of the elements depicted in the diagram, but unfortunately I don't understand why some of these elements are present at all.</p>
<p>The portion of this pathway that stands out to me the most is the presence of a Prostaglandin F receptor (FP) as well as the presence of Prostaglandin F2-alpha (PGF2α). From the research that I have done, I've gathered that PGF2α is a prostaglandin found naturally in the body that can been used as an abortifacient in medical applications. I have not found any research that covers exactly why these two elements are present in the MT1 pathway.</p>
<p>Here's what I know: the Gi beta-gamma complex (Gβγi) and the Gq alpha subunit (Gαq) both activate Phospholipase C (PLC), which in this reaction causes the hydrolysis of PIP2 to create IP3 and DAG. My current hypothesis is that FP carries PGF2α into the cell, and PGF2α helps to activate PLC in some way. Unfortunately, I haven't been able to find any information online supporting this. Has anybody done any research regarding this topic, and if so can you provide any insight? I really, really appreciate it. </p>
<p><a href="https://i.sstatic.net/og7E0.png" rel="nofollow noreferrer"><img src="https://i.sstatic.net/og7E0.png" alt="MT1 Signaling Pathway"></a></p>
| 300
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cell signaling pathways
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which signalling pathway is involved in cancer?
|
https://biology.stackexchange.com/questions/19580/which-signalling-pathway-is-involved-in-cancer
|
<p>Columnar epithelial cells from the colonic mucosa are studied to identify abnormalities in cell signaling pathways. Abnormal epithelial cells from colonic adenocarcinoma are shown to have a mutation that blocks hydrolysis of GTP-bound active RAS. Normal columnar cells have active RAS protein that undergoes hydrolysis to the inactive GDP-bound form. Which of the following signaling pathways is most likely abnormally stimulated in the carcinoma cells?</p>
<p>a. Jak-Stat Pathway<br>
b. MAP kinase pathway<br>
c. p53<br>
d. Both a & b</p>
<p>I think its MAP kinase pathway because RAS is part of MAP kinase pathway that ultimately leads to transcription and expression of genes but if GTP is not hydrolyzed then RAS will be constantly activated leading to continued gene expression that may lead to cancer but I am not completely sure about it because p53 is also a tumor suppressor gene. </p>
|
<p>The answer is b, as the mutation constantly activates the RAS protein. RAS is part of the MAP-Kinase pathway, constant signaling of it permanently activates this pathway and leads to changes in gene expression. See the image below:</p>
<p><img src="https://i.sstatic.net/d2Kms.png" alt="enter image description here"></p>
<p>If you are interested in more details, have a look at these papers:</p>
<ul>
<li><a href="http://www.nature.com/nrc/journal/v11/n11/full/nrc3106.html" rel="nofollow noreferrer">RAS oncogenes: weaving a tumorigenic web</a></li>
<li><a href="http://journal.frontiersin.org/Journal/10.3389/fgene.2011.00100/full" rel="nofollow noreferrer">RAS mutations and oncogenesis: not all RAS mutations are created
equally</a></li>
</ul>
| 301
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cell signaling pathways
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Why does Hunger lead to the aggressive behavior?
|
https://biology.stackexchange.com/questions/19508/why-does-hunger-lead-to-the-aggressive-behavior
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<p>I have observed that frequently when people are hungry; they tend to get angry more easily on pointless issues. Does this mean that our fight or flight response is more active when a person is hungry? What is a possible reason for this? Is this phenomenon linked with our cell signaling pathways? If it is, then what would be the pathway that leads to the aggressive behavior?
<p>
To summarize the question: </p>
<p><strong>When a person is hungry and they get angry, is it due to a cell signaling pathway? If so, what pathway?</strong></p>
|
<p>Brain's main energy source is glucose. It uses about 20% of total glucose [1]. Brain hypoglycemia causes depressive-like behaviors in mice through adrenergic pathways [2].</p>
<p>When it comes to humans, here is a study that claims low glucose leads to increased aggression in married couples (see <a href="http://o.canada.com/health/hunger-in-the-form-of-low-blood-sugar-leads-to-greater-aggression-and-anger-in-married-couples-study-finds" rel="noreferrer">this</a> too):</p>
<blockquote>
<p>Self-control requires energy, part of which is provided by glucose. For 21 days, glucose levels were measured in 107 married couples. To measure aggressive impulses, each evening participants stuck between 0 and 51 pins into a voodoo doll that represented their spouse, depending how angry they were with their spouse. ... As expected, the lower the level of glucose in the blood, the greater number of pins participants stuck into the voodoo doll, and the higher intensity and longer duration of noise participants set for their spouse [3].</p>
</blockquote>
<p>However, the conclusion is disputed:</p>
<blockquote>
<p>Bushman et al.'s study does not demonstrate that fluctuations in blood glucose affect individuals' self-control abilities. As an important consequence, there is no reason to assume that giving couples a sugary “boost to their self-control energy” (p. 3) will reduce intimate partner violence. Because the glucose model of self-control lacks empirical foundation, it does not qualify as a framework for scientifically based intervention strategies [4].</p>
</blockquote>
<p>What is sure, is that hypoglycemia activates sympathetic nervous system:</p>
<blockquote>
<p>... the neurogenic symptoms of hypoglycemia are largely the result of sympathetic neural, rather than adrenomedullary, activation [5].</p>
<p>Hypoglycemia increases plasma levels of both epinephrine and norepinephrine. These catechols are released primarily from the adrenal medulla. However, it is well documented that hypoglycemic increases muscle sympathetic nerve activity, and that both alpha and beta adrenergic activity increase [6].</p>
</blockquote>
<p>And this leads to behavioral changes (at least in animals):</p>
<blockquote>
<p>Noradrenaline is involved in many different functions, which all are known to affect behaviour profoundly. ... Part of these effects may arise in indirect ways that are by no means specific to aggressive behaviour, however, they are functionally relevant to it. Other effects may affect brain mechanisms specifically involved in aggression. Hormonal catecholamines (adrenaline and noradrenaline) appear to be involved in metabolic preparations for the prospective fight; the sympathetic system ensures appropriate cardiovascular reaction, while the CNS noradrenergic system prepares the animal for the prospective fight. ... It appears that neurons bearing postsynaptic alpha2-adrenoceptors are responsible for the start and maintenance of aggression, while a situation-dependent fine-tuning is realised through neurons equipped with beta-adrenoceptors [7].</p>
</blockquote>
<hr />
<p>References:</p>
<ol>
<li>Wikipedia contributors, "Human brain," Wikipedia, The Free Encyclopedia, <a href="http://en.wikipedia.org/w/index.php?title=Human_brain&oldid=615456836" rel="noreferrer">http://en.wikipedia.org/w/index.php?title=Human_brain&oldid=615456836</a> (accessed July 6, 2014).</li>
<li>Park MJ, Yoo SW, Choe BS, Dantzer R, Freund GG. <a href="http://www.ncbi.nlm.nih.gov/pubmed/21820138" rel="noreferrer">Acute hypoglycemia causes depressive-like behaviors in mice.</a> Metab. Clin. Exp. 2012 Feb;61(2):229-36. doi: <a href="http://dx.doi.org/10.1016/j.metabol.2011.06.013" rel="noreferrer">10.1016/j.metabol.2011.06.013</a>. PubMed PMID: 21820138.</li>
<li>Bushman BJ, Dewall CN, Pond RS, Hanus MD. <a href="http://www.ncbi.nlm.nih.gov/pubmed/24733932" rel="noreferrer">Low glucose relates to greater aggression in married couples.</a> Proc. Natl. Acad. Sci. U.S.A. 2014 Apr 29;111(17):6254-7. doi: <a href="http://dx.doi.org/10.1073/pnas.1400619111" rel="noreferrer">10.1073/pnas.1400619111</a>. PubMed PMID: 24733932.</li>
<li>Lange F and Kurzban R (2014) Sugar levels relate to aggression in couples without supporting the glucose model of self-control. Front. Psychol. 5:572. doi: 10.3389/fpsyg.2014.00572</li>
<li>DeRosa MA, Cryer PE. <a href="http://www.ncbi.nlm.nih.gov/pubmed/14970007" rel="noreferrer">Hypoglycemia and the sympathoadrenal system: neurogenic symptoms are largely the result of sympathetic neural, rather than adrenomedullary, activation.</a> Am. J. Physiol. Endocrinol. Metab. 2004 Jul;287(1):E32-41. doi: <a href="http://dx.doi.org/10.1152/ajpendo.00539.2003" rel="noreferrer">10.1152/ajpendo.00539.2003</a>. PubMed PMID: 14970007.</li>
<li>Hoffman RP. <a href="http://www.ncbi.nlm.nih.gov/pubmed/18220670" rel="noreferrer">Sympathetic mechanisms of hypoglycemic counterregulation.</a> Curr Diabetes Rev. 2007 Aug;3(3):185-93. PubMed PMID: 18220670.</li>
<li>Haller J, Makara GB, Kruk MR. <a href="http://www.ncbi.nlm.nih.gov/pubmed/9491941" rel="noreferrer">Catecholaminergic involvement in the control of aggression: hormones, the peripheral sympathetic, and central noradrenergic systems.</a> Neurosci Biobehav Rev. 1998;22(1):85-97. PubMed PMID: 9491941.</li>
</ol>
| 302
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cell signaling pathways
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What cells are secreting Wnt pathways and under which conditions?
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https://biology.stackexchange.com/questions/112213/what-cells-are-secreting-wnt-pathways-and-under-which-conditions
|
<p><em>Former question: Where and how happen these operations in the Wnt signaling pathway?</em></p>
<p>I have read about the signaling pathway on <a href="https://en.wikipedia.org/wiki/Wnt_signaling_pathway" rel="nofollow noreferrer">wikipedia</a>:</p>
<blockquote>
<p>Wnt comprises a diverse family of secreted lipid-modified signaling glycoproteins that are 350–400 amino acids in length.[11] The lipid modification of all Wnts is palmitoleoylation of a single totally conserved serine residue.[12] Palmitoleoylation is necessary because it is required for Wnt to bind to its carrier protein Wntless (WLS) so it can be transported to the plasma membrane for secretion[13] and it allows the Wnt protein to bind its receptor Frizzled [14][15] Wnt proteins also undergo glycosylation, which attaches a carbohydrate in order to ensure proper secretion.[16] In Wnt signaling, these proteins act as ligands to activate the different Wnt pathways via paracrine and autocrine routes</p>
</blockquote>
<p>Where happen these chemical reactions described in this quote ?
My understanding is that they happen outside of cells, before the arrival of the Wnt ligand on the receptor that is located at the surface of the cell. Am I correct?
