The Interactive Fly

Zygotically transcribed genes

Proneural and Neurogenic Genes

What is the Notch pathway?

Developmental biology of the Notch pathway

The achaete-scute complex

Embryonic central nervous system lineages

Aging neural progenitors lose competence to respond to mitogenic Notch signaling

Dampening the signals transduced through Hedgehog via microRNA miR-7 facilitates Notch-induced tumourigenesis

The bantam microRNA acts through Numb to exert cell growth control and feedback regulation of Notch in tumor-forming stem cells in the Drosophila brain

Specification and spatial arrangement of cells in the germline stem cell niche of the Drosophila ovary depend on the Maf transcription factor Traffic jam

Neurogenic genes functioning in the Notch pathway

Proneural genes of the achaete-scute complex

Additional proneural genes

What is the Notch pathway?

To understand the Notch pathway one has first to know the function of receptors and ligands. A receptor is a protein molecule located partially outside the cell and partially inside. The receptor interacts with outside signals and transmits them to the inside. Ligands are extracellular proteins or other molecules that interact with a receptor to cause it to transmit a signal to the inside of the cell. Notch is a receptor that receives signals from its ligands and transmits these signals to the inside of the cell. The Notch pathway consists of Notch and its ligands, as well as intracellular proteins that transmit the Notch signal to the nucleus. Included in the pathway are transcription factors, the proteins that bear the effector function of the pathway.

The effect of Notch signaling is the phenomenon of lateral inhibition, the singling out of one cell from a cell cluster for a given fate (neurogenesis for example), and the inhibition of those cells not elected to differentiate. Lateral inhibition occurs repeatedly in Drosophila development. Central to this process is the interaction of Notch receptor with a group of ligands including Delta, Scabrous and Serrate. Some of these ligands float free around the outside of the cell and others are bound to the surface of cells.

The interaction of Notch with its ligand triggers a chain of intracellular events resulting in lateral inhibition, the prevention of a specified fate committment on the part of most of the cells of a cluster. The first link in the chain of events is a reduction of the affinity of Suppressor of Hairless protein for the cytoplasmic tail of the Notch receptor. Following this, Su(H) is free to enter the nucleus where it assumes its role as a transcription factor. Su(H) mediates transcriptional activation of Enhancer of split complex of genes. This complex will then inhibit the proneural genes achaete, scute, lethal of scute and asense. The transcriptional inhibitory capacity of Enhancer of split complex genes results in lateral inhibition.

The process of lateral inhibition often involves cell migration. In addition to the signaling capacity of Notch receptor, a second important function of Notch and its ligand Delta is that of cell adhesion. Delta is bound to the surface of cells. The Delta-Notch interaction results in an intimate adhesive contact between fated cell, which often migrates, and inhibited cell, bearing Notch, which remains in its germ layer or cluster of origin. Thus the interaction of Notch with its ligand causes the lateral inhibition of the cell bearing Notch, and the migration and triggering of a differentiated state for the cell bearing the Notch ligand. The ligand most often turns out to be Delta.

How can the Notch pathway components be called neurogenic if the function of Notch is to repress the adoption of a differentiated state in the cell that carries Notch protein? There are two answers to this question. The first is that the term neurogenic is a misnomer, arising because the phenotype of Notch pathway mutants manifests the result of an over production of neurons. The pathway is named neurogenic because of this mutant phenotype. The second answer lies in considering the principle ligand of Notch and its function. Delta is not secreted but is cell bound. The Delta-Notch interaction serves a cell adhesive function between ligand and receptor bearing cells. While the receptor bearing cell is inhibited in assuming a differentiated state, the ligand bearing cell is free to do so. In neurogenesis this cell delaminates, and then migrates out of the epithelial cell layer. The ligand bearing cell assumes the differentiated state of neuroblast and physically resides in the developing nervous system. Thus Notch is neurogenic with respect to the cell that bears the Notch ligand.

Developmental biology of the Notch pathway

The Notch ligand receptor system functions in all three germ layers during embryonic development as well as in the germ cell line and in imaginal discs during the larval and pupal stages. The outline below lists developmental processes in which the Notch pathway functions. All are affected by Notch and where tested, all were found to be affected by Delta (Hartenstein, 1992).

The achaete-scute complex

The achaete-scute complex (AS-C) contains four related genes, achaete, scute, lethal of scute and asense. These are spread over 90 kilobases. The proteins coded for by these genes serve as transcription factors. Each has a DNA binding domain, termed a basic helix-loop-helix domain, and three of the four proteins (the exception is Asense) have a C-terminal acidic domain.

achaete and scute genes are more than 25 kb apart. lethal of scute is more than 12 kb from scute. asense is another 45 kb removed from lethal of scute.

These genes are termed proneural, since they promote neuroblast differentiation and are thus essential for the differentiation of the central and peripheral nervous system and the brain. Other functions are also served by these genes: regulation of sex determination (scute); participation in specification of muscle progenitors (lethal of scute), and regulation of sequential fates in Malpighian tubule development (achaete).

How are achaete-scute complex genes activated and inactivated? Genes like hairy, extramachrochaete and pannier help to establish patterns of achaete expression by acting as repressors. The first two genes received their names because of their interaction with achaete: mutation in either produces extra sensory elements such as hairs and machrochaete. As achaete is responsible for activating the sensory cell fate, Hairy and EMC act to restrict achaete transcription to a limited region where sensory elements will form.

NK2/Ventral nervous system defective regulates the expression of achaete and scute to the medial column of the ventral nervous system (Skeath, 1994). Specifically, first NK2 is essential for AS-C gene expression in the medial column of every other neuroblast row through regulatory elements located 3' to achaete, and second, through a 5' regulatory region, NK2 functions to increase or maintain proneural gene expression within the proneural cluster that normally gives rise to the neuroblast (Skeath, 1994).

The above information about the three repressors and NK2 provides only a nearsighted view of activation and repression by the AS-C. There is a more global answer to the question of how AS-C genes are activated. AS-C genes are activated in proneural clusters in the ventral portion of the fly. The anterior-posterior arrangement of neuroepithelial clusters is intimately connected to the process of segmentation. The proneural genes are expressed in each of the 14 segments. Pair rule genes define these segments, and therefore the placement of proneural AS-C clusters. In fact embryos that are mutant in each of the pair rule genes show an alteration of early cluster pattern.

It is not just the presence or absence of a cluster that is controlled by pair rule genes, but the arrangement of neuroblasts within each cluster, and consequently the arrangement of cells expressing AS-C genes. Each cluster is subdivided into subdomains, and each subdomain has its particular arrangement of neuroblasts. Segment polarity genes act here too, especially gooseberry and naked. They help maintain the AP borders of the cells within each cluster (Skeath, 1992).

See Chris Doe's Hyper-Neuroblast map site for information on the origin and lineage of specific neuroblasts in the CNS and for information about the expression of specific genes regulating neuroblast origin.

achaete and scute are controlled by combinations of axis-patterning genes through a common enhancer or locus control region. The pair rule genes act through this region, situated in the intervening chromosomal segment between achaete and scute. Thus this region is under the initial control of products of the pair rule genes and is later maintained by selected segment polarity genes.

Consider the amazing patterning present in insect epidermis. Flies have hairs that appear as reproducable patterns from one individual to another. These patterns are species specific and therefore distinguish one species from another. How are these patterns made?

Further analysis of the 90 kD region between achaete and asense has defined nine site-specific enhancer-like elements that function to regulate achaete and scute expression in proneural clusters of imaginal discs. Ultimately it is the capacity of site specific enhancers to kindle AS-C transcription at very localized regions in the proneural clusters of imaginal discs that is responsible for the patterning of sensory elements seen in adults.

Each of these enhancers, regulated by a specific set of transcription factors defines a prepattern for achaete and scute expression. The enhancers are named after the sensilla in which they direct achaete and scute expression. For example the L3/TSM enhancer, just upstream of scute directs achaete and scute to third vein sensilla campaniforma (L3) and twin sensilla of the anterior wing margin (TSM) (Gomez-Skarmeta, 1995). It is rather remarkable that each sensory organ or group is specificied by a special enhancer selected through evolution for just that purpose. For a historical view of this incredible work see Ghysen, 1988.

Embryonic central nervous system lineages

In Drosophila, central nervous system (CNS) formation starts with the delamination from the neuroectoderm of about 30 neuroblasts (NBs) per hemisegment. These give rise to approximately 350 neurons and 30 glial cells during embryonic development. Understanding the mechanisms leading to cell fate specification and differentiation in the CNS requires the identification of the NB lineages. Each segment is also subdivided along its dorsal-ventral axis. The delamination of the neuroblasts (NBs) from the neuroectoderm occurs between embryonic stages 8 and 11 and is divided into five phases (S1-S5). On each side of the ventral midline there are three longitudinal columns of cells: medial, intermediate and lateral. S1-S3 NBs form these three longitudinal columns. The positions of these three columns is regulated by genes defining the dorsal-ventral axis: twist, snail decapentaplegic and tolloid. These genes likewise act through the enhancer located between achaete and scute (Skeath, 1992). S4 and S5 NBs become interspersed between the existing columns of NBs.

Perpendicular to the columns, NBs are numbered in each segment from anterior to posterior as rows 1-7. Thus NB 1-2 is in the medial column (1) and represents the 2nd neuroblast in each segment. There are a total of thirty identified neuroblasts in each (right or left) half of a segment (hemisegment), and they give rise to a total of 350 progeny cells. For instance, NB 1-1 delaminates preferentially as a S1 NB. Around mid stage 11 the anterior-posterior position of NB1-1 is in line with, or slightly anterior to, the tracheal pits. At this stage it has given rise to a cluster of six to eight daughter cells on its dorsal side. At the beginning of germ band shortening (end stage 11) two of the progeny, the aCC and pCC leave the cluster and move anteriorly. They are the first progeny of NB 1-1. The aCC neuron innervates a dorsal muscle. Each thoracic NB 1-1 gives rise to 8 - 14 cells including ipsilaterally projecting interneurons and 1 or 2 motoneurons. In the abdomen NB 1-1 gives rise to a smaller cluster comprising four to six ipsilaterally projecting interneurons and no motoneurons; in addition the abdominal clone comprises 3 subperineural glial cells (Bossing, 1996). Different segmental fates of each neuroblast is determined by homeotic genes.