If yes, were does this happen precisely in the body? Where the ligand comes from?</p>
<hr />
<p><strong>EDIT:</strong>
From comments: The Wnt ligand is secreted by other cells and transmitted from cell to cell, this is an autocrine process.</p>
<p><strong>The question is: Under which conditions does this process start in one cell?</strong></p>
| 303
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cell signaling pathways
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Cell Signaling: What is meant by "sustained tonal induction"?
|
https://biology.stackexchange.com/questions/114836/cell-signaling-what-is-meant-by-sustained-tonal-induction
|
<p>I am reading a <a href="https://elifesciences.org/articles/32893" rel="nofollow noreferrer">journal paper</a> about the insulin-like growth factor 1 receptor. I have a question about the following statement in the paper:</p>
<blockquote>
<p>IGF/IGF-IR stimulates the PI3K-Akt pathway in a stereotypical way –
sustained tonal induction. Sustained induction is thought to define
the specific biological outcomes of IGF signaling, and distinguish the
function of the IGF ligand from other RTKs/ligands that access the Akt
cascade (Gross and Rotwein, 2016; Kubota et al., 2012). In particular,
sustained activation of the PI3K-Akt pathway, mediated by IGF-IR,
induces cell proliferation in multiple types of cells, cell survival
in neural cells, and protein homeostasis in skeletal muscle cells
(Fernandez and Torres-Alemán, 2012; Fukushima et al., 2012; Ness and
Wood, 2002; Sacheck et al., 2004; Stewart and Rotwein, 1996). To date,
the mechanism by which IGF-IR produces sustained signaling remains
poorly understood.</p>
</blockquote>
<p>What is meant by <strong>"sustained tonal induction"</strong>? I have searched online but could not find any specific results.</p>
<p>Any advice is greatly appreciated.</p>
| 304
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cell signaling pathways
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Specific examples of signalling pathway using logical 'OR' and 'AND'?
|
https://biology.stackexchange.com/questions/64573/specific-examples-of-signalling-pathway-using-logical-or-and-and
|
<p>I have read <a href="https://www.khanacademy.org/science/biology/cell-signaling/mechanisms-of-cell-signaling/a/intracellular-signal-transduction" rel="noreferrer">here</a> that
"<strong>signals from two different pathways may be needed to activate a response, which is like a logical "AND." Alternatively, either of two pathways may trigger the same response, which is like a logical "OR."</strong>
But no example is mentioned. I want to know some specific examples in which cell signalling uses logical OR and Logical AND. Any references will be appreciated.</p>
|
<p>There are thousands of examples, here I list just a few.</p>
<p>1) Macrophage activation. This is a complex case with many proteins acting as AND/OR. The following paper depicts a nice scheme that helps to understand the circuit.</p>
<p><a href="https://bmcsystbiol.biomedcentral.com/articles/10.1186/1752-0509-2-36" rel="nofollow noreferrer">https://bmcsystbiol.biomedcentral.com/articles/10.1186/1752-0509-2-36</a></p>
<p>2) The Lac operon that follows the logic:</p>
<pre><code>if low_glucose AND lactose:
express(lac_genes)
if (high_glucose OR low_glucose) AND no_lacotse:
inhibit(lac_genes)
if high_glucose AND lacotse:
express_at_low_level(lac_genes)
</code></pre>
<p><a href="https://en.wikipedia.org/wiki/Lac_operon" rel="nofollow noreferrer">https://en.wikipedia.org/wiki/Lac_operon</a></p>
<p><a href="https://en.wikipedia.org/wiki/Synthetic_biological_circuit" rel="nofollow noreferrer">https://en.wikipedia.org/wiki/Synthetic_biological_circuit</a></p>
<p>3) Phosphorylation and ubiquitination pathways. For example, </p>
<blockquote>
<p>... proteins primed through phosphorylation by one protein
kinase are often phosphorylated processively on the N-terminal side of
the priming phosphate by GSK3 at a series of Ser/Thr spaced by three
residues, with the cluster of phosphates regulating protein activity
<strong>(e.g., glycogen synthase, β-catenin)</strong>. If the two sites are
phosphorylated by different protein kinases, then this can in
principle provide a logical <strong>AND</strong> gate in a downstream response.</p>
</blockquote>
<p><a href="http://www.sciencedirect.com/science/article/pii/S1097276507007988" rel="nofollow noreferrer">http://www.sciencedirect.com/science/article/pii/S1097276507007988</a></p>
<p>4) Neurotransmitter signaling pathways.
Figure 2 of the following paper</p>
<p><a href="http://www.sciencedirect.com/science/article/pii/S0022519306003675" rel="nofollow noreferrer">http://www.sciencedirect.com/science/article/pii/S0022519306003675</a></p>
<p>describes the boolean logic underlining the signaling pathway</p>
<blockquote>
<p>Tyrosine hydroxylase activates itself in this model.
There is an “AND NOT” gate between
tyrosine hydroxylase and COMT to activate dopamine as
tyrosine hydroxylase and not COMT activates dopamine...</p>
<p>...adenylate cyclase is activated by dopamine
receptor 1 and not by dopamine receptor 2, which has been
represented by the “AND NOT” gate between the input nodes.</p>
<p>...</p>
<p>DARPP32, is activated by protein kinase A, and
not by calcineurin, therefore protein kinase A “AND NOT”
calcineurin activates DARPP32.</p>
<p>DARPP32 inhibits protein phosphatase1, which is
represented by the “NOT” gate from DARPP32 for protein
phosphatase1.
...
The activation of glutamate receptor, needs the presence of both
protein kinase A and the ligand, glutamate therefore, a
“AND” gate between protein kinase A and glutamate, “AND NOT”
protein phosphatase 1. ...</p>
</blockquote>
<p><a href="http://bioinformatics.ac.cn/synbiolgdb/" rel="nofollow noreferrer">Here the link</a> to a database of natural and artificial biological logic gates and <a href="https://www.nature.com/articles/srep08090" rel="nofollow noreferrer">here the article</a> presenting the database</p>
<p>To conclude, I would like to highlight <a href="http://science.sciencemag.org/content/340/6132/599.full" rel="nofollow noreferrer">this work</a> on the engineering of a biological transistor.</p>
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Do we know how the different functions are selected when Wnt pathway is activated?
|
https://biology.stackexchange.com/questions/112215/do-we-know-how-the-different-functions-are-selected-when-wnt-pathway-is-activate
|
<p>The Wnt signaling pathway is <a href="https://en.wikipedia.org/wiki/Wnt_signaling_pathway" rel="nofollow noreferrer">said</a> to allows multiple functions:</p>
<ul>
<li>Axis patterning</li>
<li>Cell differentiation</li>
<li>Cell proliferation</li>
<li>Cell fate specification</li>
<li>Cell migration</li>
</ul>
<p>But how are these functions "distinguished by the cell" when the Wnt signaling pathway is activated by the receptor on the cell's surface? I mean, do only specific environment conditions (and if yes, do we know which one?) triggers the duplication of the cell, or its differentiation? Or is there a co-signalling that triggers the cell into one or the other mechanism?</p>
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cell signaling pathways
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Exact definition of 'convergent' and 'divergence' in cell signalling?
|
https://biology.stackexchange.com/questions/59477/exact-definition-of-convergent-and-divergence-in-cell-signalling
|
<p>From what I understand, we refer to 'signal convergence' as being when two different ligands/stimuli lead to the same (at least in part) responses inside a single cell. This may or may not be due to activation of the same pathway, so for example we can refer to the response of a muscle cell to adrenaline and to stimulation by the nervous system via acetylcholine as convergent because they both lead to the release of calcium. We can also refer to the activation of the phosphoinositide pathway by both activation of beta phospholipase C via a heterotrimeric Gq protein, and activation of gamma phospholipase c via the tyrosine kinase pathway, as signal convergence.</p>
<p>So it seems to me that 'convergence' is defined <em>on the level of the cell</em> with it being, in a nutshell, <em>different stimuli resulting in the same effect</em> regardless of at which point in some internal signalling pathway the two responses 'converge' (at least in part).</p>
<p>Now with divergence I am getting slightly more stuck, and I have not seen a single, clearly stated, definition. Firstly, <em>does divergence refer only to the level of the cell, or also for the whole organism</em>? In particular, I am thinking of cases where a single ligand has different responses in different cells, such as adrenaline leading to increased contraction in muscle cells and glycogen breakdown in the liver (I know this isn't the best example because adrenaline technically leads to glycogen breakdown in both cell types). </p>
<p>Perhaps instead divergence is only referred to on the level of the cell, as with convergence? So for example generally tyrosine kinase stimulation activates several different proteins, which trigger different responses: ras, gamma phospholipase C etc. Even the phospholipase C pathway produces DAG and IP3, which have different responses. But then it seems to me like practically all pathways are divergent if this is the case, because generally pathways lead to several responses inside the cell. So with the case of phospholipase C which cleases PIP2 to DAG and IP3, the DAG can regulate metabolism and transcription etc via protein kinase C and the IP3 leads to calcium ion release that causes muscle contraction. Would this be referred to as divergence?</p>
<p>I would greatly appreciate if someone could just state what exactly convergence and (especially) divergence refer to. I am relatively new to cellular and molecular biology.</p>
|
<p>In my experience those terms are primarily used intracellularly, but I wouldn't argue that it is wrong to use them more broadly, it's just that essentially everything released extracellularly is going to have some level of divergence, so it makes more sense to use a separate classification scheme (i.e., endocrine/paracrine). Between cells, the terms are also used in other contexts such as the nervous system to refer to inputs from multiple neurons synapsing on one neuron (convergence), and outputs from one neuron synapsing on many targets (divergence).</p>
<p>Convergence just means multiple signalling pathways converging on the same target: like multiple pathways that can activate phospholipase C.</p>
<p>Divergence just means that one effector, which could be a protein such as a kinase or a second messenger like IP3, has multiple targets.</p>
<p>You are quite likely to get both convergence and divergence within any given signalling pathway. In my opinion, this is one of the ways that biology is not taught well in schools: textbooks might make it seem like convergence and divergence are these concrete, specific classifications, but they are really just descriptive terms. Phospholipase C pathways are a great example that displays both. If you are talking from the perspective of activation of phospholipase C, you are probably talking about convergence because there are so many ways to activate phospholipase C, but at the same time, if you talk from the perspective of the G protein subunit that is activating phospholipase C, that G protein probably has other targets as well, so that's divergence. And downstream of phospholipse C you have divergent effects of DAG and IP3.</p>
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cell signaling pathways
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What does it mean for a chemical pathway to be conserved?