Thirteen lineages derived from the dorsal (lateral) part of the neuroectoderm are described here and 12 of them are assigned to identified NBs. Together, the 13 lineages comprise approximately 120 neurons and 22 to 27 glial cells which have been included in a systematic terminology. Therefore, NBs from the dorsal neuroectoderm produce about 90% of the glial cells in the embryonic ventral ganglion. Two of the NBs give rise to glial progeny exclusively (NB 6-4A, GP); five NBs give rise to glia as well as neurons (NBs 1-3, 2-5, 5-6, 6-4T, 7-4). These seven NBs are arranged as a group in the most lateral region of the NB layer. The other lineages (NBs 2-4, 3-3, 3-5, 4-3, 4-4, 5-4, clone y) are composed exclusively of neurons (interneurons, motoneurons, or both). It has been possible to link the lateral cluster of even-skipped expressing cells (EL) to the lineage of NB 3-3. Along with the previously described clones, the vast majority (more than 90%) of cell lineages in the embryonic ventral nerve cord (in the thorax and abdomen) are now known. Previously identified neurons and most glial cells are now linked to certain lineages and, thus, to particular NBs. This complete set of data provides a foundation for the interpretation of mutant phenotypes and for future investigations on cell fate specification and differentiation (Schmidt, 1997).

Aging neural progenitors lose competence to respond to mitogenic Notch signaling

Drosophila neural stem cells (neuroblasts) are a powerful model system for investigating stem cell self-renewal, specification of temporal identity, and progressive restriction in competence. Notch signaling is a conserved cue that is an important determinant of cell fate in many contexts across animal development; for example, mammalian T cell differentiation in the thymus and neuroblast specification in Drosophila are both regulated by Notch signaling. However, Notch also functions as a mitogen, and constitutive Notch signaling potentiates T cell leukemia as well as Drosophila neuroblast tumors. While the role of Notch signaling has been studied in these and other cell types, it remains unclear how stem cells and progenitors change competence to respond to Notch over time. Notch is required in type II neuroblasts for normal development of their transit amplifying progeny, intermediate neural progenitors (INPs). This study finds that aging INPs lose competence to respond to constitutively active Notch signaling. Moreover, reducing the levels of the old INP temporal transcription factor Eyeless/Pax6 allows Notch signaling to promote the de-differentiation of INP progeny into ectopic INPs, thereby creating a proliferative mass of ectopic progenitors in the brain. These findings provide a new system for studying progenitor competence and identify a novel role for the conserved transcription factor Eyeless/Pax6 in blocking Notch signaling during development (Farnsworth, 2015).

Notch signaling plays a crucial role in the control of proliferation and differentiation of stem and progenitor cells during embryogenesis or organogenesis, but its regulation is incompletely understood. BLOS2, encoded by the Bloc1s2 gene (see Drosophila Blos2), is a shared subunit of two lysosomal trafficking complexes, biogenesis of lysosome-related organelles complex-1 (BLOC-1) and BLOC-1 related complex. Bloc1s2-/- mice are embryonic lethal and exhibit defects in cortical development and hematopoiesis. Loss of BLOS2 results in elevated Notch signaling, which consequently increases the proliferation of neural progenitor cells and inhibits neuronal differentiation in cortices. Likewise, ablation of bloc1s2 in zebrafish or mice leads to increased hematopoietic stem and progenitor cell production in the aorta-gonad-mesonephros region. BLOS2 physically interacts with Notch1 in endo-lysosomal trafficking of Notch1. These findings suggest that BLOS2 is a novel negative player in regulating Notch signaling through lysosomal trafficking by controlling multiple stem and progenitor cell homeostasis in vertebrates (Zhou, 2016).

Fine-tuned Notch and Hedgehog signalling pathways via attenuators and dampers have long been recognized as important mechanisms to ensure the proper size and differentiation of many organs and tissues. This notion is further supported by identification of mutations in these pathways in human cancer cells. However, although it is common that the Notch and Hedgehog pathways influence growth and patterning within the same organ through the establishment of organizing regions, the cross-talk between these two pathways and how the distinct organizing activities are integrated during growth is poorly understood. An unbiased genetic screen in the Drosophila melanogaster eye has found that tumour-like growth was provoked by cooperation between the microRNA miR-7 and the Notch pathway. Surprisingly, the molecular basis of this cooperation between miR-7 and Notch converged on the silencing of Hedgehog signalling. In mechanistic terms, miR-7 silenced the interference hedgehog (ihog) Hedgehog receptor, while Notch repressed expression of the brother of ihog (boi) Hedgehog receptor. Tumourigenesis was induced co-operatively following Notch activation and reduced Hedgehog signalling, either via overexpression of the microRNA or through specific down-regulation of ihog, hedgehog, smoothened, or cubitus interruptus or via overexpression of the cubitus interruptus repressor form. Conversely, increasing Hedgehog signalling prevented eye overgrowth induced by the microRNA and Notch pathway. Further, it was shown that blocking Hh signal transduction in clones of cells mutant for smoothened also enhance the organizing activity and growth by Delta-Notch signalling in the wing primordium. Together, these findings uncover a hitherto unsuspected tumour suppressor role for the Hedgehog signalling and reveal an unanticipated cooperative antagonism between two pathways extensively used in growth control and cancer (Da Ros, 2013).

A challenge to understand oncogenesis produced by pleiotropic signalling pathways, such as Notch, Hh, and Wnts, is to unveil the complex cross-talk, cooperation, and antagonism of these signalling pathways in the appropriate contexts. Studies in flies, mice, and in human cell cultures have provided critical insights into the contribution of Notch to tumourigenesis. These studies highlighted that Notch when acting as an oncogene needs additional mutations or genes to initiate tumourigenesis and for tumour progression, identifying several determinants for such co-operation. The identification of these co-operative events has often been knowledge-driven, although unbiased genetic screens also identified known unanticipated tumour-suppressor functions. In this sense, this study describes a conserved microRNA that cooperates with Notch-induced overproliferation and tumour-like overgrowth in the D. melanogaster eye, miR-7. Alterations in microRNAs have been implicated in the initiation or progression of human cancers, although such roles of microRNAs have rarely been demonstrated in vivo. In addition, by identifying and validating functionally relevant targets of miR-7 in tumourigenesis, this study also exposed a hitherto unsuspected tumour suppressor role for the Hh signalling pathway in the context of the oncogenic Notch pathway. Given the conservation of the Notch and Hh pathways, and the recurrent alteration of microRNAs in human cancers, it is speculated that the genetic configuration of miR-7, Notch, and Hh is likely to participate in the development of certain human tumours (Da Ros, 2013).

In human cancer cells, miR-7 has been postulated to have an oncogene or a tumour suppressor functions that may reflect the participation of the microRNA in distinct pathways, due to the regulation of discrete target genes in different cell types, such as Fos, IRS-2, EGFR, Raf-1, CD98, IGFR1, bcl-2, PI3K/AKT, and YY1 in humans (Da Ros, 2013).

In Drosophila, multiple, cell-specific, targets for miR-7 have been previously validated via luciferase or in vivo eGFP-reporter sensors or less extensively via functional studiest. Although microRNAs are thought to regulate multiple target genes, when tested in vivo it is a subset or a given target that predominates in a given cellular context. Indeed, of the 39 predicted miR-7 target genes tested by direct RNAi, only downregulating ihog with several RNAi transgenes (UAS-ihog-IR) fully mimicked the effect of miR-7 overexpression in the transformation of Dl-induced mild overgrowth into severe overgrowth and even tumour-like growth. Moreover, it was confirmed that endogenous ihog is directly silenced by miR-7 and that this silencing involves direct binding of the microRNA to sequences in the 3'UTR of ihog both in vivo and in vitro (Da Ros, 2013).

Nevertheless, other miR-7 target genes may contribute to the cooperation with Dl-Notch pathway along with ihog, such as hairy and Tom. While miR-7 can directly silence hairy in the wing, this effect has been shown to be very modest, and thus, it is considered that while hairy may contribute to such effects, it is unlikely to be instrumental in this tumour model. Indeed, the loss of hairy is inconsequential in eye development, although retinal differentiation is accelerated by genetic mosaicism of loss of hairy and extramacrochaetae that negatively sets the pace of MF progression. It is unclear how Hairy might contribute to Dl-induced tumourigenesis (Da Ros, 2013).

The RNAi against Tom produced overgrowth with the gain of Dl albeit inconsistently and with weak penetrance, where one RNAi line did not modify the Dl-induced overgrowth and the other RNAi line caused tumours in less than 40% of the progeny. Tom is required to counteract the activity of the ubiquitin ligase Neuralized in regulating the Notch extracellular domain, and Dl in the signal emitting cells. These interactions are normally required to activate Notch signalling in the receiving cells through lateral inhibition and cell fate allocation. However, although it remains to be shown whether similar interactions are active during cell proliferation and growth, the moderate enhancement of Dl that is induced when Tom is downregulated by RNAi suggests that miR-7-mediated repression of Tom may contribute to the oncogenic effects of miR-7 in the context of Dl gain of function, along with other targets such as ihog (Da Ros, 2013).