|
https://biology.stackexchange.com/questions/16300/what-does-it-mean-for-a-chemical-pathway-to-be-conserved
|
<p>In many papers the MAPK pathway, (along with many others) is referred to as being conserved: </p>
<p><a href="http://www3.aiche.org/Proceedings/Abstract.aspx?PaperID=133783" rel="nofollow">Example</a>: "The mitogen-activated protein kinase (MAPK) cascades are ubiquitous in eukaryotic signal transduction, and these pathways are conserved in cells from yeast to mammals"</p>
<p><a href="http://en.wikipedia.org/wiki/Hippo_signaling_pathway" rel="nofollow">example 2</a> "The Hippo signaling pathway appears to be highly conserved."</p>
<p>What does it mean for a pathway to be conserved?</p>
|
<p>A conserved pathway is a pathway that exists in a variety of species, by virtue of that pathway being conserved throughout the evolution of those species. A pathway must by neccessity have appeared for the first time in one particular species. If that species gives rise to new species, but the pathway in question is identical or very similar in all the new species, it is sensible to say that pathway has been evolutionary conserved. This may imply that the pathway in its conserved form is important for the fitness of those species.</p>
<p>For some examples, here is a page on evolutionary conserved pathways in Drosophila: <a href="http://www.sdbonline.org/sites/fly/aimain/aadevinx.htm" rel="nofollow">http://www.sdbonline.org/sites/fly/aimain/aadevinx.htm</a></p>
| 308
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cell signaling pathways
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Textbook on molecular basis of memory
|
https://biology.stackexchange.com/questions/40465/textbook-on-molecular-basis-of-memory
|
<p>Looking at the rules in the meta, it seems book-recs are a little on the iffy side for on-topic so I hope this is okay. </p>
<p>I am looking for a (graduate-level) textbook that has a thorough treatment of the molecular basis of learning and memory. The issue I am having is that a lot of textbooks seem to cover bits and pieces or just focus on one area. Usually, they just have a few small sections on the molecular events and the rest of the textbook covers channel properties, pharmacology, systems neuro, and behavior. </p>
<ul>
<li><a href="http://rads.stackoverflow.com/amzn/click/B004L2KP58" rel="nofollow">Synapses</a> - This one comes close but is now quite dated (2003)</li>
<li><a href="http://rads.stackoverflow.com/amzn/click/1605352306" rel="nofollow">The Neurobiology of Learning and Memory</a> - A significant section on the molecular aspect but is not particularly detailed. Very basic idea.</li>
</ul>
<p>Review articles tend to be difficult because they are more specialized for specific kinases (mTOR, CaMKII etc), events, or channels.</p>
<p>I am trying to get a sense of the broader picture of the just the molecular side of all of this. Specifically the molecular and cell biology that underlies:</p>
<ul>
<li>Early-phase LTP (receptor trafficking, signaling pathways)</li>
<li>Proteins expressed from local translation in dendritic spines, signaling pathways that control this</li>
<li>Details on synaptic tagging hypothesis</li>
<li>Late-phase LTP events, pathways, proteins, and regulation</li>
<li>Downstream effects of activation of different channels, how they control pathways below, how they all intersect and how this differs by brain area and cell type</li>
<li>LTD events</li>
<li>Events at the post-synaptic density (receptors, ion-channels, protein composition, scaffolding, actin network)</li>
<li>Synaptogenesis, plasticity, maintenance during long-term memory</li>
<li>Kinases in learning and memory</li>
<li>Transcriptional control</li>
<li>Post-translational modifications and consequences for learning and memory</li>
<li>Scaffolding proteins and their contribution to signal stability</li>
<li>Pre-synaptic events</li>
<li>Contributions of glia/auxiliary neuronal cells</li>
<li>Bonus: Disease focus - what pathways and proteins underlie various well-studied genetic diseases?</li>
<li>etc.</li>
</ul>
<p>I know this is a tall order but with 1000+ page textbooks out there on other neuroscience topics, surely someone has written something? Review article are great too, perhaps I just haven't found the right one yet.</p>
| 309
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cell signaling pathways
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What is the role of RAGEs?
|
https://biology.stackexchange.com/questions/30447/what-is-the-role-of-rages
|
<p>According to articles I read, AGEs (advanced glycation end products) activate RAGEs (receptors for AGEs). This activation increases the ROS (reactive oxygen species) levels in the cells.</p>
<p><img src="https://i.sstatic.net/CnVk4.gif" alt="Reactive Oxygen Species-Regulated Signaling Pathways in Diabetic Nephropathy"></p>
<ul>
<li><a href="http://jasn.asnjournals.org/content/14/suppl_3/S241/F3.expansion" rel="nofollow noreferrer">2003 - Reactive Oxygen Species-Regulated Signaling Pathways in Diabetic Nephropathy</a></li>
</ul>
<p>Free radicals can cause damage in cells, which is definitely not good. What do you think, what is the role of RAGEs, which is more important than cellular damage?</p>
| 310
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cell signaling pathways
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How does evolution work at the level of a caterpillar mimicking a snake?
|
https://biology.stackexchange.com/questions/110588/how-does-evolution-work-at-the-level-of-a-caterpillar-mimicking-a-snake
|
<p>I am referencing <a href="https://www.youtube.com/watch?v=_5CRKTgFal0&t=35s" rel="nofollow noreferrer">this video</a>, where a caterpillar turns into what looks like a snake when it gets frightened, presumably to ward off predators.</p>
<p>Now how can this evolve?</p>
<p>I majored in Molecular Neurobiology and studied evolution, and random molecular evolution makes sense to some degree. Every now and then there are random mutations, sometimes which affect gene / cell-signaling pathways, which could cause reproductive advantages, leading to evolutionary adaptations. But to go so far as to be able to mimic an external structure like a snake, how is that possible? I don't see how that can randomly evolve.</p>
<p>It is as if as a species we have a plan on what we want to become, and we slowly <em>realize</em> that plan by somehow tuning the genes to, say, look like a snake. But that is just me trying to fill in the gaps with imaginative thinking, how something like this can evolve. There are many other examples of incredible things evolving, but this one demonstrates the question perfectly.</p>
<p>Briefly, can you explain how this sort of thing could evolve?</p>
|
<p>Because vision is not perfect, a predator could confuse a caterpillar for a snake under different conditions at different distances. is it cloudy out, is the caterpillar in shadow, partially obscured. At every step of looking more like a snake the caterpillar is less likely to be eaten in more and more situations. this is fairly easy to see because there are a variety of caterpillars that resemble snakes to varying degrees. Many are just simple spots that look like eyes. Even if it only fools only a few predators under only a few conditions it is still an advantage. This is why mimicry in general is so common.</p>
<p><a href="https://i.sstatic.net/4MvAb.png" rel="nofollow noreferrer"><img src="https://i.sstatic.net/4MvAb.png" alt="enter image description here" /></a></p>
<p><a href="https://i.sstatic.net/KOo5O.jpg" rel="nofollow noreferrer"><img src="https://i.sstatic.net/KOo5O.jpg" alt="enter image description here" /></a></p>
<p>Consider the certainty of a predator, if the predatory is just confused about whether something is or is not a snake, it can still be a big advantage. That might be a snake but that other thing is definitely a caterpillar. Which one is the predator going to investigate the possible threat or the easy meal? Then the caterpillar that does not resemble a snake gets eaten and the poor mimic gets spared. Prey selection is a risk assessment and a certainty assessment at the same time, thus is exploitable.</p>
| 311
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cell signaling pathways
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Specificity in MAPK/ERK pathway and PC12 Cells
|
https://biology.stackexchange.com/questions/58674/specificity-in-mapk-erk-pathway-and-pc12-cells
|
<p><strong>Background</strong></p>
<p>PC12 Cell stimulation leads to distinct outcomes upon stimulation with either EGF or NGF (epidermal and nerve growth factors). The outputs are transmitted through the MAPK/ERK signaling pathway; stimulation with EGF causes transient activation of the ERK, via a negative feedback loop, leading to cell proliferation whereas stimulation with NGF leads to sustained activation of ERK, via a positive feedback loop, leading to differentiation. </p>
<p><strong>Question</strong></p>
<p>How does the signaling pathway know which kind of feedback loop to initiate at the ERK level from upstream stimulation by EGF or NGF? </p>
<p><strong>Helpful Image to explain question</strong></p>
<p>To reiterate: I want to figure out how EGF binding at the membrane level triggers negative feedback downstream, and NGF binding triggers positive feedback? </p>
<p>A link to the image is (<a href="http://www.nature.com/nrm/journal/v12/n2/fig_tab/nrm3048_F4.html" rel="nofollow noreferrer">http://www.nature.com/nrm/journal/v12/n2/fig_tab/nrm3048_F4.html</a>)</p>
<p><a href="https://i.sstatic.net/rQhIg.jpg" rel="nofollow noreferrer"><img src="https://i.sstatic.net/rQhIg.jpg" alt="enter image description here"></a></p>
| 312
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cell signaling pathways
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Does the cellular response to every receptor work the same way?
|
https://biology.stackexchange.com/questions/23898/does-the-cellular-response-to-every-receptor-work-the-same-way
|
<p>I heard somewhere that activating any receptor results in the same intracellular response (signaling) which involves <a href="http://en.wikipedia.org/wiki/NF-%CE%BAB" rel="nofollow noreferrer">NF-κB</a>. If that is true, I hardly understand how the cells distinguish between different types of stimulis coming from different types of receptors. I guess I am missing the point... :S</p>
<p><img src="https://i.sstatic.net/d2mZ4.png" alt="signal transduction"></p>
<ul>
<li>Figure 1 - signal transduction - <a href="http://en.wikipedia.org/wiki/Signal_transduction" rel="nofollow noreferrer">source</a></li>
</ul>
<p>If you check this picture about signal transduction. What you can see that there are about 10 arrows pointing to the inside of the nucleus, and there are about 10 receptor categories involved. Afaik. there are much more genes in the nucleus, so how is it possible to regulate the expression of so many genes with only a few signaling pathways?</p>
|
<p>It's a complicated answer. More than 200 cell types, each type of cell inherits a unique expression of receptors, internal and external. Diffusion of signals through the plasma membrane and/or nuclear compartment may act directly as cofactors, activators, etc. The specific sequestration, and pattern of expression of external receptors also influence what signals become bound and relayed. The cell will also have a motif of internal/cytoplasmic/nuclear proteins expressed that complex or become directly activated by interaction with any of the above, etc. So the currently expressed set of internal, and external proteins determine not only how the cell interprets the signal, but also how they respond. </p>
<p>As for the actual genetic material, you'll find inherited motifs like DNA methylation, histone acetylation patterns etc. influence what parts of the DNA can actually be accessed. Per the above schema of signaling pathways, the signals received by each individual cell differ based on the actual cell type. In any case there are patterns of activators/repressors, enhancers/silencers that are switched on or off by these signals. Some journals are also introducing evidence insoluble receptors like EGFR+ligand can actually internalize and translocate to the nucleus and act as transcription regulators. </p>
<p>The final note, is that due to alternative splicing and transcription of proteins that promote or repress splicing sites, in a combination with the above concepts a cell can regulate for thousands of genes. This is a very general answer, however. The concept to take home, though, is that differentiated cells have their own specific motifs they express that make "reading" different signals possible, and also makes differing responses to what could be the same signal possible. These motifs also allow for the accurate expression of the relevant portions of the genome (not all genes are expressed at all times in all cells).</p>
| 313
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cell signaling pathways
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How do signal transduction pathways utilize transcription factors to express a specific gene?