Conversely, while the target genes of the Notch pathway, E(spl)m3 and E(spl)m4 as well as E(spl)mγ, Bob, E(spl)m5, and E(spl)mδ, have been identified as direct targets of miR-7 in the normal wing disc via analysis of 3'UTR sensors, there was no evidence that HLHm3, HLHm4, HLHm5, Bob, and HLHmγ are biological relevant targets of miR-7 in the Dl overexpression context. HLHmδ RNAi produced inconsistent phenotypes in the two RNAi transgenic lines available, causing tumour-like growth at very low frequency in only one of the lines. No evidence was obtained that miR-7 provoked overgrowth by targeting the ETS transcription factor in the EGFR pathway AOP/Yan, a functionally validated target of the microRNA miR-7 during retinal differentiation. Neither was any evidence obtained that RNAi of atonal provoked eye tumours with Dl overexpression, although a strong inhibition via expression of a fusion protein Atonal::EN that converts Atonal into a transcriptional repressor has been shown to be sufficient to trigger tumorigenesis together with Dl. Thus, it was reasoned that given that microRNA influenced target genes only subtly (even when using ectopic expression), it is possible that downregulation of atonal contributes to the phenotype along with the other targets (Da Ros, 2013).

In conclusion, this study has identified cooperation between the microRNA miR-7 and Notch in the D. melanogaster eye and identified and validated ihog as a direct target of the miR-7 in this context and have identified boi as a target of Notch-mediated activity at the DV eye organizer, although it remains whether this regulation is direct or indirect. A hitherto unanticipated tumour suppressor activity was uncovered of the endogenous Hh signalling pathway in the context of gain of Dl-Notch signalling that is also apparent during wing development (Da Ros, 2013).

Hh tumour suppressor role is revealed when components of the Hh pathway were lost in conjunction with a gain of Dl expression in both the eye and wing discs. Hh and Notch establish signalling centres along the AP and DV axes, respectively, of the disc to organize global growth and patterning. Where the organizer domains meet, the Hh and Notch conjoined activities specify the position of the MF in the eye disc and the proximodistal patterning in the wing disc. This study also unvailed that in addition antagonistic interaction between the Hh and Notch signalling might help to ensure correct disc growth. Thus, it was shown that Hh signalling limits the organizing activity of Dl-Notch signalling. Although it is often confounded whether Dl-Notch signalling instructs overgrowth by autonomous or nonautonomous (i.e., DV organizers) mechanisms, these findings uncover that loss of Hh signalling enhances a non-cell autonomous oncogenic role of Dl-Notch pathway (Da Ros, 2013).

To date, Hh has not yet to be perceived as a tumour suppressor, although it is noteworthy that human homologs of ihog, CDO, and BOC were initially identified as tumour suppressors. Importantly, both CDO and BOC are downregulated by RAS oncogenes in transformed cells and their overexpression can inhibit tumour cell growth in vitro. Since human RAS regulates tumourigenesis in the lung by overexpressing miR-7 in an ERK-dependent manner, it is possible that RAS represses CDO and BOC via this microRNA. Indeed, the 3'UTR of both CDO and BOC like Drosophila ihog contains predicted binding sites for miR-7. There is additional clinical and experimental evidence connecting elements of the Hedgehog pathway with tumour-suppression. The function of Growth arrest specific gene 1 (GAS1), a Hh ligand-binding factor, overlaps that of CDO and BOC, while its overexpression inhibits tumour growth . More speculative is the association of some cancer cells with the absence of cilium, a structure absolutely required for Hh signal transduction in vertebrate cells (Da Ros, 2013).

Given the pleiotropic nature of Notch, Wnts, BMP/TGFβ, Ras, and Hh signalling pathways in normal development in vivo, it is speculated that competitive interplay as that described in this study between Notch and Hh may not be uncommon among core growth control and cancer pathways that act within the same cells at the same or different time to exert multiple outputs (such as growth and cell differentiation). Moreover, context-dependent tumour suppressor roles could explain the recurrent, unexplained, identification of somatic mutations in Hh pathway in human cancer samples. Indeed, the current findings stimulate a re-evaluation of the signalling pathways previously considered to be exclusively oncogenic, such as the Hh pathway (Da Ros, 2013).

Notch (N) signaling is central to the self-renewal of neural stem cells (NSCs) and other tissue stem cells. Its deregulation compromises tissue homeostasis and contributes to tumorigenesis and other diseases. How N regulates stem cell behavior in health and disease is not well understood. This study shows that Notch regulates bantam (ban) microRNA to impact cell growth, a process key to NSC maintenance and particularly relied upon by tumor-forming cancer stem cells. Notch signaling directly regulates ban expression at the transcriptional level, and ban in turn feedback regulates N activity through negative regulation of the Notch inhibitor Numb. This feedback regulatory mechanism helps maintain the robustness of N signaling activity and NSC fate. Moreover, this study shows that a Numb-Myc axis mediates the effects of ban on nucleolar and cellular growth independently or downstream of N. These results highlight intricate transcriptional as well as translational control mechanisms and feedback regulation in the N signaling network, with important implications for NSC biology and cancer biology (Wu, 2017).

By revealing the involvement of the miRNA pathway, this study highlights the complexity of the N signaling network in normal NSCs and tumor-forming cancer stem cell (CSC)-like NSCs. Previous studies implicated critical roles for both canonical and non-canonical N signaling pathways in NSCs and CSC-like NSCs, and revealed particular dependence of CSC-like NB growth on non-canonical N signaling, which involves PINK1, mTORC2, and mitochondrial quality control. The current study reveals a particular requirement for ban in CSC-like NBs induced by N hyperactivation. The CSC-like NB overproliferation induced by hyperactivation of N or N pathway component Dpn can all be assumed to be of type II NB origin, since previous studies have clearly established that Notch signaling is essential for the development and/or maintenance of type II NBs, but dispensable for type I NBs, and that hyperactivation of Notch or its downstream effector Dpn induced ectopic CSC-like NB growth by altering the lineage homeostasis of the type II but not type I NBs. It would be interesting to test whether, in addition to ban's role in canonical N signaling, there exists a link between ban and non-canonical N signaling. The data indicate that the ban-Numb signaling motif regulates NSC/CSC behavior through at least two mechanisms. On one hand, it regulates cell growth and particularly nucleolar growth, through Myc, a known regulator of cellular and nucleolar growth. Consistently, negative regulation of Myc protein level by Numb was observed through E3 ubiquitin-protein ligase, Huwe1, and the UPS. c-Myc is an essential regulator of embryonic stem cell (ESC) self-renewal and cellular reprogramming, and Myc level and stability can be controlled in stem cells through targeted degradation by the UPS, suggesting conserved mechanisms. A key function of the nucleolus is the biogenesis of ribosomes, the cellular machinery for mRNA translation, and previous studies in Drosophila have supported the critical role of nucleolar growth in NSC self-renewal and maintenance. On the other hand, the ban-Numb axis feedback regulates the activity of N by a double negative regulation, with the end result being positive feedback regulation. This feedback mechanism may help transform initial not so dramatic differences in N activity between NB and its daughter cell generated by the asymmetric segregation of Numb during NB division [33] into 'all-or-none' decision of cell fates. Feed-forward regulatory loops, both coherent and incoherent, are frequently found in gene regulatory networks, and although ban miRNA is not conserved in mammals, miRNAs have been implicated in an incoherent feed-forward loop in the Numb/Notch signaling network in colon CSCs in mammals (Wu, 2017).

Given the role of ban in a positive feedback regulation of N and the potency of N hyperactivity in inducing tumorigenesis, one may wonder why ban overexpression is not sufficient to cause tumorigenesis. As in any biological systems, feedback regulation is meant to increase the robustness and maintain homeostasis of a pathway. Feedback alone, either negative or positive, should not override the main effect of the signaling pathway. Thus, in the NB system feedback regulation by ban is built on top of the available N signaling activity in a given cell and serving to maintain N activity. Because of ban's 'fine-tuning' rather than 'on/off switching' of Numb expression, its effect on N activity during feedback regulation will also be 'fine-tuning', serving to maintain N activity in NB within a certain range. Overexpression of ban in a wild type background may not be sufficient to cause tumorigenesis because N activity is not be elevated to the level sufficient to induce brain tumor as in N-v5 overexpression condition. Consistent with this, the extent of Numb inhibition by ban is also modest, not reaching the threshold level of Numb inhibition needed to cause tumorigenesis. Consistent with the notion that feedback regulation by ban is built on top of the available N signaling activity in a given cell, and that there is dosage effect of N activity in tumorigenesis, overexpression of ban in N-v5 overexpression background further enhanced N-v5 induced tumorigenesis. It is likely that ban or other miRNAs may participate in additional regulatory mechanisms in the N signaling network in Drosophila. Of particular interest, it would be interesting to test whether miRNAs may impinge on the asymmetric cell division machinery to influence the symmetric vs. asymmetric division pattern, a key mechanism employed by NSCs and transit-amplifying IPs to balance self-renewal with differentiation (Wu, 2017).

The results emphasize the critical role of translational control mechanisms in NSCs and CSC-like NSCs. Compared to the heavily studied transcriptional control, knowledge of the translational control of NSCs and CSCs is rather limited. As fundamental regulators of mRNA translation, miRNAs can interact with both positive and negative regulators of translation to influence gene expression. Thus, miRNA activity can be regulated context-dependently at both the transcriptional and translational levels, which may account for the opposite effect of N on ban activity in the fly brain and wing disc, although the ban genomic locus is bound by Su(H) in both tissues. Whether N regulates the transcription of ban or its activity as a translational repressor in the wing disc remains to be tested. With regard to the translation of numb mRNA, the conserved RNA-binding protein (RNA-BP) Musashi has been shown to critically regulate the level of Numb protein in mammalian hematopoietic SCs and leukemia SCs. Further investigation into the potential interplay between miRNAs and RNA-BPs in the translational control of Numb in NBs and CSC-like NBs promises to reveal new mechanisms and logic in stem cell homeostasis regulation, with important implications for stem cell biology and cancer biology (Wu, 2017).

Germline stem cells in the Drosophila ovary are maintained by a somatic niche. The niche is structurally and functionally complex and contains four cell types, the escort, cap, and terminal filament cells and the newly identified transition cell. The large Maf transcription factor Traffic jam (Tj) is essential for determining niche cell fates and architecture, enabling each niche in the ovary to support a normal complement of 2-3 germline stem cells. In particular, this study focused on the question of how cap cells form. Cap cells express Tj and are considered the key component of a mature germline stem cell niche. It is concluded that Tj controls the specification of cap cells, as the complete loss of Tj function caused the development of additional terminal filament cells at the expense of cap cells, and terminal filament cells developed cap cell characteristics when induced to express Tj. Further, it is proposed that Tj controls the morphogenetic behavior of cap cells as they adopted the shape and spatial organization of terminal filament cells but otherwise appeared to retain their fate when Tj expression was only partially reduced. The data indicate that Tj contributes to the establishment of germline stem cells by promoting the cap cell fate, and controls the stem cell-carrying capacity of the niche by regulating niche architecture. Analysis of the interactions between Tj and the Notch (N) pathway indicates that Tj and N have distinct functions in the cap cell specification program. It is proposed that formation of cap cells depends on the combined activities of Tj and the N pathway, with Tj promoting the cap cell fate by blocking the terminal filament cell fate, and N supporting cap cells by preventing the escort cell fate and/or controlling the number of cap cell precursors (Panchal, 2017).

Stem cells retain the capacity for development in differentiated organisms, which is important for tissue growth, homeostasis and regeneration, and for long-term reproductive capability. Stem cells are often associated with a specialized microenvironment, a niche that is essential for the formation, maintenance, and self-renewal of stem cells by preventing cell differentiation and controlling rate and mode of cell division. The niche for the germline stem cells (GSCs) in Drosophila serves as an important model for the analysis of interactions between niche and stem cells. The astounding fecundity of Drosophila females that can lay dozens of eggs per day over several weeks depends on approximately 100 GSCs that are sustained by 40 stem cell niches. To understand the formation and maintenance of these GSCs, it is important to understand how stem cell niches form and how they function (Panchal, 2017).

The GSC niche of the Drosophila ovary consists of three somatic cell types: cap cells, escort cells, and terminal filament (TF) cells. GSCs are anchored to cap cells by DE-cadherin-mediated adhesion and require close proximity to cap cells to retain stem cell character. Cap cells secrete the BMP homolog Decapentaplegic (Dpp), activating the TGFβ signaling pathway in adjacent GSCs, which leads to the repression of the germline differentiation factor Bag-of-Marbles (Bam). Through Hedgehog (Hh) signaling, cap cells also appear to stimulate escort cells to secrete Dpp. The combined pool of Dpp from cap and escort cells, together with mechanisms that concentrate Dpp in the extracellular space around GSCs, promotes the maintenance of 2-3 GSCs, whereas the adjacent GSC daughter cells that have lost the contact to cap cells will enter differentiation as cystoblasts. In contrast, TFs are not in direct contact with GSCs but serve important functions in the development and probably also in the maintenance and function of GSC niches (Panchal, 2017).

Formation of GSC niches begins with the progressive assembly of TFs by cell intercalation during the 3rd larval instar. The process of TF cell specification is not understood but might start in 2nd instar when the first TF precursor cells appear to leave the cell cycle. TF morphogenesis depends on the Bric à brac transcriptional regulators that control the differentiation of TF cells and their ability to form cell stacks, and involves the Ecdysone Receptor (EcR), Engrailed, Cofilin, and Ran-binding protein M (RanBPM). The number of TFs that form at the larval stage determine the number of GSC niches at the adult stage, and are regulated by several signaling pathways that control cell division and timing of cell differentiation in the larval ovary, including the EcR, Hippo and Jak/Stat, Insulin and Activin pathways. Despite the recent advance in elucidating mechanisms that control the number of GSC niches and the temporal window in which they form, relatively little is known about the origin and specification of the somatic cell types of the GSC niche (Panchal, 2017).

Notably, the origin and specification of cap cells, the main component of an active GSC niche is little understood. Cap cells (also called germarial tip cells) are first seen at the base of completed TFs at the transition from the 3rd larval instar to prepupal stage. They appear to derive from the interstitial cells (also called intermingled cells) of the larval ovary that are maintained by Hh signaling from TFs. The formation of cap cells is accompanied by the establishment of GSCs. The N pathway contributes to the development of cap cells. A strongly increased number of functionally active cap cells per niche form in response to overexpression of the N ligand Delta (Dl) in germline or somatic cells, or the constitutive activation of N in somatic gonadal cells. The ability of N to induce additional cap cells seems to depend on EcR signaling. Loss of Dl or N in the germline had no effect on cap cells. However, loss of N in cap cell progenitors or Dl in TF cells caused a decrease in the number of cap cells. A current model suggests that Dl signaling from basal-most TF cells to adjacent somatic cells together with Dl signaling between cap cells allows for a full complement of cap cells to form. Furthermore, N protects cap cells from age-dependent loss as long as its activity is maintained by the Insulin receptor. The Jak/Stat pathway, which operates downstream or in parallel to the N pathway in the niche, is not required for cap cell formation. As cap cells were reduced in number but never completely missing when the N pathway components were compromised, the question remains whether N signaling is the only factor that is important for cap cell formation. Furthermore, no factor that operates downstream of N has been identified that is crucial for cap cell formation (Panchal, 2017).

This study finds that Traffic jam (Tj) is both required for cap cell specification and for the morphogenetic behavior of cap cells, enabling them to form a properly organized niche that can accommodate 2-3 GSCs. Tj is a large Maf transcription factor that belongs to the bZip protein family. Its four mammalian homologs control differentiation of several cell types and are associated with various forms of cancer. Tj is essential for normal ovary and testis development, and is only expressed in somatic cells of the gonad. Interestingly, Tj is present in cap cells and escort cells but not in TFs. This study shows that Tj is essential for the formation of the GSC niche. First, Tj regulates the behavior of cap cells, enabling them to form a cell cluster instead of a cell stack, which appears to be important for the formation of a normal-sized GSC niche with the capacity to support more than one GSC. Second, cap cells adopt the fate of TF cells in the absence of Tj function, and TF cells develop cap cell-like features when forced to express Tj, indicating that Tj specifies the cap cell fate. Genetic interactions suggest that Tj and N are required together for cap cell formation, but have different functions in this process. For somatic gonadal cells to adopt the cap cell fate, it is proposed that Tj has to be present to inhibit the TF cell fate and N has to be present to prevent the escort cell fate and/or produce the correct number of cap cell precursors (Panchal, 2017).

Loss of Tj has a profound negative effect on the establishment, number, and maintenance of GSCs. Effects of Tj on the germline were previously shown to be indirect as Tj is neither expressed nor cell-autonomously required in the germline. Therefore, it is proposed that the dramatic change in the structure of the somatic niche affects GSCs when Tj function is compromised. An inverse causal relationship, where a reduced number of GSCs would trigger the somatic niche defects was ruled out by showing that cap cells can still look and behave normally in the absence of any germ cells. It is concluded that Tj controls GSCs indirectly by controlling somatic cell fate and cell arrangement in the stem cell niche (Panchal, 2017).

By controlling the morphology and behavior of the cap cells, Tj regulates the GSC-carrying capacity of the niche. When Tj expression was moderately reduced, the number of GSCs per niche was reduced, with the remaining GSC properly maintained over several weeks. The decrease of GSCs per niche correlated with a decrease of cap cells in the germarium. Two cap cells were on average required to sustain one GSC, similar to what has been proposed for a wild-type ovary. The data indicate that the reduced niche capacity is due to a reduction in the available contact surface between cap cells and GSCs. Tj-depleted cap cells that convert from forming a cluster inside the germarium to forming a stalk outside the germarium minimize their availability for GSC attachment. A connection between the GSC-cap cell contact area and niche capacity is similarly reflected in the increased number of GSCs that accompanies an increase in cap cell size due to loss of RanBPM. This study shows that the spatial arrangement of the cap cells has a crucial impact on the number of stem cells per niche (Panchal, 2017).

When Tj function was completely abolished, the number of GSCs was drastically reduced, as expected in the absence of cap cells. The very few pMad-positive GSC-like cells in tj mutant prepupal ovaries were always associated with a TF, suggesting that TFs might temporarily provide enough Dpp to activate Mad in a few germline cells, consistent with the finding that Dpp is expressed in TFs at the late larval stage\. This is not sufficient, however, to maintain GSCs and adult ovaries rarely contain pMad-positive germline cells. This is in agreement with the finding that Dpp is not detected in adult TFs, and corroborates that cap cells are required for GSC maintenance. In addition, the rapid loss of the entire germ cell pool in Tj-depleted ovaries during the pupal stage might be precipitated by loss or defects in escort cells. Escort cell precursors are not properly intermingled with germ cells at the larval stage and differentiated escort cells appear to be missing in adult ovaries that lack Tj. As escort cells are crucial for germ cell differentiation, the defect in escort cell differentiation could be responsible for the demise of the germline in tj mutants (Panchal, 2017).