|
https://biology.stackexchange.com/questions/37003/how-do-signal-transduction-pathways-utilize-transcription-factors-to-express-a-s
|
<p>I have an inquiry regarding the regulation of genes via extracellular signaling.</p>
<p>To my knowledge, in autocrine, paracrine, and endocrine cellular communication, large protein ligands that cannot directly diffuse through the plasma membrane of the target cell(s) use surface receptors to perform their desired action on the target cell(s).</p>
<p>I have learned that some of these ligands activate signal transduction pathways such as the MAPK/ERK and JAK/STAT pathway and drive the expression of specific genes by utilizing transcription factors (of course, in eukaryots). A simple example of this would be the action of epinephrine (adrenaline) on hepatocytes (liver cells), where the amino acid-based hormone uses the transcription factor CREB to express the gene coding for Glycogen Phosphorylase to engage in glycogenolysis.</p>
<p>Here I have two questions:</p>
<ol>
<li><p>How does the transcription factor chemically indicate which gene to express? (Is there a gene indexing system like a computer filesystem?)</p></li>
<li><p>How does the transcription factor locate and bind to the promoter of the gene it is trying to express?</p></li>
</ol>
<p>Thank you.</p>
|
<p>This is a combination of multiple regulatory systems. Most genes are not regulated by a single factor, but by many. Moreover in eukaryotic organisms there is also epigenetic, which "inactivates" permanently certain areas of the genome by compacting these zones forming <a href="https://en.wikipedia.org/wiki/Heterochromatin" rel="nofollow">heterochromatin</a>. </p>
<p>Moreover, even when we are interested in a single function a TF might have, that does not mean that it only has this function in the cell. We view it as humans, and maybe for us it is logical that a response to a high metabolite in the environment simply leads to expressing the protein that metabolizes it. However, probably it is interesting for the cell to also activate other pathways and gene networks (i.e. anabolic pathways that feed off this metabolite, compensatory pathways to mantain homeostasis...). Because that normally TFs affect more than one gene naturally. </p>
<p>The mechanisms of action of the TFs are multiple, but they tend to simply allow or deny the access to the adequate RNA polymerase. For more information about this check the <a href="https://en.wikipedia.org/wiki/Transcription_factor#Structure" rel="nofollow">wikipedia article on TF structure</a> <img src="https://upload.wikimedia.org/wikipedia/commons/thumb/8/80/Transcription_Factors.svg/1572px-Transcription_Factors.svg.png" alt=""> </p>
<p><em>In this image we see a typical mechanism of action, DNA bending</em> </p>
<p>Different domains interact in activating a gene, <strong>enhancers</strong> which are usually long distance, <strong>promoters</strong> which tend to be in the upstream region, though they can be downstream or in the gene sequence, and <strong>epigenetic factors</strong> that change the compaction of DNA, making it inaccessible to proteins. </p>
<p>Just as a last thing to point out, you should acknowledge that the biological systems are far from exact and yes, a single TF might activate 200 genes, even when its main objective in that moment is to activate one, but it won't really matter as long as the those 199 activated genes express at a very low rate (for example by not having the right RNA polymerase subunit accessible, being marked as inactive by epigenetic systems or having their own inhibitors). </p>
| 314
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cell signaling pathways
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Effect of single-gene overexpression in the cell's response
|
https://biology.stackexchange.com/questions/2505/effect-of-single-gene-overexpression-in-the-cells-response
|
<p>Which are the factors that modify the overall gene differential expression by introducing a vector for single-gene overexpression?</p>
<p>If you overexpress a gene for a protein involved in signal transduction (e.g., a kinase, scaffold, or receptor) by vector cell transfection, then you overdrive the cell using this signaling pathway, it's useful to isolate the pathway and study them.</p>
<p>Is there any way to modify the overall gene expression or cell differential expression pattern by gene transfection? I think this would work if you delivered a gene for overexpression in proteins involved for RNA processing (e.g., splicing, ribosomal proteins, etc.), RNA transcription (e.g., TFs) or protein translation.</p>
|
<blockquote>
<p>Which are the factors that modify the overall gene differential
expression by introducing a vector for single-gene overexpression</p>
</blockquote>
<p>This is a very relevant question and the field of experimental biology needs to revisit the experimental strategies.</p>
<p>One of the explanations that you gave is correct- that the overexpressed protein might override the cellular processes. </p>
<p>Another problem that this practice gives rise to is incorrect inference. We are mostly interested in knowing what a protein does in a cell at general physiological conditions. If the stoichiometry is altered wrong results will definitely pop up.
For example in the case of a repressor/activator with multiple targets: At its general physiological concentration it may not really activate gene-X because of low affinity but in high concentrations it just may, and we end up concluding that gene-x is a target of repressor-1 by an OVEREXPRESSION experiment. </p>
<p>A synthetic biological approach is very good in studying small signaling/transcriptional <strong>modules</strong>. The entire module can be cloned and expressed in a cell under an inducible/constitutive promoter. This can be implemented in a heterologous cell also. A mathematical model generally helps in making certain predictions. </p>
<p>Other strategies which can be used instead of overexpression:</p>
<ol>
<li>using sensor constructs harboring a functional or non-functional
target sites instead of overexpressing the upstream protein</li>
<li>downregulation experiments</li>
</ol>
| 315
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cell signaling pathways
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What is the present tense verb form of apoptosis?
|
https://biology.stackexchange.com/questions/28497/what-is-the-present-tense-verb-form-of-apoptosis
|
<p>For example, if I want to say something along the lines of "this signaling pathway causes a cell to go through the process of apoptosis", but I want to shorten the phrase "go through the process of apoptosis" to one word, what would that word be? I've been saying "apoptose" so far but I'm not sure if that's correct.</p>
|
<p>According to Google Scholar, there were <a href="http://scholar.google.co.uk/scholar?q=apoptose&btnG=&hl=en&as_sdt=0%2C5" rel="noreferrer">~40000 hits</a> for "apoptose" and ~<a href="http://scholar.google.co.uk/scholar?q=%22undergo%20apoptosis%22&btnG=&hl=en&as_sdt=0%2C5" rel="noreferrer">120k hits</a> for "undergo apoptosis" in published literature, both of which had significant numbers of high-impact articles. </p>
<p>Therefore, it is clear that both expressions are sufficiently used in literature for either to be used. </p>
<p>Personally, I would say "undergo apoptosis" being both shorter than your original phrase and not risking coining new word forms.</p>
| 316
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cell signaling pathways
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Bioinformatics Prediction vs. Experimental Results: miRNA Regulation of AMPK/WNT Pathway in High-Glucose Conditions
|
https://biology.stackexchange.com/questions/116162/bioinformatics-prediction-vs-experimental-results-mirna-regulation-of-ampk-wnt
|
<p>I am investigating the role of a specific miRNA in high-glucose conditions, particularly its effects on cell proliferation, migration, and metabolism. However, I have encountered a significant discrepancy between bioinformatics predictions and experimental results, and I would appreciate insights on potential explanations and next steps.</p>
<h2>Background & Bioinformatics Predictions:</h2>
<p>Using TargetScan and miRDB, my bioinformatics analysis suggests that this miRNA targets PRKAA1 (AMPK) and WNT pathway components, implying that it may suppress autophagy and cell growth.</p>
<p>In a high-glucose environment, RNA-seq data shows that this miRNA is downregulated, which theoretically should:</p>
<ul>
<li>Release AMPK inhibition → promote autophagy.</li>
<li>Suppress WNT signaling → inhibit proliferation.</li>
</ul>
<h2>Experimental Results (Contradictory to Predictions):</h2>
<p>Overexpressing the miRNA in high-glucose conditions reduces cellular damage, contrary to the expectation that it should further suppress AMPK/WNT.</p>
<p>Inhibiting the miRNA in normal conditions leads to reduced proliferation and migration, contradicting the assumption that it suppresses growth.</p>
<p>This contradicts previous literature where AMPK activation is typically protective in high-glucose stress conditions.</p>
<p>Methods Used for Validation:</p>
<ul>
<li>qPCR: Confirmed miRNA expression changes in high-glucose conditions.</li>
<li>CCK-8 Assay: Measured proliferation.</li>
<li>Transwell Assay: Evaluated cell migration.</li>
</ul>
<h2>Key Questions:</h2>
<p>What could explain this discrepancy between bioinformatics predictions and experimental results?</p>
<p>Could this be due to context-dependent miRNA regulation, indirect compensatory mechanisms, or database biases?</p>
<p>Are there known cases where AMPK suppression is protective in a high-glucose environment rather than harmful?</p>
<p>Could WNT signaling be altered differently in this metabolic condition?</p>
<p>Should I continue pursuing this pathway if no prior literature supports this specific mechanism?</p>
<p>Is it common for miRNA bioinformatics predictions to significantly differ from in vitro results?</p>
<p>Should I validate additional targets or check for compensatory signaling pathways?</p>
<p>Any insights, relevant studies, or alternative explanations would be greatly appreciated!</p>
|
<p>The single key question in the poster’s shopping list can, I think, be reduced to</p>
<blockquote>
<p>“Why don’t the experimental results bear out the predictions?”</p>
</blockquote>
<p>The answer would seem self-evident:</p>
<blockquote>
<p>“Because the predictions, based as they are on sequence comparison, are incorrect.”</p>
</blockquote>
<p>It would seem rather naive to be surprised about this as the graveyard of biology is populated by the remains of many a beautiful hypothesis.</p>
<p>The purpose of this site is not to try to provide detailed advice on research programmes, but in similar circumstances I personally would accept the experimental results and either look for another target for the mir or give up the idea that its changes are the cause of the biological changes observed.</p>
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cell signaling pathways
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How does the cell regulate different metabolic pathways?
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https://biology.stackexchange.com/questions/23900/how-does-the-cell-regulate-different-metabolic-pathways
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<p>I heard somewhere that cells use different nucleosides bound to triphosphates e.g. ATP, GTP, CTP and other modified compounds: NADH, NADPH to distinguish between different metabolic pathways and so they regulate where they use up the energy. I heard that kinases play an important role in the regulation. Is there a connection (I guess there is if I check NAD<strong>P</strong>H)? Is this regulation mapped? I mean is there a simple map which contains the main processes and the energy carrier and regulatory compounds?</p>
<p>I am looking for something like this map (of receptor responses), but for metabolic regulation:</p>
<p><img src="https://i.sstatic.net/kFkt1.png" alt="signal transduction"></p>
<ul>
<li>Figure 1 - signal transduction - <a href="http://en.wikipedia.org/wiki/Signal_transduction" rel="nofollow noreferrer">wikipedia</a></li>
</ul>
<p>So it possibly contains mitochondria, O<sub>2</sub>, CO<sub>2</sub>, flows, ATP, NADPH, etc... I understand that different cell types can have different energy producer and consumer organelles and it is not possible to create something that is universally true, so I would be satisfied with a map of your favorite human cell type.</p>
|
<blockquote>
<p>I heard somewhere that cells ………………………… so they regulate where they use up the energy.</p>
</blockquote>
<p>Yes NADP/H is primarily employed in anabolic pathways such as fatty acid synthesis, while NAD/H is employed in catabolic pathways such as glycolysis.</p>
<p>I don't think there is a general rule for other "<em>energy-currency</em>" molecules (pyrimidine triphosphates are not used except in some rare cases such as glycogenesis).</p>
<blockquote>
<p>I mean is there a simple map which contains the main processes and the energy carrier and regulatory compounds?</p>
</blockquote>
<p>You can search for the term "<em>metabolic network</em>". It would be too huge so it is better to look at specific sub-networks. KEGG is a good site for finding metabolic networks. There are other representations like hive plots, to visualize very huge graphs.</p>
| 318
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cell signaling pathways
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Examples of natural graded transcriptional responses to extracellular ligands
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https://biology.stackexchange.com/questions/108509/examples-of-natural-graded-transcriptional-responses-to-extracellular-ligands
|
<p><a href="https://pubmed.ncbi.nlm.nih.gov/11571757/" rel="nofollow noreferrer">In this paper</a> (1) from 2001 the authors show that the mating pathway in budding yeast yields a graded transcriptional response to increasing concentrations of pheromone, and claim that:</p>
<blockquote>
<p>To our knowledge, this is the first example in eukaryotes that extracellular signals can be propagated in a simple graded fashion.</p>
</blockquote>
<p><a href="https://i.sstatic.net/Dd3h3.png" rel="nofollow noreferrer"><img src="https://i.sstatic.net/Dd3h3.png" alt="Dose-response" /></a></p>
<p><a href="https://pubmed.ncbi.nlm.nih.gov/16170311/" rel="nofollow noreferrer">Here's another paper</a> (2) from 2005 showing the same thing, but with single-cell fluorescence microscopy instead of flow cytometry.</p>
<p>Given that these papers are around 20 years old, I assume other pathways with similar signaling dynamics have since been found, but I'm struggling with finding good examples.</p>
<p>Is anyone aware of other pathways where the dose-response relationship between an extracellular stimulus and a transcriptional response is graded, as opposed to bistable? The organism is irrelevant, as long as it's a eukaryote, it's a natural (i.e. not synthetic nor mutated) pathway and the results have been shown by some single-cell analysis technique.</p>
<ol>
<li>Poritz et al. (2001) PMID: 11571757</li>
<li>Colman-Lerner et al. (2005) PMID: 16170311</li>
</ol>
| 319
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cell signaling pathways
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What are the short/long term effects of chloroquine on fundamental cell processes?