GSCs have broad cellular protrusions, which they use to reach and tightly ensheath the accessible surface of cap cells. In wild type, relatively short protrusions are sufficient to make extensive contact with more than one cap cell. However, when cap cells formed a stalk, GSCs were often observed to produce unusually long extensions that allowed them not only to contact the immediate cap cell neighbor but also a more distantly located cap cell. This suggests that GSCs respond to a chemotactic signal from cap cells and send protrusions toward this signal. It remains to be investigated whether this is a response to Dpp signaling or signaling through another pathway. The importance of cellular protrusions in signaling events in the stem cell niche has recently come to light with the discovery of nanotubes that mediate Dpp signaling between GSCs and hub cells in the Drosophila testes, and cytonemes that contribute to Hh signaling from cap to escort cells in the ovary (Panchal, 2017).

This analysis shows that Tj is required for the specification of cap cells. In the absence of Tj function, additional TF cells form at the expense of cap cells, resulting in unusually long TFs while the cap cell fate is not established. Whereas the formation of cap cell precursors appears not to require Tj, this transcription factor is essential for the ability of these precursors to take on the cap cell fate and to prevent the TF cell fate that is otherwise adopted as a default state. The following findings support this conclusion: (1) In the absence of Tj function, cap cells were missing while additional cells that displayed TF cell-characteristic morphology, behavior and marker expression were integrated into the TF. The number of additional TF cells was comparable to the normal number of cap cells. (2) Prospective cap cells cell-autonomously adopted a TF-specific morphology and behavior in the absence of functional Tj. (3) A hypomorphic tj mutant provided direct evidence for the incorporation of cap cells into TFs, forming the basal portion of these stalks. (4) Ectopic expression of Tj in TF cells caused a change toward cap cell-typical marker expression and morphology. Together, these data demonstrate that Tj promotes cap cell specification (Panchal, 2017).

The expression pattern of Tj supports the notion that Tj has a function in cap cells but not in TF cells. Tj is continuously expressed in cap cells. Tj is also present in the anterior interstitial cells of the larval ovary, which are thought to develop into cap cells. In contrast, Tj is neither detected in the cell population that gives rise to TFs during 3rd larval instar, nor in differentiated TFs. Interestingly, even in the absence of Tj function, the tj gene remains differentially expressed in the anterior niche, being inactive in regular TF cells but active in the additional TF cells, which form the apical and basal portion of a TF, respectively. This differential expression of Tj indicates that a regionally or temporally regulated mechanism operates upstream of Tj that initiates differences in anterior niche cells. Although it is conspicuous that Tj expression from 3rd instar onwards is restricted to cells that are in direct contact with germline cells, which includes cap cells but excludes TF cells, it has previously been shown that Tj expression is not dependent on the germline. This suggests that a soma-specific mechanism is responsible for the differential expression of Tj in anterior niche cells. Interestingly, a recent study uncovered the importance of Hh signaling from TFs to neighboring interstitial cells in the larval ovary and proposes that tj is a direct target of the Hh signaling pathway (Panchal, 2017).

The current findings suggest the presence of a new cell type in the GSC niche that has been named 'transition cell' as it is located between the cap cell cluster and the TF, connecting these two structures of the niche. Notably, the one or occasionally two transition cells have the morphology of TF cells and align with neighboring TF cells despite displaying a cap cell-like marker profile that includes the expression of Tj—although Tj expression is substantially lower than in cap cells. Interestingly, cap cells from ovaries with reduced Tj expression (tjhypo) similarly displayed a TF cell-like morphology and behavior while their expression profile remained cap cell-like. A similar, although weaker effect was noted in a tj hemizygous condition, suggesting that Tj function is haplo-insufficient in cap cells. Thus, when Tj levels are reduced, cap cells adopt very similar molecular and morphogenetic properties as the transition cell in a wild-type niche, and might have adopted this cell fate (Panchal, 2017).

Together, the current findings indicate that Tj has an important role in the establishment of three cell types in the GSC niche: TF cells, transition cells, and cap cells. As lack of Tj function seems to cause a transformation of cap and transition cells into TF cells, and a mild reduction of Tj a cap to transition cell transformation, it is proposed that different Tj expression levels establish different cell fates and morphogenetic traits. It is proposed that a high concentration of Tj leads to the formation of cap cells and a lower concentration to the formation of the transition cell, whereas absence of Tj is required for the formation of TF cells. This model implies that different levels of Tj have different effects on target genes. It is predicted that Tj has at least one target gene that only responds to high levels of Tj and that specifically controls the morphogenetic behavior of cap cells, allowing them to adopt a round morphology and organize into a cell cluster. Whether this relates to an effect of Tj on the expression of adhesion molecules as observed in other gonadal tissues awaits further analysis (Panchal, 2017).

This study identifies Tj as essential for cap cell formation. In addition, this process depends on the N pathway. Therefore, it was asked how the functions of Tj and N in cap cell formation relate to each other. A comparison between the loss and gain-of-function phenotypes suggests that Tj and N have different functions in the establishment of cap cells. In the absence of Tj function, cap cell precursor cells are present but take on the fate of TF cells, whereas depletion of N leads to a loss of cap cells but does not cause the formation of additional TF cells. Ectopic activation of N can induce a strong increase in the number of cap cells, whereas overexpression of Tj did not appear to affect the number of cap cells. Therefore, both factors are important for cap cell formation but contribute differently to this process. The questions then are: What is the respective contribution of Tj and N to cap cell formation, and how are their functions related (Panchal, 2017)?

The function of N in cap cell formation is still not fully understood. The observation that depletion of N reduces the number of cap cells confirms previous findings. However, neither in this nor any previously published experiments were cap cells lost completely when the N pathway was compromised, and it remains therefore unclear whether N is de facto essential for cap cell formation or primarily functions in regulating the size of the cap cell pool. Interestingly, evidence amounts to a function of the N pathway in a decision between the cap cell and escort cell fate: First, Dl signal from TF cells activates the N pathway in adjacent interstitial cells, inducing them as cap cells, whereas the remaining interstitial cells are thought to develop into escort cells. Second, escort cells expressing activated N can develop into cap cells. Third, when tj-Gal4 was used to express active N in interstitial cells, the number of cap cells dramatically increased while the escort cell region became smaller, and some germaria seemed to lack escort cells all together. These germaria also lacked germline cells, although a larger pool of cap cells was expected to increase the number of GSCs. However, the absence of germline cells is consistent with an absence of escort cells, as escort cells have been shown to be important for maintaining the germline. Together, these observations support the hypothesis that N is involved in a cap cell versus escort cell fate decision, and suggest that the N pathway might promote the formation of cap cells by inhibiting the escort cell fate (Panchal, 2017).

To determine how the functions of Tj and N depend on each other, genetic interactions were examined. The N pathway seems to be still functional in tj mutants. First, the expression of N and Dl appeared unaffected and E(spl) was activated in the additional TF cells (= transformed cap cells) similarly to normal cap cells. Second, the formation of additional TF cells in the absence of Tj depended on the presence of N, as only very few additional TF cells formed in a N compromised background. These findings indicate that the N pathway is still active in cap cell precursors when Tj function is abolished. This together with the observation that constitutively active N cannot suppress the tj mutant phenotype suggests that Tj does not act upstream of N in regulating cap cell fate (Panchal, 2017).

Therefore, it was asked whether Tj might operate downstream of N. Loss of Tj was not detected upon N depletion, and this together with the finding that Tj is expressed in all interstitial cells, and not only in those that receive Dl signaling argues against a requirement of N signaling for tj expression. If at all, one would expect tj to be negatively regulated by N as cap cells express a lower level of Tj than escort cells. The maintenance of somatic cell types in N mutant ovaries that are lost in tj mutant ovaries, including the escort cells is also not consistent with a linear relationship. Nevertheless, the ability of Tj to promote the formation of cap cells appears to depend on the activity of the N pathway in cap cell precursors. Again, this is suggested by the finding that when N and Tj were both compromised, the number of additional TF cells were much smaller than when N was fully active. Therefore, it is proposed that N activity sets aside a pool of percursor cells that in the presence of Tj take on the cap cell fate, and in its absence the TF fate (Panchal, 2017).

Similar to the ovary, N is important for the formation of the GSC niche (= hub) in the Drosophila testis. Interestingly, N contributes to hub cell specification by downregulating the expression level of Tj. Not only is the hub still present in tj mutant testes but additionally, ectopic hub cells form in the absence of Tj. Thus, Tj seems to have opposing functions in testes and ovaries, suppressing the niche cell fate in the testis, while promoting it in the ovary (Panchal, 2017).

The interplay between Tj and N seems not restricted to the cap cell fate in the ovary. Whereas neither factor alone is required for TF cell formation, as TF cells formed normally in the absence of either Tj or N, the combined loss of Tj and N led to a strong reduction in the number of TFs and number of TF cells within stalks. This suggests that their combined action is already required at an earlier stage of ovary development, when Tj is still expressed in all somatic cells of the ovary. Moreover, Tj knockdown combined with expression of activated N caused TF cells to be the only cell type remaining of the ovary, indicating that several cell types in the ovary require proper input from both factors. Taken together, the findings support a model, in which both Tj and N operate together to promote the cap cell fate but have separate functions. It is proposed that Tj and N promote the cap cell fate by blocking the TF cell fate and escort cell fate, respectively, and that the combined actions of Tj and the N pathway are required to establish the cap cell fate (Panchal, 2017).

In contrast to vertebrates that have a single Pax6 gene, the Drosophila genome contains two Pax6 homologs, ey and toy. Both genes are expressed broadly throughout the entire eye-antennal disc but are later limited to a far more restricted domain within the undifferentiated cells of the eye field. Whereas most studies on Pax6 in the eye-antennal disc have focused on the developing compound eye, several reports have hinted at a role for both genes outside of the eye. However, the underlying mechanism of how Ey/Toy promote eye-antennal disc development has been elusive. This is, in part, because of the use of single Pax6 mutants to study development. The phenotypes associated with individual mutants are variable and often restricted to the eye. Several studies have suggested that Ey and Toy function redundantly to each other. This finding most likely explains the variability of phenotype severity and penetrance. Thus, the combined loss of both Ey/Toy may be a more accurate reflection of the effect that Pax6 loss has on Drosophila development. Indeed, this appears to be the case as it is reported that the combined loss of both ey and toy leads to the complete loss of all head structures that are derived from the eye antennal disc. This study attempted to determine the mechanism by which Ey/Toy support eye-antennal disc development (Zhu, 2017).

Previous studies in the fly eye proposed that Pax6 is concerned solely with eye specification, whereas Notch signaling and other retinal determination proteins, such as Eyg, Tsh, and Hth, control cell proliferation and tissue growth. This study proposes an alternate model in which Ey/Toy are in fact required for cell survival and proliferation in addition to eye specification. The data indicate that Ey/Toy regulate growth of the eye-antennal disc through Tsh, N/Eyg, and additional N-dependent proliferation promoting genes. It is proposed that on simultaneous removal of Ey and Toy the eye-antennal disc fails to develop, in part, because the expression of eyg and tsh is lost in complete absence of Pax6. Expression of tsh and activation of the N pathway are sufficient to restore tissue growth to the eye-antennal disc. Support for this model linking Ey/Toy to cell proliferation via Eyg and Tsh comes from studies showing that eyg loss-of-function mutants display a headless phenotype identical to that seen in the ey/toy double knockdowns, that cells lacking eyg do not survive in the eye disc, and overexpression of Tsh causes overproliferation (Zhu, 2017).

The results also show that the combined loss of Ey and Toy affects the number of cells that are in S and M phases of the cell cycle. This observation directly supports the model that Ey/Toy control growth of the eye-antennal disc and is consistent with studies in vertebrates that demonstrate roles for Pax6 in the proliferation of neural progenitors within the brain. Earlier studies observed cells undergoing apoptosis in Pax6 single-mutant eye-antennal discs and showed that blocking cell death alone can partially rescue the head defects of the eyD and toyhdl mutants. Although this study shows that retinal progenitor cells lacking both Pax6 proteins undergo even greater levels of apoptosis, blocking cell death does not restore the eye-antennal disc. What accounts for the differences in the two experiments? In the eyD and toyhdl rescue experiments, each genotype contained wild-type copies of the other Pax6 paralog, but this study has knocked down both Pax6 genes simultaneously. Another possible difference is that Pax6 levels are being reduced while the eyD and toyhdl mutants are likely functioning as dominant negatives. It is concluded from these results that a reduction in cell proliferation but not elevated apoptosis levels is the proximate cause for the complete loss of the eye-antennal disc (Zhu, 2017).

Although the activation of Tsh and the Notch pathway can restore antennal and head epidermal development, neither factor is capable of restoring eye development to the ey/toy double-knockdown discs. This is most likely because both Pax6 genes are also required for the specification of the eye. In particular, Ey/Toy are required for the activation of several other retinal determination genes, including so, eya, and dac. Thus, the results suggest that Notch signaling, Eyg, and Tsh can restore nonocular tissue growth to the eye field but cannot compensate for the Pax6 requirement in eye specification (Zhu, 2017).

Finally, the results using the double knockdown of ey/toy are consistent with the dosage effects that are seen in mammalian Pax6 mutants. Although mutations in ey have just eye defects, the combined loss of ey/toy lacks all head structures. Mice that are heterozygous for Pax6 mutations have small eyes, whereas those that are homozygous completely lack eyes, have severe CNS defects, and die prematurely. Similarly, human patients carrying a single mutant copy of Pax6 suffer from aniridia, whereas newborns that are homozygous for the mutant Pax6 allele have anophthalmia, microcephaly, and die very early as well. As a master control gene of eye development, Pax6 appears to initiate both retinal specification and proliferation. These data demonstrate that the functions of Ey and Toy in the eye-antennal disc are redundant and dependent upon gene dosage, thereby making the roles of Pax6 in the Drosophila similar to what is observed in vertebrates where Pax6 controls both specification and proliferation of the brain and retina in a dosage-sensitive manner (Zhu, 2017).

list of proneural and neurogenic genes

Bossing, T., et al. (1996). The embryonic central nervous system lineages of Drosophila melanogaster. I. Neuroblast lineages derived from the ventral half of the neuroectoderm. Dev. Biol 179: 41-64. 8873753

Cummings, C.A. and Cronmiller, C. (1994). The daughterless gene functions together with Notch and Delta in the control of ovarian follicle development in Drosophila. Development 120: 381-94. 8149916

Da Ros, V. G., Gutierrez-Perez, I., Ferres-Marco, D. and Dominguez, M. (2013). Dampening the signals transduced through Hedgehog via microRNA miR-7 facilitates Notch-induced tumourigenesis. PLoS Biol 11(5): e1001554. PubMed ID: 23667323

Farnsworth, D. R., Bayraktar, O. A. and Doe, C. Q. (2015). Aging neural progenitors lose competence to respond to mitogenic Notch signaling. Curr Biol 25: 3058-3068. PubMed ID: 26585279

Ghysen, A. and Dambly-Chaudiere, C. (1988). From DNA to form: The achaete-scute complex. Genes Dev. 2: 495-501

Gomez-Skarmeta, J. L., et al. (1995). Cis-regulation of achaete and scute: shared enhancer-like elements drive their coexpression in proneural clusters of the imaginal discs. Genes Dev. 9: 1869-1882. 7649474

Gonzalez-Gaitan, M. and Jackle, H. (1995). Invagination centers within the Drosophila stomatogastric nervous system anlage are positioned by Notch-mediated signaling which is spatially controlled through wingless. Development 121: 2313-25. 7671798

Hartenstein, A.Y., Rugendorff, A., Tepass, U. and Hartenstein, V. (1992). The function of the neurogenic genes during epithelial development in the Drosophila embryo. Development 116(4): 1203-1220. 1295737

Panchal, T., Chen, X., Alchits, E., Oh, Y., Poon, J., Kouptsova, J., Laski, F. A. and Godt, D. (2017). Specification and spatial arrangement of cells in the germline stem cell niche of the Drosophila ovary depend on the Maf transcription factor Traffic jam. PLoS Genet 13(5): e1006790. PubMed ID: 28542174

Schmidt, H., et al. (1997). The embryonic central nervous system lineages of Drosophila melanogaster. II. Neuroblast lineages derived from the dorsal part of the neuroectoderm. Dev. Biol. 189(2): 186-204

Schweisguth, F. (1995). Suppressor of Hairless is required for signal reception during lateral inibition in the Drosophila pupal notum. Development 121: 1875-1884. 7601001

Skeath, J. B. et al. (1992). Gene regulation in two dimensions: the proneural achaete and scute genes are controlled by combinations of axis-patterning genes through a common intergenic control region. Genes Dev 6: 2606-19. 7601001

Skeath, J. B., Panganiban, J. F. and Carroll, S. B. (1994). The ventral nervous system defective gene controls proneural gene expression at two distinct steps during neuroblast formation in Drosophila. Development 120: 1517-1524. 8050360

Wu, Y. C., Lee, K. S., Song, Y., Gehrke, S. and Lu, B. (2017). The bantam microRNA acts through Numb to exert cell growth control and feedback regulation of Notch in tumor-forming stem cells in the Drosophila brain. PLoS Genet 13(5): e1006785. PubMed ID: 28520736

Zhou, W., He, Q., Zhang, C., He, X., Cui, Z., Liu, F. and Li, W. (2016). BLOS2 negatively regulates Notch signaling during neural and hematopoietic stem and progenitor cell development. Elife [Epub ahead of print]. PubMed ID: 27719760

Zygotically transcribed genes

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Topic accent causes letter to change font

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Keyboard Details Galaxie BibleScript Phonetic Keyboards Keyboard Downloads and Details

# accent causes letter to change font   2009-09-13 16:35:28.653
Marcia Moore
OK, so I can now type accents, but whenever I put an accent over a letter, the font of the letter changes from whatever font I was typing in (e.g., Cambria, or Arial, or Book Antiqua, or Times New Roman) to the "Tahoma" font. I do not want to use the Tahoma font.

How can I accent letters and keep to the same font?
# RE: accent causes letter to change font   2009-09-14 14:46:56.980
I have a simple solution to your issue.

Fonts like Cambria and Ariel do not offer the best support for polytonic Greek, which is why your system searches for other fonts with better support. For the Greek accents to display correctly in your document, you need to use a font with good support for polytonic Greek. Fonts like Galatia SIL ( and Gentium SIL are two good choices (

You can find other good fonts for Greek from the Font Helper tool of Keyman's On Screen Keyboard. Simply click on the Keyman Menu, select the On Screen Keyboard, and choose Font Helper from the buttons along the top. You will see a list of the fonts on your system which offer the best support for Greek.


Yoga Basics

Asanas 0 8 limbs of yoga

Movement is the center of our world and the center of our being.  It’s what creates action and in turn produces change and growth within society. Movement within the yoga practice is referred to as Asana; it is the third petal of the eight limbs of yoga. For many people, the physical postures are their first glimpse into yoga and why this ancient practice has become one of the most popular forms of exercise.

Millions of people have found yoga through a physical fitness journey. The intricate postures, the beautiful people, and the image of what yoga is, have made a profound effect on present society.  Whether the practice was introduced at a gym, by a friend, or by purchasing a Groupon, it doesn’t matter. What matters is the relationship cultivated between yourself and the yoga practice. For yoga’s benefits to take effect it requires constant nourishment; nourishment in the form of a loving attitude, a daily commitment to move the body, to only digest pure nutrients, to shed layers that no longer serve you, and to practice wholehearted truthfulness.