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https://biology.stackexchange.com/questions/92873/what-are-the-short-long-term-effects-of-chloroquine-on-fundamental-cell-processe
|
<p>Does chloroquine, which affects the endosomal membrane traffic pathway (by affecting the acid environment used for fundamental endosomal reactions), have short/long-term effects on cell growth/proliferation/signaling?</p>
|
<p>This <a href="https://www.ncbi.nlm.nih.gov/pmc/articles/PMC6103682/" rel="nofollow noreferrer">paper</a> provides many experiments about changes to the endosomal system and autophagy and should be a good starting point.<br>
Most of the treatments are done in the 5h to 24h range typical for cell culture studies. In theis relatively short time frame Chloroquine is well tolerated by cells. However, long term treatment will very likely be different.</p>
| 320
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cell signaling pathways
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How to find out what biological effects a molecule has, without having a specific mechanism or pathway in mind?
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https://biology.stackexchange.com/questions/113483/how-to-find-out-what-biological-effects-a-molecule-has-without-having-a-specifi
|
<p>I'm interested in finding what biological effects a (small) molecule has in a high-throughput and "low-assumption" way. I'm mainly interested in cell-based assays.</p>
<p>Background: There are easy/fast assays of high level phenotype effects eg cell proliferation or differentiation, but those wouldn't pick up lots of molecules as being active at all, unless they're at toxic levels. There are also a lot of assays which look at a specific pathway, eg using a Western blot with an antibody to a phosphorylated protein (phospho-Akt comes to mind). But those don't help if one doesn't already know the relevant pathways; they're more useful if screening molecules that target a known pathway, or confirming a known/suspected mechanism of action (and also most really aren't high throughput).</p>
<p>I'm interested in assays which can produce information on <em>everything</em> a molecule does when interacting with a cell, if possible in a single experiment.</p>
<p>Some things that come to mind:</p>
<ul>
<li>Gene expression profiling (eg using microarrays or RNAseq): gives information on the expression of everything; it is comprehensive and reasonably high throughput. The main limitation is that expression is usually a few steps downstream of what the molecule would interact with, unless it is literally interacting with transcription factors; so it would usually require tracing changes back up various signalling pathways</li>
<li>Proteomics (eg 2D-PAGE, shotgun proteomics): gives information on protein abundance, and potentially post-translational modifications. Seems hard; for 2D-PAGE for example typically a couple of thousands of spots can be identified (a fraction of the whole proteome), and the amount of protein in each measured to within ~20% accuracy (semi-quantitative). Possibly also needs a fairly large sample</li>
<li>Metabolomics (eg LCMS): gives information on all metabolites</li>
</ul>
<p>Are there any other types of assays that would give information across all/many genes/proteins/enzymes/signalling pathways?</p>
|
<p>Personally I think this is a poor question, because the answer is immediately obvious to anyone who was worked with current technologies for both proteins and nucleic acids, but as always, it is hard to prove a negative.</p>
<p>The short answer is NO, not even close to a single experiment or assay that can tell us EVERYTHING a single protein can do/interact with. If there were, we would be in a situation where almost all of cell biology could be done in a few thousand experiments.</p>
<p>Let's take a well known protein that people have been studying for quite some time now - the "Guardian of the Genome", <a href="https://www.ncbi.nlm.nih.gov/gene?cmd=search&term=7157" rel="nofollow noreferrer">Tumour Protein p53</a> (TP53, AKA p53). This is admittedly an integral protein to cellular function, so has a lot of interactions - but it serves to illustrate the point.</p>
<p>Now, let's head to the <a href="https://string-db.org/network/9606.ENSP00000269305" rel="nofollow noreferrer">STRING database for this protein</a>. STRING is a database that charts protein-protein interactions through a number of sources, mostly from the literature, but also through predicted interactions, with varying levels of confidence. Check out the legend and settings tabs for the page to see how those work.</p>
<p>The initial page for p53 should show you p53 and 10 other interacting proteins - these are the most highly significant interactions, with the greatest confidence in them being real. Now, click on the "+More" tab, and you should see 20 interactions - do this a few more times and watch that number and the complexity of interactions grow - it'll show interactions, not just with p53 but between other displayed proteins too. This is a fairly simplistic way to show the number of interactions a single protein has inside the cell. You should finally come up with a few hundred different proteins that have been found to interact with p53. Now, don't forget these are protein-protein interactions only.</p>
<p>p53 is also a DNA binding protein - it is predicted to interact with the genome at about 55,000 binding sites (see sect 2.1, para 2 in <a href="https://www.ncbi.nlm.nih.gov/pmc/articles/PMC6888028/" rel="nofollow noreferrer">this publication</a>) - What does it do at each of those sites? Short answer is - we don't know. Maybe upregulates something, maybe downregulates, maybe nothing, maybe something else.</p>
<p>There's also some evidence that <a href="https://www.ncbi.nlm.nih.gov/pmc/articles/PMC6314037/" rel="nofollow noreferrer">p53 interacts with various types of RNA molecules</a>, some of which are only relatively recently discovered, so assays for them are still fairly primitive. Again, what it does there, we don't know.</p>
<p>If you have ever worked with proteins and DNA, you will know that the methods of working with each are usually quite distinct - which also puts a technological barrier to your idea of a single unified assay. Of course, it is entirely possible to do DNA or RNA-protein interaction studies (e.g. ChIP), but to do these at the same time as protein-protein in a single assay - no chance.</p>
<p>There's also a second (third?) problem - sensitivity. Currently, with most technologies, you can have it one of two ways, but generally not both! You can either detect the rare interactions - the 1 in 1,000,000, by removing the common ones; or you can see the common ones but not the rare, because the common ones swamp the rare. An example of this is in RNA sequencing. In a cell, <a href="https://en.wikipedia.org/wiki/Ribosomal_RNA" rel="nofollow noreferrer">ribosomal RNAs</a> (rRNA) make up about 80% of cellular RNA, despite not coding for proteins (that I know of). So, if you want to look at a cellular coding RNA by next-gen sequencing, most of the time you will do a rRNA depletion or else 80% of the raw signals will be rRNA, which is a waste of your useful information achievable.</p>
<p>Edited to add:</p>
<p>As has been clarified in comments, OP is not asking for a single assay to do all, but rather types of assays that might be suitable to produce similar results - basically things like Next Gen sequencing, but for small-molecule (not necessarily protein-protein) interactions, which essentially turns this into a shopping question, which are generally not considered appropriate for this site as tools/techniques change quite frequently. It might be more appropriate on bioinformatics.</p>
<p>As always, the content I wrote above applies, particularly the bits about sensitivity and technological barriers.</p>
<p>As far as I know, no such assays exist for proteins, but the one that might come the closest is <a href="https://en.wikipedia.org/wiki/Protein_mass_spectrometry" rel="nofollow noreferrer">protein mass spectrometry</a> by MALDI-ToF with <em>de-novo</em> peptide sequencing. These are (in my very limited experience with them) by no means as trivial as setting up a Next Gen sequencing run, but I am sure the technology and associated bioinformatics tools will improve and mature over the next few years.</p>
| 321
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cell signaling pathways
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Importance of fate maps
|
https://biology.stackexchange.com/questions/53465/importance-of-fate-maps
|
<p>I did some normal google.book search and found the two importance of fate maps-</p>
<p><a href="https://books.google.co.in/books?id=2R_fCQAAQBAJ&pg=PA99&dq=fate%20map%20importance&hl=en&sa=X&ved=0ahUKEwiigb3vi7DQAhWLLo8KHdA9A9MQ6AEIOzAG#v=onepage&q=fate%20map%20importance&f=false" rel="nofollow noreferrer">1.</a> They helped establish the idea that communication between different parts of an embryo leads to the formation of new cell types and have thus helped discover different signalling pathways.</p>
<blockquote>
<p><a href="https://books.google.co.in/books?id=a1_mBwAAQBAJ&pg=PA87&dq=fate+map+importance&hl=en&sa=X&ved=0ahUKEwiigb3vi7DQAhWLLo8KHdA9A9MQ6AEIIDAB#v=onepage&q=fate%20map%20importance&f=false" rel="nofollow noreferrer">2.</a> They enable us to interpret experiments in which cells of the embryo are exposed to conditions that may alter their developmental fate.</p>
</blockquote>
<p>I really don't know where to look for the experiments that support these merits of fate maps. So is my question,<strong><em>what are some experiments that support these merits?</em></strong></p>
|
<p>A good place to begin is with the experiments of Hans Speymann and Hilde Mangold, done in the 1920s. They transplanted a piece of the dorsal lip in a newt gastrual to the ventral side of another newt gastrula with different pigmentation. Because of this transplant, a "secondary" embryo formed, on what would have been the belly, in addition to the standard embryo expected. (Some picture show this to be rather like a second head.) Use of differently pigmented embryos allowed the researchers to note that the transplanted dorsal lip material developed into a second notochord. But the interesting facet is the newly developing notochord <strong>induced</strong> the development of a new neural tube in the adjacent cells which were otherwise destined to become "belly stuff".</p>
<p>My appologies for not finding a great reference with diagrams, my source is the 7th edition of Biology, Campbell and Reece. Here's a very thorough reference, with diagrams, if you're up for it.
<a href="https://app.shoreline.edu/kwennstrom/spemannmangold.pdf" rel="nofollow noreferrer">https://app.shoreline.edu/kwennstrom/spemannmangold.pdf</a>
This work contributed to a Noble Prize for Speymann. The bottom line here is that tissues receive information from adjacent tissues that influence their developmental fate.
Later experiments showed that the inducing agent was chemical in nature</p>
| 322
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cell signaling pathways
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Role of the CD3 proteins and ζ chain
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https://biology.stackexchange.com/questions/109059/role-of-the-cd3-proteins-and-%ce%b6-chain
|
<p>Could someone please explain which of the following is correct and why?</p>
<blockquote>
<p>The role of the CD3 proteins and ζ chain on the surface of the cell is to:</p>
<p>a) transduce signals to the interior of the T cell</p>
<p>b) bind to antigen associated with MHC molecules</p>
<p>c) bind to MHC molecules</p>
<p>d) bind to CD4 or CD8 molecules</p>
<p>e) facilitate antigen processing of antigens that bind to the surface of T cells</p>
</blockquote>
<p>From what I have read I believe <strong>a)</strong> to be the correct answer but I am unsure if I have understood it correctly. This is based on this paragraph in the book "The Immune System 4th Edt":</p>
<blockquote>
<p>"By inducing this organization of molecules in the T-cell membrane, the interactions
of MHC ligands with T-cell receptors activate cytoplasmic protein
tyrosine kinases, which phosphorylate particular tyrosine residues in the cytoplasmic
tails of the CD3 cell-surface proteins and the associated ζ chain
(CD247), a purely cytoplasmic protein (see Figure 5.6, p. 118). The tyrosine residues
that become phosphorylated are part of short amino-acid sequence
motifs called immunoreceptor tyrosine-based activation motifs (ITAMs).