Ironically, ancient yoga tradition only practiced four seated postures and all were in preparation for meditation. This is a drastic change from our modern yoga practices that are filled with creative sequences, trendy music, heated rooms, and designer yoga apparel. While the setting and context of our Asana practice has changed, the end goal has not: to be free from suffering and experience bliss.

Experience bliss – say what? Yes, that’s right. All yogis and yoginis of the world can glimpse into a world of bliss: a world that is filled with kind, generous, and like-minded people; a world where gossip and judgment are foreign; and a world where peace and love are the only answers. You might not experience this every moment BUT you will experience this within the yoga community and within yourself. The physical practice is a means to turn inward, to dim the exterior, to reconnect with our true nature, and to ultimately achieve that beautiful state of bliss.

So how do we do this? How can you or I feel the yoga high? The answer lies within the physical asana practice. Do the work and the rest will come. Make time everyday to step onto the mat, whether it’s five minutes or ninety minutes, simply drop into the moment. Allow the moment to be filled with breath and movement. Let the mind become quiet, let the skin begin to soften, let your inhibitions and fears drip away like rain falling from the sky. LET YOGA DO YOU!

A yoga practice can mean whatever it needs to be. Some days it will consist of a sweaty, heart pumping, creative sequence with inversions and backbends. Other days it will be gentle with longer holds, deeper breathing, and an airy softness to it. Regardless of the movement, the outcome will remain the same: a glimpse into bliss.

There’s that word again – bliss.

Upon first reaction, this word conjures up far-fetched images that seem impossible to experience. But that’s the beauty of yoga, bliss can have 1,00 different meanings and yet still be a state that everyone achieves. A friendship that comes from joining a yoga studio, a peaceful savasana at the end of practice, a handstand hold for five breaths, a moment where the mind is completely still, and a realization that our purpose in life is greater than it appears – this is all bliss.

With this understanding of what the yoga practice can offer, now is the time to step onto your mat. Join a local yoga studio, pair up with a friend and practice outdoors, enroll in one of the many online yoga communities, go on a yoga retreat somewhere beautiful, or simply practice in the comfort of your home. The how doesn’t matter; it’s the quality of the movements and the intention that is put forth Make a commitment to better yourself and start practicing yoga. One breath, one movement, and bliss will be around the corner shining brightly.

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The single dependency in configobj so...

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Setup Config File

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How Do I Create or Incorporate Text In An Oil Painting?
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How Do I Create or Incorporate Text In An Oil Painting?

Ideas on Incorporating text into a painting or along side a painting.

If you are making an oil painting and want to include a short poem or other text in the work or along side the work you have a number of options.

Incorporate the Text as Part of the Painting

Depending on your style of painting and your subject matter you will want to consider whether your text will be in a formal style font or a more painterly style font. You might want to first print and cut out the exact text from your computer and play around with compositions and configurations on the canvas. Rough in with pencil your picture composition.  Consider incorporating the poem or text into a border around the main subject part of the painting. Choose a canvas that is bigger than what you want for the main subject then mark off a border according to the size and area you will need for the text and paint the border area with a layer of oil paint in a color that will work with your painting. Use a drying agent and make sure that the surface is dry before proceeding. Smooth it out so you can write out the text all around the main subject in an artistic hand. Do your writing before the painting. Use a really nice set of markers such as Prismacolor or one of the great scrapbooking markers available that have different styles of tips from a fine tip to a "brush" style tip or paint the letters using a small brush. Understand that you have reserved the central part of the canvas for the main subject. You may want to put a hard line painted border between the main subject and the text for a more formal look or you may want to blend the background of the painting softly into the text area. Paintings with text incorporated can be really beautiful and compelling. Make the text a part of the painting rather than a separate element. After your painting has dried for several months you can varnish the entire work with the appropriate oil painting varnish including the text area.

Carve the Words Into the Wet Paint

If you are using an impasto technique you can use a stylus and carve the words into the paint while it is still wet. This will be a more painterly or informal look.

Use a Shadow Box Alongside your Painting

If your painting is a painting to commemorate a loved one you can have a brass plate with the name and dates on it and have it adhered to the frame at the bottom. The text or poem can be framed under glass and hung separately. Most frame shops have a vendor that will custom make a small plate for the frame. Be sure to choose a flat frame if you are doing this or one that has a somewhat flat area on it for the plate.

For a more formal approach you might choose to do a separate frame for the poem or text. I would recommend you consider creating a framed and glassed shadow box and include memorobilia along with the written document. A pair of glasses, gloves, jewelry or a necktie or in the case of an animal a collar or toy. Consult a good independent framer for shadow boxes and design options. See my article on Picture Framing Tips for some ideas on selecting the appropriate frame and colors.

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Great article. When I saw this question, I was hoping someone would write a factoid about it!

Thanks Kathleen!


Handy Plans In Golfing – Some Updated Tips

If your golfing putts need work, there are a couple of easy drills to get them up to snuff. One easy exercise is to throw a dime a short ways and try to get the ball to land on it. Another drill is to try to sink five one-foot shots in a row, and then increase distance.

To improve your score fast, focus on your short shots because they make up the greater percentage of the game. New golfers often put a lot of focus on driving, but this won’t bring up your score as quickly as getting really good at getting the ball into the hole.

Golfing can be loads of fun or a relaxing getaway. The way you play the game can change depending on the circumstances involved. This is why golfing is one of the most popular sports around the world. Golf has a learning curve that allows you to really get good at the game, so here are some tips to help you on your way.

Good golfers never forget that the perfect golf swing is a full-body motion. The hips and legs need to work in concert with the arms to maximize power and maintain accuracy. At the same time that the arms bring the club down, the hips should be swiveling, and once the shot is made the legs should pivot for full follow-through.

It is wise to avoid buying golf clubs over the internet, even if you have seen a picture of it. This is because although the club may seem to be in good condition, there could be flaws or problems with the club. Instead, buy a new, durable one at a sporting goods store.

Hone your skills and sharpen your game by applying the lessons in this article. If you make it a priority to adapt the knowledge you learn about the game into your practice sessions and weekend jaunts, even the driving range can become a theater for improvement. Use this article to boost your performance on the court and send your handicap into the basement.

A helpful tip when it comes to golf is to make sure that the last club you use in practice before heading out on the golf course is the first club you use when you tee off. This will ensure that you are ready for your first shot – the most important one of the day.

There are many ways to putt a golf ball, but for the beginner golfer the most simple way to remember proper putting is the word “center”. All puts should originate from the center of the putter while you are still learning. Once you become more experienced, you can begin to learn about zones of the putter and how that will affect your shot.

If you want to improve your golf game, it is crucial that you hold your club in the correct way. First of all, you should place the club handle onto your palms. Then, you should grab the club with thumbs going down. Your hands should be touching. If you want the ball to go to its maximum distance, one technique is to move your hands up on the handle a little. For a shorter distance, you should move your hands down a little.

The back swing of a golf swing is just as important as the front swing- it must be gradual and controlled. Amateurs will tend to bring the club back quickly and jerkily with the wrists, which compromises the whole swing. Bring it back in a controlled fashion with the arms to start your swing off right.

When performing a golf swing, the right leg is the root of everything. It’s essential to keep weight on your right leg, with the knee slightly bent, during the back swing. If your right leg is weak, it means a major lack of power. Imagine your knee braced against a solid wall while swinging.

If you are right-handed, swing the putter with the left hand in front of the ball at the point of impact. Maintain this position as you follow through with your swing. This helps to keep the ball from bouncing too hard off the club face, and will also hold the club firmly in your grasp.

It doesn’t matter if you are a golf novice or seasoned pro, you will hit sand traps and bunkers from time to time. Sand often times gets spread around carelessly trying to free the ball from a trap. Follow golf etiquette and properly rake the bunker prior to leaving. By leaving the trap raked cleanly, you allow players behind you the chance to play their shot without the frustration of dealing with an unfair lie.

Examining The Facts For Fundamental Elements Of Golfing

When you walk around the golf source, your feet typically expand a good bit due to the swelling caused by walking. Therefore, you should purchase golf shoes one size bigger than usual to compensate for this. Shoes that fit snugly in the store may end up uncomfortably constrictive by the eighteenth hole.

The way your golf game ends depends on your putting capabilities. Putting is one of the most difficult things to do correctly in golf. It is recommended that you go to the putting greens regularly just to practice your putting and nothing else. This will be very beneficial to you as you continue to play rounds of golf.

Understand when to use a long and short iron. A long iron needs a steady tempo to achieve a long distance. Don’t forget to swing slowly when using a long iron: smoother is better. On the other hand, a short iron is best used when you are within 150 yards of the green. Pay attention to the speed with which you hit the golf ball, and once again, swing smoothly. There is a tendency to swing a short iron fast, but this will not get the best results.

Many people complain about not being able to improve their game once they hit a certain point. This is a good time to go back to the basics and maybe even take some golfing lessons from a pro. Usually there are certain things that can be noticed and improved upon. This will enable you to go play your next round with a few tricks up your sleeve that improve your game.

Be careful of an overly aggressive or choppy golf swing- even the most powerful swing should be smooth and fluid. The key point here is the top of the swing between ending the back swing and beginning the forward swing. Don’t jerk the club from one to the other- accelerate gradually for a good, controlled swing.

In order to be good at golf, one of the basic things that you need to take care of is your grip. It is good to know how the ball will act in response to your grip. The tighter you hold the golf club the more tendency that the ball will go to the right. The looser the grip is the more tendency that the ball will go to the left.

When you are putting and the ball consistently misses to the left or the right, there is a reason for this. Your eyes are supposed to directly be positioned right over the ball. If they are positioned slightly inside the ball, then your shot will miss to the right. If they are positioned outside the ball, then your shot will consistently miss to the left.