Enzymes and other signaling molecules bind to the phosphorylated tyrosine
residues and thus also become activated. In this way, the extracellular binding
of antigen to the T-cell receptor initiates pathways of intracellular signaling
that lead to alterations in gene expression and end with T-cell differentiation."</p>
</blockquote>
|
<p>A is most likely the answer they're looking for. However, CD3<span class="math-container">$\epsilon$</span> and either CD3<span class="math-container">$\delta$</span> or CD3<span class="math-container">$\gamma$</span> associate with CD4 or CD8 (depending on cell type) in the TCR complex, bringing in kinases like Lck to phosphorylate the ITAMs on the CD3 chains' intracellular domains, leading to further downstream signaling. So, CD3 <em>does</em> associate/"bind" CD4/CD8, making D correct as well, at least in my opinion.</p>
| 323
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cell signaling pathways
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If inhibiting S6 kinase decreases protein translation, then could inhibiting S6 kinase could possibly slow down long-term potentiation in neurons?
|
https://biology.stackexchange.com/questions/1369/if-inhibiting-s6-kinase-decreases-protein-translation-then-could-inhibiting-s6
|
<p>From <a href="http://en.wikipedia.org/wiki/P70S6_kinase" rel="nofollow">http://en.wikipedia.org/wiki/P70S6_kinase</a>...</p>
<blockquote>
<p>Phosphorylation of S6 induces protein synthesis at the ribosome.</p>
<p>P70S6 kinase is in a signaling pathway that includes mTOR (the
mammalian target of rapamycin). mTOR can be activated in distinct
ways, thereby activating p70S6K. For example, branched chain amino
acids such as leucine are sufficient to activate mTOR, resulting in an
increase in p70S6K phosphorylation (and thereby activating it). mTOR
is also in a pathway downstream of the kinase Akt. Akt is typically
activated upon stimulation of a cell with a growth factor (such as
IGF-1). Akt then activates mTOR (by inhibiting the Tsc complex),
leading to p70S6K activation.</p>
</blockquote>
<p>We also found a paper showing that downregulation of S6 kinase also results in a decrease in protein translation in yeast (but I'm waiting for Matt Kaeberlein to email me the slides from yesterday).</p>
|
<p>I can't rule it out, but it sounds a lot like trying to tune a piano with sledgehammer. </p>
<p>Neuronal LTP depends on protein translation, but so does absolutely everything else in the cell. Inhibiting protein synthesis at the ribosome will block the formation of all proteins, not just the ones responsible for LTP. Unless there's a link I don't know about between LTP and <em>total</em> levels of protein translation, you're really going to want to look into inhibiting the production of proteins specifically responsible for LTP and not protein synthesis in general.</p>
| 324
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cell signaling pathways
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Confusion related to a term probe-by-background interaction
|
https://biology.stackexchange.com/questions/7800/confusion-related-to-a-term-probe-by-background-interaction
|
<p>I was reading <a href="http://www.nature.com/tpj/journal/v12/n5/full/tpj201135a.html" rel="nofollow">a paper</a> related to bioinformatics where it uses the drug response on the cancer cells and the gene expression of the individual cells are studied to find any useful insights. Specially, using the gene expression of the cells a predictor of the drug response is created.</p>
<p>They have stated that just using the correlation between the gene expression and the drug response might not be a good predictor. But the genes interact through signaling pathways to drive a particular drug response.</p>
<p>What these guys have done is like used PCA on the gene expressions of the cancer cells to use the components which preserve the greatest variance.</p>
<p>Actually, I didn't get what they mean by probe-by-background interaction and how it is calculated.</p>
<p>Can anyone please explain. I googled for a while but didn't get it.</p>
<p>Here are some quotes from the paper where the term is used</p>
<blockquote>
<p>Towards this end, we have compared how well drug response can be
predicted by simple statistical models, which either directly relate
<strong>probe and background</strong> networks to drug response or consider
probe-by-background network interactions.</p>
<p>To generalize this approach, the term ‘probe’ could be
replaced by individual transcript expression levels measured through
other gene expression methods. Similarly, ‘background networks’ and
principal components are used interchangeably. Generally, ‘background
networks’ could be represented by any data reduction method that
summarizes the expression of a gene network. We demonstrate that
<strong>probe-by-background</strong> network interactions significantly enhance drug
response predictions, over and above the predictive power garnered
through utilizing individual probes and background networks alone.</p>
</blockquote>
|
<p>The "probe by background" interaction is the response of different probes as a function of background gene expression. For example, depending on which of the 6 backgrounds a probe is in, the drug response may go up or down. Probes as a function of background is probably easier to imagine than background as a function of probe (which is equally valid). For 39,115 probes and 6 background networks, there are 234,690 interactions.</p>
<p>In a technical sense, "probe by background" is the interaction term in the linear model that the authors fit for each drug. The details of the analysis are in the supplemental methods. The model they fit is</p>
<pre><code>Drug_Response ~ Probe + Probe * Background + Background
</code></pre>
<p>which translates to "Drug response is modeled by Probe, the interaction of Probe and Background, and Background". The middle term (with the <code>*</code>) is the interaction term. <a href="http://www.wilderdom.com/statistics/Interaction.html" rel="nofollow">Here is a page</a> that helps to explain how to understand significant interaction terms in linear models.</p>
| 325
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cell signaling pathways
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Advantage of GCPRs over RTKs or other receptor protein kinases
|
https://biology.stackexchange.com/questions/42818/advantage-of-gcprs-over-rtks-or-other-receptor-protein-kinases
|
<p>My book lists two important differences between GCPRs and receptor protein kinases:</p>
<ul>
<li>GCPRs do not directly activate a signal transduction pathway. It only does so indirectly, via a G protein. On the other hand, RTKs directly activate a signal transduction pathway, bypassing the mediation of a G protein. (Any G proteins involved with RTKs act as relay molecules in themselves and are part of the signal transduction pathway.)</li>
<li>GCPRs only activate one signal transduction pathway, while RTKs can activate many pathways.</li>
</ul>
<p>From what I can tell, receptor protein kinases are faster and more versatile than GPCRs. They do not require as many intermediate steps and can activate many pathways at once. However, GPCRs make up the most largest class of ligand receptors in human cells. This leads me to think there is a great advantage GPCRs have over RTKs that I’m not seeing.</p>
<p>If RTKs are more advantageous than GPCRs, why are there more GPCRs than RTKs?</p>
|
<p>I think GPCRs are evolutionary more older since tyrosine kinase signalling is relative recent evolved system. This could be one explanation for why their is more diversity in GPCRs</p>
<p>By the way GPCRs can actually signal g-protein independent, for example via B-arrestin. Also their are many types of g-proteins witch can all induce signaling via different signaling pathways. So GPCRS are actually quite versitile.</p>
<p>And then their is also a lot of crosstalk between GPCRs and RTKs </p>
| 326
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cell signaling pathways
|
How does the Lgr5 receptor contribute to maintaining stemness in the intestine?
|
https://biology.stackexchange.com/questions/72925/how-does-the-lgr5-receptor-contribute-to-maintaining-stemness-in-the-intestine
|
<p>I don't understand the connection between Lgr5 receptor and Wnt between Paneth cells and stem cells. And how does this link to the EphB-EprinB inhibition between transit amplifying cells and differentiated cells?</p>
<p>My book "The molecular biology of the cell" 6th edition (Garland Science) only states that the Lgr5 receptor in the stem cells makes them able to create a mini-gut (organoid), but I am asked to present factors that makes the gut able to totally renew itself and maintain its structure.</p>
<p>I have already answered the Delta-Notch lateral inhibition pathway and its connection to Wnt signaling from the paneth cells in determinating differentiation and maintaining in ability to divide.</p>
| 327
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cell signaling pathways
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What are the response frequencies of sensory neurons?
|
https://biology.stackexchange.com/questions/67829/what-are-the-response-frequencies-of-sensory-neurons
|
<p>Both visual and auditory stimuli are sent to the brain via ganglion cells (<a href="https://en.wikipedia.org/wiki/Retinal_ganglion_cell" rel="nofollow noreferrer">retinal</a> resp. <a href="https://en.wikipedia.org/wiki/Spiral_ganglion" rel="nofollow noreferrer">spiral</a>). Both are the first cells along their resp. pathways that produce action potentials.</p>
<p>My question concerns typical frequencies of action potentials sent along the axons of the visual vs. auditory ganglion cells as a reaction to a "typical stimulus", i.e. a medium long, medium strong signal of some fixed frequency (e.g. light: red, sound: 440Hz) against a white resp. silent background.</p>
<p>Are these frequencies of comparable range, or does one type of ganglion cell (retinal vs. spiral) fire with a significantly higher or lower rate than the other?</p>
<p>(The question would not make sense, if the physical frequencies of light and sound - which trigger the receptor cells - would be coded by frequencies of action potentials. But I assume that this is not the case, is it?)</p>
|
<p>The auditory brainstem shows "phase-locking" typically up to 1-3Khz at most; 3000Hz is an incredibly high firing rate for a single neuron, but this phase-locking is achieved <strong>not by individual cells firing in-phase with an auditory stimulus</strong>, but rather with a <strong>population of cells</strong> that tend to fire in-phase, such that if you average across the population you get a phase-locked population volley.</p>
<p>In some cases, in some animals, this phase locking can even get to the higher frequencies (<a href="http://www.jneurosci.org/content/jneuro/17/9/3312.full.pdf" rel="nofollow noreferrer">see here</a> for example).</p>
<p>However, this phase locking seems primarily important for <em>sound localization</em> via <a href="https://en.wikipedia.org/wiki/Interaural_time_difference" rel="nofollow noreferrer">interaural time differences</a>. Frequency itself is encoded by which population of hair cells is activated, according to the properties of the <a href="https://en.wikipedia.org/wiki/Basilar_membrane" rel="nofollow noreferrer">basilar membrane</a>. Firing <em>rates</em> of individual spiral ganglion cells <a href="https://link.springer.com/content/pdf/10.1007%2FBF00694467.pdf" rel="nofollow noreferrer">are only faster than 100 Hz at very high stimulus intensities</a>.</p>
<p>Similar to the spiral ganglion cells, retinal ganglion cells primary encode intensity information in their firing rates.</p>
<p>However, in both cases, it's important to recognize how crucial <em>adaptation</em> is in sensory systems. RGCs in particular fire primarily to <em>transients</em>, so it is typical to use light flashes, drifting gratings, or other dynamic stimuli. The response to a "medium long, medium strong signal of some fixed (wavelength)" is going to be brief, followed by silence, not a constant response like you imply.</p>
| 328
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cell signaling pathways
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Is Signal Transduction Unidirectional from the Stimuli to the Final Receptor?