If you need to make a longer than normal bunker shot, try taking a nine iron instead of a sand wedge. By taking a nine iron you increase the distance you are able to carry the ball out of the sand and decrease the likelihood that you will skull the ball by taking a sand wedge and swinging too hard.

One way to ensure that your swing is strong and consistent is to swing all the way through without breaking your wrist. You can place a tee about 6 to 8 inches in front of your ball. After you hit the ball, you want to hit the tee in front of it as well.

If there are things you don’t understand about the game of golf, don’t be afraid to ask! Ask your your fellow golfers, and friends at your club. Try to go for a round with a really good player and ask them to analyze your game. Read up on golf and watch golf tournaments and training videos; however, remember that you will create your own unique style as you learn and apply the advice.

If you are looking to hone your putting skill, you should roll a golf ball across a carpet in your home. By doing this, you will begin to understand speed and line without even having to pick up your club. This is an easy way to hone your mental understanding of the golfing game.

Have you recently taken up the sport of playing golf, or perhaps you have been playing golf for years? Either way, there are always things that you can learn about golf that you never knew existed. What you are going to read about golf in the following article may take you by surprise.

Golfing is a fun and easy sport. It gets you outside and keeps you active. That is why people young and old can play the sport, and you can too! Just stick with it! You will not be any good at first, but that is okay. You may get embarrassed because your swing is different than others, or you take longer, but don’t! You are out there to have fun and challenge yourself, remember that.

A tip for practicing difficult golf putts is to try them with your eyes closed. It’s easy to get distracted from your technique when you want to watch the ball go to the hole- if you shut your eyes, it’s easier to feel the pendulum effect and the movement of your shoulders.

Have a sense of humor. If you take a horrible shot, laugh about it instead of getting angry or sulking. While golf is a fairly serious game, having a bad attitude can affect more than just your mood. When you are upset or angry, you will lose focus, leading to a downward spiral. Instead, brush off the bad shot, and concentrate on having a good time on the course.

Sensible Golf Products – Some New Guidance

Once you have taught yourself to consistently hit the ball, next is to get your swing down right. Think of the club as the pendulum in a grandfather clock. Practice swinging your club from side to side like the pendulum. Remember, it doesn’t take a lot of force to drive the ball far. It takes consistency and accuracy when hitting the ball. People as old as 80 can out drive someone twice their strength because it is all about consistency and accuracy.

As you can see from the practical ideas in the article above, there is no reason to have your golf game looking old and boring. When you start using the information from the article above, your game will have a whole new look, and you will find that every swing will be fresh.

Golf is one of the most popular leisure activities in the world. Playing a round of golf is a great way to have fun, challenge yourself and get some exercise all at the same time. There is nothing better than spending a beautiful, sunny day out on the course. This article contains a few hot tips to help improve your golf game.

Are you having trouble slicing the ball away from you? Did the ball leave you in the shape of a banana? This is easy to fix. Move 2 inches closer to the ball so that your ball is just an inch away from the hosel. People’s first reaction during a swing is to pull yourself away from the ball. This will be corrected by moving up on the hosel.

Updates To Consider On Aspects Of Golfing

The sport is actually a very fun and rewarding activity, having numerous health benefits. Golf is a sport anyone can do, you just need to get out there. Read this article for some tips on doing just that.

There is no magic formula, so you will need to find out for yourself what works well for you. Based on a variety of physical attributes, finding your proper stance may look much different than the best stance for other golfers you play with. Taking the time to perfect your stance will greatly lower your average score.

If you want to Golfing improve the quality of your swing you should learn to put the full force of your body into the motion. Amateurs often think that power comes from the arms alone. The reality is that if you rely solely on your arms, your swing will be weak and awkward. It’s better to use the entire body along with the motion from your club.

Make sure that you are holding the club correctly when you are a beginner. The most common problem when trying to hit the ball far is they tend to have a hard grip on the club. The best grip is actually soft, yet firm. Hold your club similar to how you would a bird.

Because of this, go through some stretches before you tee off, and also stay well hydrated. When you’re in good shape, your game will improve.

Before you decide to enter on the course, get a set of golf footwear. Make it a point to read several product reviews for golfing shoes well before you actually buy. The cheapest golfing footwear ought to be avoided, as they often have negative review articles.

Each golf club is made with a “sweet spot,” which is the exact location on the club that provides you the most accuracy and distance if you strike the ball at that point. Take the time to recognize where this ‘sweet spot’ can be found on each of your clubs in order to maximize your game.

Always focus all of your concentration on the next shot. Don’t let the outcome of the previous shot, or any upcoming challenges, influence the way you swing that club. Anxiety about earlier poor plays can really have an effect on your game, so do your best to push them from your mind completely, rather than dwelling on them.

Before you take your shot, make sure that you align the rear of your forward facing foot with the ball. For most of your shots, you will want your feet to align with the ball. Imagine that you are making a triangle with your feet and the ball acts as the top of the pyramid. The only exception to this is when your ball lies on a slope.

Transfer your weight from your back foot to your front foot by rapidly swinging through your hips after assuming your golf stance. This will increase the power of your swing and the distance the ball will travel.

Nearly anyone can enjoy a good game of golf, but only people who take golf seriously will strive to make their swing perfect. By adding these tips to your golf repertoire, you will become an expert golfer in no time.

Prudent Golf Systems – Straightforward Ideas

Golf Golfing is wonderful pastime that is enjoyed by everyone of all ages. Although players come in all shapes in sizes, there is some advice that will make anyone a better golfer. The following article will offer plenty of great advice that just about anyone who loves golf can benefit from.

You can implement this subtle research into your own style, which helps create a stance and methodology that is perfect for you. The right stance can vary a lot depending on your body type, weight, height and gender. A good stance will do a lot for your game.

If you have the opportunity, ask a professional his or her opinion on a club, or certainly on a set of new clubs you are considering purchasing. They will have intimate knowledge of all the clubs available, being able to select for you, exactly what you need for the best shot possible.

An easy exercise, a “toe-wiggling” exercise, can show golf stance faults. Anyone who is leaning inward too far will have difficulty wiggling their toes. Lean back until your feet are loose to ensure you have the correct posture.

Before you decide to set foot on the course, get a set of golf footwear. Make sure you read through several review articles about golfing footwear before you actually buy. That way you’re going to get a high quality set.

Make sure you stretch and stay hydrated. Giving proper attention to your body will help you make improvements in your golf game.

There is a reason that the game of golf has endured for generations. Now you just need to work it into your game.

Quick Products Of Golf – Ideas

Solutions For Golfing – A Helpful Breakdown


Thoughtful Parenting: Applied Behavior Analysis useful therapeutic tool |

Thoughtful Parenting: Applied Behavior Analysis useful therapeutic tool

Deirdre Pepin/For Steamboat Today

Thoughtful Parenting First Impressions

Applied Behavior Analysis, the science of behavior change, is a proven effective method for changing undesirable behavior. Behavior analysis is an approach to understanding behavior and its relationship with the environment.

Behavior refers to a person's actions and skills. Environment refers to anything that has the power to change or be changed by a person's behavior — it can be psychological, emotional, or physical. At the heart of behavior analysis is understanding how learning happens.

Applied Behavior Analysis, or ABA, uses techniques to reduce inappropriate or unwanted behavior and increase appropriate or wanted behavior. ABA can take place in a structured classroom setting, as well as informal real life situations. It can help children and adults improve communication, relationships, school and job performance and participation in family and community events.

Studies show ABA is helpful for children with autism spectrum disorders in acquiring language, self-help and play skills and decreasing aggressive, self-injurious and self-stimulatory behavior. Preschool children with autism spectrum disorders who engage in early intensive ABA for at least two years benefit from substantial gains and may need little or no support as they continue through their school years.

ABA targets the improvement of socially significant behaviors, including communication, social skills, academics, reading, gross and fine motor skills, toileting, dressing, eating, personal self-care, domestic skills and work skills. But ABA is not only for people with developmental disabilities or autism spectrum disorders. Its principles can be used to alter any observable behavior. While ABA can be used as a treatment for anxiety, addiction and ADHD, it can also be used for professional development by enhancing productivity and engagement, for example.

If you are a parent of a child with challenging or inappropriate behaviors, Applied Behavior Analysis might be a great option. It's important to remember, however, that there is no single plan or program that constitutes ABA. There is no single study that says how many sessions it will take to change your child's behavior.

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The most successful ABA programs are those that are woven into the fabric of the child's and family's lifestyle. As the parent of a child in ABA, your role is critical. You know your child better than anyone else, and you experience your child's behavior frequently enough to track it (helping to determine why or when it occurs). Engaged parents ensure that prompts and reinforcements continue throughout daily routines and are fundamental to a successful program.

The first step for parents who want to start ABA is to get an assessment of the child's current skill level. Assessments provide a baseline for the ABA provider and help measure progress going forward. Options include a diagnostic assessment, developmental assessment, domain specific assessment, neuropsychological assessment, criterion-referenced assessment, preference assessment, functional behavioral assessment, and skill probes.

As your local community centered board, Horizons hopes to continue providing information about different kinds of behavior services. ABA is but one of many; others include social cognition, synergetic play therapy and cognitive behavioral therapy. While ABA has attracted attention as the best method for children with autism spectrum disorders, each child's unique situation requires individual evaluation and treatment.

Currently, ABA is in high demand in Northwest Colorado, creating the need for more local providers. Behavior services, not just ABA, should be available through insurance or Medicaid (you do not need to be on a Medicaid Waiver to receive behavior services). Some schools offer behavior services as part of a child's individualized education program.

If you have questions about your infant or toddler's development, call Horizons Specialized Services' Early Intervention office at 970-871-8558.

Deirdre Pepin works in resource development and public relations at Horizons Specialized Services.