|
https://biology.stackexchange.com/questions/106689/is-signal-transduction-unidirectional-from-the-stimuli-to-the-final-receptor
|
<p>I wonder if signal transduction in biological systems including visual, olfactory, tactile or any other biological system, is unidirectional. Suppose that <span class="math-container">$X_i$</span> is the <span class="math-container">$ith$</span> cell in the signal transduction pathway and <span class="math-container">$X_{i+1}$</span> is the next cell down the pathway. To be more clear let me rephrase my question. Suppose that,</p>
<p><span class="math-container">$X_i \rightarrow X_{i+1}$</span></p>
<p>means that the signal is sent from <span class="math-container">$X_i$</span> to <span class="math-container">$X_{i+1}$</span> and,</p>
<p><span class="math-container">$X_i \rightleftharpoons X_{i+1}$</span></p>
<p>means that the signal is being sent both from <span class="math-container">$X_i$</span> to <span class="math-container">$X_{i+1}$</span> and vice versa but the signal amplitude in the <span class="math-container">$X_i$</span> to <span class="math-container">$X_{i+1}$</span> direction is larger so on average we can say that effectively the signal is being transmitted from <span class="math-container">$X_i$</span> to <span class="math-container">$X_{i+1}$</span>. Which one of these cases is true for biological systems? Or can both occur depending on the system?</p>
<p>The question might not be a very good one because I don't have any background in biology. I would appreciate any answer or guidance. Thanks in advance.</p>
|
<p>No.</p>
<p>I'll just use one example, the retina, for a proof by contradiction.</p>
<p>In the retina, the principle "forward"/unidirectional pathway would be from photoceptor cells (rods/cones) to bipolar cells (BC) to retinal ganglion cells (RGCs/GCs). RGCs are the projection neurons whose axons make up the optic nerve from eye to the rest of the brain.</p>
<p>However, there are other cell types in the retina as well that convey information horizontally and in reciprocal/feedback directions. Here are some examples depicted in a review paper:</p>
<p><em>(image from Demb & Singer 2015)</em>
<a href="https://i.sstatic.net/KV1nC.png" rel="nofollow noreferrer"><img src="https://i.sstatic.net/KV1nC.png" alt="Retinal pathways" /></a></p>
<p>In (a), on the left, you see connectivity between a bipolar cell (BC) and an amacrine cell (AC), a type of interneuron in the retina. The AC is excited by the BC but also inhibits the BC; this is not unidirectional. In the middle, you see a horizontal connection between two BCs through an AC. On the right, you see another horizontal connection among BCs, in this case BCs of different types, through an AC.</p>
<p>Horizontal and feedback connectivity patterns are ubiquitous in the mammalian nervous system; you'll find them everywhere you look, so it's not reasonable to expect signal transduction to be unidirectional.</p>
<p>I'll add that it is not straightforward to determine the "strength" of signals in the nervous system, especially when inhibition is involved. For only excitatory connections, you might measure the excitatory potential (voltage change) at the cell body as a measure of "strength", but this is complicated because this strength depends on lots of other things: you can have nonlinear interactions in dendrites such that multiple signals sum linearly, sublinearly, or supralinearly; the rate of presynaptic activation matters, so even a "strong" synapse may not be particularly influential if the presynaptic neuron rarely fires, etc.</p>
<p>For inhibition, it's even more complicated, because many inhibitory potentials involve <a href="https://en.wikipedia.org/wiki/Shunting_inhibition" rel="nofollow noreferrer">shunting</a> and may not affect the membrane potential at the soma at all unless they are paired with an excitatory input. It's possible to have an inhibitory input so strong that a cell can effectively never fire, and yet the effect of that inhibitory input alone has no detectable influence on the cell's voltage, it merely provides a shunt that fixes the membrane potential at that voltage.</p>
<p><a href="https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5749398/" rel="nofollow noreferrer">Demb, J. B., & Singer, J. H. (2015). Functional circuitry of the retina. Annual review of vision science, 1, 263-289.</a></p>
| 329
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cell signaling pathways
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Can GFP reporting be used to track localization of peptides in the ER, Golgi, and plasma membrane?
|
https://biology.stackexchange.com/questions/100194/can-gfp-reporting-be-used-to-track-localization-of-peptides-in-the-er-golgi-an
|
<p>Suppose I want to study the trafficking of a peptide throughout the ER, Golgi, and plasma membrane. An idea I had was labeling a secreted or plasma membrane integral protein with GFP and using time-series live cell microscopy to track it through the secretory pathway.</p>
<p>However, I am concerned that by the time everything is set up, the cell will be at steady state, which means GFP signal would be present throughout the secretory pathway, preventing me from doing this tracking.</p>
<p>How might I overcome this, without changing the fundamental technology (GFP reporting)?</p>
|
<p>Historically speaking, the use of VSVG (Vesicular Stomatitis Virus G protein) labeled with GFP was indeed a practical method to define the secretory pathway. (@tyersome comment)</p>
<blockquote>
<p>However, I am concerned that by the time everything is set up, the cell will be at steady state, which means GFP signal would be present throughout the secretory pathway, preventing me from doing this tracking.</p>
</blockquote>
<p><em>Sec</em> mutations in <em>S.cerevisiae</em>: Many biochemical pathways are initially identified owing to a particular mutation in the synthesis process. Recall <a href="https://en.wikipedia.org/wiki/One_gene%E2%80%93one_enzyme_hypothesis" rel="nofollow noreferrer">Beadle and Tatum experiment</a> and the logic they used to clarify the existence of 3 enzymes used in arginine metabolism.
<a href="https://i.sstatic.net/GPKD0.png" rel="nofollow noreferrer"><img src="https://i.sstatic.net/GPKD0.png" alt="" /></a>
(Image taken From <a href="https://www.chegg.com/homework-help/questions-and-answers/following-experiments-beadle-tatum-searching-mutations-disrupt-arginine-biosynthesis-pathw-q41797559" rel="nofollow noreferrer">https://www.chegg.com/homework-help/questions-and-answers/following-experiments-beadle-tatum-searching-mutations-disrupt-arginine-biosynthesis-pathw-q41797559</a>)</p>
<p>A similar approach was taken back then to identify the distinct parts of the secretory pathway. Five different mutations in S.cerevisiae were identified named from A to E. Culturing all A to E mutated strains plus VSVG-GFP assay would clearly demonstrate the entire route.
<a href="https://i.sstatic.net/yCzqJ.jpg" rel="nofollow noreferrer"><img src="https://i.sstatic.net/yCzqJ.jpg" alt="" /></a>
(Image taken from <a href="https://slideplayer.com/slide/4382105/" rel="nofollow noreferrer">https://slideplayer.com/slide/4382105/</a>)</p>
<p>For example, even if the mutated class B, reaches the steady/equilibrated state, transport vesicles, Golgi, and the rest will not show the fluorescence activity (characteristic of VSVG-GFP) because all of the proteins have accumulated in the previous step(i.e budding from Rough ER)</p>
<p><strong>Further Readings :</strong></p>
<p><a href="https://pubmed.ncbi.nlm.nih.gov/2188733/" rel="nofollow noreferrer">https://pubmed.ncbi.nlm.nih.gov/2188733/</a></p>
<p><a href="https://www.sciencedirect.com/science/article/abs/pii/0092867481900647" rel="nofollow noreferrer">https://www.sciencedirect.com/science/article/abs/pii/0092867481900647</a></p>
<p>MCB Lodish et al 8th edition chapter 14 section 14.1</p>
<p><a href="https://www.nature.com/articles/35073068" rel="nofollow noreferrer">https://www.nature.com/articles/35073068</a></p>
| 330
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cell signaling pathways
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How do membrane proteins find their target locations?
|
https://biology.stackexchange.com/questions/68249/how-do-membrane-proteins-find-their-target-locations
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<p>The question might be asked for any kind of "bound" proteins, but I'd like to restrict it to <a href="https://en.wikipedia.org/wiki/Membrane_protein" rel="noreferrer">membrane proteins</a>.</p>
<p>Assuming membrane proteins (or their main parts) don't (or aren't) build <em>in situ</em> but at some distance of the membrane, I wonder by which mechanisms they travel from their generation site to their final destination inside the membrane (inner or outer).</p>
<p>Proteins that are distributed roughly evenly (or randomly) over the membrane don't pose a big conceptual problem: they could have gone their just <em>by diffusion</em>, possibly from many generation sites, distributed roughly evenly (or randomly) inside the cell.</p>
<p>But what about uneven distributions, where some proteins are more densely and non-randomly packed (significant and functional) at some sites of the membrane than at others, e.g. </p>
<ul>
<li><p>receptors at <a href="https://en.wikipedia.org/wiki/Postsynaptic_density" rel="noreferrer">postsynaptic densities</a></p></li>
<li><p>Na<sup>+</sup> channels at action potential initiation sites like the <a href="https://en.wikipedia.org/wiki/Axon_hillock" rel="noreferrer">axon hillock</a> or the <a href="https://en.wikipedia.org/wiki/Axon#Initial_segment" rel="noreferrer">axon initial segment</a></p></li>
<li><p>Na<sup>+</sup> channels at <a href="https://en.wikipedia.org/wiki/Node_of_Ranvier" rel="noreferrer">nodes of Ranvier</a>?</p></li>
</ul>
<p>By which mechanisms (forces, signals or structures) are these proteins led to
their targets?</p>
<p>Maybe it depends, and there are different mechanisms. These I came up with (by contemplating <a href="https://en.wikipedia.org/wiki/First_principle" rel="noreferrer">first principles</a>):</p>
<ul>
<li><p>uneven distribution of generation sites inside the cell (due to what?)</p></li>
<li><p>uneven distribution of origins of attracting signals inside the membrane (due to what?)</p></li>
<li><p>some "self-attracting" forces or signals (leading to accumulation by <a href="https://en.wikipedia.org/wiki/Positive_feedback" rel="noreferrer">positive feedback</a>)</p></li>
<li><p><a href="https://en.wikipedia.org/wiki/Microtubule" rel="noreferrer">microtubules</a></p></li>
</ul>
<p>Which mechanism is — possibly — predominant?</p>
<hr>
<p>Related questions:</p>
<ul>
<li><p><a href="https://biology.stackexchange.com/questions/67679/life-cycle-of-proteins">Life cycle of proteins</a></p></li>
<li><p><a href="https://biology.stackexchange.com/questions/67865/pathways-of-ligand-gated-ion-channels">Pathways of ligand-gated ion channels</a></p></li>
<li><p><a href="https://biology.stackexchange.com/questions/66489/distribution-of-synapses-of-ca1-neurons">Distribution of synapses of CA1 neurons</a></p></li>
<li><p><a href="https://biology.stackexchange.com/questions/66121/distribution-of-dendritic-spike-generating-ion-channels-on-the-dendritic-tree">Distribution of dendritic spike generating ion channels on the dendritic tree</a></p></li>
<li><p><a href="https://biology.stackexchange.com/questions/65550/visual-maps-of-the-neuronal-membrane">Visual maps of the neuronal membrane</a></p></li>
<li><p><a href="https://biology.stackexchange.com/questions/65219/distribution-of-sodium-potassium-pumps">Distribution of sodium–potassium pumps</a></p></li>
</ul>
|
<p>This is a great question. A comprehensive answer would be beyond the scope of an answer on a forum like this. I will summarize the best I can here, but if you are really interested in this you should look at some of the work by <a href="https://mcb.berkeley.edu/labs/schekman/pages/publications.html" rel="noreferrer">Randy Schekman</a> and <a href="http://rapoport.hms.harvard.edu/" rel="noreferrer">Tom Rapoport</a>, who have done a lot of pioneering work in this field and have papers from more than two decades ago on their lab websites. I'll talk about membrane proteins generally, but I'm not sure what the state of the field is for Na+ channels specifically, so I can't comment too much on that particular case. Many of the details of the processes I will mention are still areas of active research, so I will try to stick mainly to what has been well-characterized (to the best of my knowledge). </p>
<p>To restate the problem, proteins are generally synthesized in the lumen of the <a href="https://en.wikipedia.org/wiki/Endoplasmic_reticulum" rel="noreferrer">endoplasmic reticulum</a>, which is an aqueous environment similar to the cytosol in many (but not all) ways. However, membrane proteins, which are not stable in aqueous environments, must:</p>
<p>1) Find a way from the ER lumen into a membrane.<br>
2) Get from the ER into the correct membrane so they may perform their cellular function. </p>
<p>We will start with step 1, but the key to both is a critical but often underappreciated aspect of protein biology called the <a href="https://en.wikipedia.org/wiki/Signal_peptide" rel="noreferrer">signal peptide</a>. The signal peptide is simply an N-terminal sequence of amino acids that precedes what we would normally think of as the beginning of a mature protein. It is relatively short, usually only ~30 amino acids in length. It is cleaved off the mature protein by a protease once the protein is folded and in the membrane. Until that time, the signal peptide serves as a molecular marker that indicates where the nascent protein should be heading and how it should be handled. Not surprisingly, there are many different signal peptides that serve multiple functions, and they are not only used for membrane proteins. </p>
<p>So let's say we are in the ER lumen, and have some mRNA coding for a membrane protein that is destined for the plasma membrane. The first amino acids that will emerge from the ribosome is the signal sequence, in this case a specific signal sequence indicating that this is to be a plasma membrane protein. Once the signal sequence emerges from the ribosome, it is recognized by a ribonucleoprotein complex (that is, a complex of RNA and protein) called the <a href="https://en.wikipedia.org/wiki/Signal_recognition_particle" rel="noreferrer">signal recognition particle</a>. Once the SRP binds to the signal peptide, translation is halted, and the whole complex moves to the ER membrane, where it forms a complex with another large protein complex called the <a href="https://en.wikipedia.org/wiki/Translocon" rel="noreferrer">translocon</a>. I can't go into the intricacies of these complexes and their functions in this answer alone, but the simple description is that the translocon contains an ATPase that can insert the membrane protein into the ER membrane as it is translated, with the correct orientation. The hydrolysis of ATP provides energy to move the emerging polypeptide chain into the hydrophobic membrane, where chaperones help it fold. This process is in part driven by the recognition of hydrophobic transmembrane regions of the proteins by the translocon. It can also move soluble, cytosolic proteins across the membrane through a similar mechanism. </p>
<p>Now that the protein has been translocated, a peptidase will cleave the signal sequence off the protein. From here, sorting signals will take over. Generally, these are <a href="https://en.wikipedia.org/wiki/Protein_targeting" rel="noreferrer">simple sequence motifs in the first transmembrane domain that act similarly to a signal sequence, but they are not cleaved</a>. However, <a href="http://www.annualreviews.org/doi/abs/10.1146/annurev.biochem.72.121801.161800?journalCode=biochem" rel="noreferrer">sorting signals can be stretches of peptide throughout the protein too</a>, in some cases. </p>
<p>These sequence motifs will be recognized by cell trafficking machinery. Without going into too much detail, the proteins will be gathered into vesicles, and transported to other organelles. Usually, the first stop for proteins is the <a href="https://en.wikipedia.org/wiki/Golgi_apparatus" rel="noreferrer">golgi apparatus</a>, which is typically where many post-translational modifications, such as glycosylation, take place. I am a biochemist and not a cell biologist, so I am not the most qualified to go into the details of subcellular trafficking. Suffice it to say, once the protein is finished being processed in the golgi, it will be trafficked into other organelles, such as the plasma membrane using vesicle transport as before. From my understanding, the protein will be sorted into the proper vesicles based on its sorting signals, as well as other markers (in some cases, certain post-translational modifications on certain proteins can influence its trafficking). The vesicles recognize the proper destination membrane in part by the lipid composition of that membrane. For example, <a href="https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3340507/" rel="noreferrer">phosphoinositides have extensive influence on membrane trafficking</a>, and many membranes can be differentiated by their phosphoinositide signature. </p>
<p>Anyway, that is a very broad overview of the answer to your question. I'm sorry I can't comment too much on the intricacies of cellular trafficking, I don't quite have the expertise to go through that literature quickly enough to answer your question in a reasonable time frame. I hope this is helpful in pointing you in the right direction, and good luck!</p>
| 331
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cell signaling pathways
|
What is basis of multifunctionality of "master glands" in the endocrine system?
|
https://biology.stackexchange.com/questions/35745/what-is-basis-of-multifunctionality-of-master-glands-in-the-endocrine-system
|
<p>I have just started reading about the endocrine system and I am having some difficulty understanding the basis of distribution of glands and associated hormones.</p>
<p>I am using multifuntionality to describe the amount of relatively independent input and output that are shared through a same organ or proximal space. Many of these organs have a heterogenous mixture of endocrine cells sharing the same glandular space.</p>
<p>There appear to be some glands that are considered "master glands" such that they take a variety of relatively independent input, apply some control logic via a diverse set of endocrine cells and tissue and secrete a diverse hormonal output. The pituitary and hypothalamus are the best examples, however the adrenal cortex and the pancreas also fit this definition to a certain extent. Many times the diversity of hormones and cells within an organ have no obvious connection (eg thyroid). Some glands appear more specialized with a relatively straightforward endocrine axis of input and output (eg testes).</p>
<p>Some of this multifunctionality seems unnecessary, in that insulin secretion could occur anywhere and be effective in the manner of an insulin pump, or contain risks due to excessive centralization as in a pituitary tumor or renal artery stenosis, so there must be some advantages I do not understand.</p>
<p>My textbook (Headley, Levine, 6th) seems to minimally cover multiplexing, signal cross-talk and competition between diverse endocrine cells sharing an organ. Its focus on discrete pathways and axes suggests these are evolutionarily unfavorable.</p>
<p>Are there synergistic advantages in shared endocrine infrastructure such as vascularity or metabolic pathways?</p>
<p>Is there some overlying logic that can be used to explain this distribution of glands and hormones regarding multifuntionality-specialization, or is it necessary to view the distribution of vertebrate endocrine system as the outcome of evolutionary contingencies?</p>
<p>Thanks</p>
|
<p>It would seem to me that in the examples that you have listed that proximity to necessary input is the overriding logic behind gland geography. Take the hypothalamus as the first example. This gland receives <a href="http://medschool.slu.edu/anatomy/guide_ms/chapter_13.html" rel="nofollow noreferrer">input from diverse regions of the brain</a> from the amygdala and hippocampus to the retina and brainstem. The brain can tweak levels of various releasing hormones entering the <a href="http://arbl.cvmbs.colostate.edu/hbooks/pathphys/endocrine/hypopit/anatomy.html" rel="nofollow noreferrer">hypothalamic-hypophyseal portal veins</a> in order to produce a response that is appropriate to the situation perceived by the brain. It makes sense to have all of these co-located because they are all receiving input from the brain. Similarly, it makes sense to have all of cells that are uptaking the releasing hormones close-by to protect these vital peptides from degradation in the systemic circulation. Their collectively unique circumstance makes sharing a specialized vasculature a logical choice.</p>
<p>The pancreas is a good example of the importance of cross-talk within some <a href="http://hpcreu.umbc.edu/summer2014/projects/team3/" rel="nofollow noreferrer">endocrine tissue</a>. Within the islet, alpha, beta, and delta cells are feeding back on each other to get the balance of insulin and glucagon just right. To whit, glucagon secretion provides a paracrine signal for insulin secretion. This makes sense because the body knows that once glucagon is released the blood sugar will soon rise. Ergo, it makes sense to trigger some insulin secretion as well in preparation. Their co-location makes that possible.</p>
<p>In short, I would say the overriding logic to the placement of each of these glands is their proximity to the necessary inputs and outputs that each of these glands interacts with.</p>
| 332
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cell signaling pathways
|
Do different chiral centers on ligands cause different confirmational changes and effects in their target proteins?
|
https://biology.stackexchange.com/questions/62557/do-different-chiral-centers-on-ligands-cause-different-confirmational-changes-an
|
<p>Say pathogenic bacteriaA makes toxinA, which had D-amino acids instead of L-amino aids, does this difference in chirality <strong>cause</strong> a different conformational change in the receptor or enzyme, thus leading to either deactivation of the enzyme or signal transduction pathway or activation of a different pathway?</p>
<p>I understand what chirality is in the concept of organic chemistry — rotating plane polarized light, ingold-prelog system etc; however I never leaned what structural feature of chiral molecules changes the way they react inside a cell.</p>
<p>I do NOT understand HOW changes in chirality can be associated with cellular toxicity.</p>
<p>links:
<a href="https://www.ncbi.nlm.nih.gov/pubmed/24752840" rel="nofollow noreferrer">https://www.ncbi.nlm.nih.gov/pubmed/24752840</a></p>
<p><a href="http://www.jomb.org/uploadfile/2014/0113/20140113053743849.pdf" rel="nofollow noreferrer">http://www.jomb.org/uploadfile/2014/0113/20140113053743849.pdf</a></p>
<p><a href="https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3960212/" rel="nofollow noreferrer">https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3960212/</a></p>
|
<p>There a few things you have to take care of with chirality in biological systems:</p>
<p>First note, that for amino acids we usually use L and D instead of r and s nomencalture. </p>
<p>In chemistry the difference between chiral enantionemeres is usually really just a difference in light polarisation like you described, otherwise they can be thought of as identical, since they have identical chemical properties. This way of thinking does <em>not</em> apply to biochemistry - the moment enzymes (large molecules with very defined 3D structures) are involved the structural difference between the two forms becomes very important for each and every biological reaction.</p>
<p>In the case of <em>single</em> amino acids this means that D amino acids are toxic to eukaryotic cells, because their similarity to the D form allows them to interact with some of the enzymes that usually bind it, but the binding is not a perfect match so the enzymes get inhibited, which makes the D form toxic. (This is already described in the second paper you linked:)</p>
<blockquote>
<p>In animal studies, administration of D-amino acids to rats and chicks resulted in growth inhibition [18]. In addition, serious damages such as suppression of the synthesis of glutamate oxaloacetate transaminase, glutamic pyruvic transaminase, and lactate dehydrogenase resulted from D-amino acids accumulation in animal tissues [18].</p>
</blockquote>
<p>Now in your question you are asking about the difference between a toxin made from the 'wrong' amino acid. The thing is, this change of the protein will most likely mean, that its not a toxin anymore. A protein toxin is toxic because it can interact with other proteins, based on its 3D structure. This structure is based on the chemical and structural properties of the amino acids building it. If you change the underlying structure of every amino acid it will fold differently and you will get completely different protein - and <em>not</em> a mirrored version of the original protein/toxin.</p>
| 333
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