PMCID string | Title string | Sentences string |
|---|---|---|
PMC10589262 | Identification of novel pathogenic roles of BLZF1/ATF6 in tumorigenesis of gastrointestinal stromal tumor showing Golgi-localized mutant KIT | One-way ANOVA with a post-hoc test was performed to compare multiple means (*p < 0.05, **p < 0.01, ***p < 0.001). |
PMC10589262 | Identification of novel pathogenic roles of BLZF1/ATF6 in tumorigenesis of gastrointestinal stromal tumor showing Golgi-localized mutant KIT | Error bars in G and H represent the standard deviation (SD) of the mean of three independent western blots. |
PMC10589262 | Identification of novel pathogenic roles of BLZF1/ATF6 in tumorigenesis of gastrointestinal stromal tumor showing Golgi-localized mutant KIT | A, B Immunocytochemistry (ICC) analysis of KIT expression in gastrointestinal stromal tumor (GIST) (GIST430 and GIST882) and colon cancer (CC) cells (DLD-1 and Colo320DM) with or without permeabilization using triton X-100. |
PMC10589262 | Identification of novel pathogenic roles of BLZF1/ATF6 in tumorigenesis of gastrointestinal stromal tumor showing Golgi-localized mutant KIT | C ICC analysis of KIT and calnexin (endoplasmic reticulum (ER) marker) in GIST cells. |
PMC10589262 | Identification of novel pathogenic roles of BLZF1/ATF6 in tumorigenesis of gastrointestinal stromal tumor showing Golgi-localized mutant KIT | D ICC analysis of KIT, GM130 (cis-Golgi), mannosidase ll (medial-Golgi), and Golgin-97 (trans-Golgi) in GIST cells. |
PMC10589262 | Identification of novel pathogenic roles of BLZF1/ATF6 in tumorigenesis of gastrointestinal stromal tumor showing Golgi-localized mutant KIT | E Lysates from GIST and CC cells were deglycosylated using endoglycosidase H (Endo-H) or PNGase-F, and analyzed by western blotting. |
PMC10589262 | Identification of novel pathogenic roles of BLZF1/ATF6 in tumorigenesis of gastrointestinal stromal tumor showing Golgi-localized mutant KIT | F Biotinylation, streptavidin pull-down, and western blotting were performed using GIST and CC cells. |
PMC10589262 | Identification of novel pathogenic roles of BLZF1/ATF6 in tumorigenesis of gastrointestinal stromal tumor showing Golgi-localized mutant KIT | Because the surface expression of MT-KIT was low, twice the amount of biotin-labeled lysates was used for GIST cells than in CC cell lysates. |
PMC10589262 | Identification of novel pathogenic roles of BLZF1/ATF6 in tumorigenesis of gastrointestinal stromal tumor showing Golgi-localized mutant KIT | G Quantification of the band intensities for the western blots as shown in F. H Stem cell factor (SCF)-treatment-mediated KIT degradation was measured by western blot lysates in GIST and CC cells. |
PMC10589262 | Identification of novel pathogenic roles of BLZF1/ATF6 in tumorigenesis of gastrointestinal stromal tumor showing Golgi-localized mutant KIT | Band intensities of the western blots were quantified. |
PMC10589262 | Identification of novel pathogenic roles of BLZF1/ATF6 in tumorigenesis of gastrointestinal stromal tumor showing Golgi-localized mutant KIT | One-way ANOVA with a post-hoc test was performed to compare multiple means (*p < 0.05, **p < 0.01, ***p < 0.001). |
PMC10589262 | Identification of novel pathogenic roles of BLZF1/ATF6 in tumorigenesis of gastrointestinal stromal tumor showing Golgi-localized mutant KIT | Error bars in G and H represent the standard deviation (SD) of the mean of three independent western blots. |
PMC10589262 | Identification of novel pathogenic roles of BLZF1/ATF6 in tumorigenesis of gastrointestinal stromal tumor showing Golgi-localized mutant KIT | Although ICC analysis showed Golgi localization of MT-KIT, membranous MT-KIT expression in GISTs and CCs was quantitatively measured by biotin labeling and western blot analysis of PM proteins. |
PMC10589262 | Identification of novel pathogenic roles of BLZF1/ATF6 in tumorigenesis of gastrointestinal stromal tumor showing Golgi-localized mutant KIT | Since the membranous MT-KIT expression in GIST cells was too low, the amount of GIST whole-cell lysate used for biotin-labeled protein pulldown was doubled than that in CCs. |
PMC10589262 | Identification of novel pathogenic roles of BLZF1/ATF6 in tumorigenesis of gastrointestinal stromal tumor showing Golgi-localized mutant KIT | The ratios between membranous KIT levels compared with whole-cell lysates were approximately 0.40 and 0.20 for GIST430 and GIST882 cells, respectively, while those for CC cells were approximately 1.0 (Fig. 1F, G). |
PMC10589262 | Identification of novel pathogenic roles of BLZF1/ATF6 in tumorigenesis of gastrointestinal stromal tumor showing Golgi-localized mutant KIT | The surface expression of MT-KIT in GIST430 cells was higher than in GIST882 cells due to the heterozygous mutation in GIST430 cells . |
PMC10589262 | Identification of novel pathogenic roles of BLZF1/ATF6 in tumorigenesis of gastrointestinal stromal tumor showing Golgi-localized mutant KIT | Despite a small fraction of the total MT-KIT, weak membranous KIT expression was detected in GIST cells (Fig. 1F). |
PMC10589262 | Identification of novel pathogenic roles of BLZF1/ATF6 in tumorigenesis of gastrointestinal stromal tumor showing Golgi-localized mutant KIT | Therefore, a ligand tolerance assay was performed to test if the membranous MT-KIT responds to its ligand, the stem cell factor (SCF). |
PMC10589262 | Identification of novel pathogenic roles of BLZF1/ATF6 in tumorigenesis of gastrointestinal stromal tumor showing Golgi-localized mutant KIT | As previously reported , SCF treatment rapidly downregulated WT-KIT expression in CC cells. |
PMC10589262 | Identification of novel pathogenic roles of BLZF1/ATF6 in tumorigenesis of gastrointestinal stromal tumor showing Golgi-localized mutant KIT | However, MT-KIT levels barely changed in GIST cells following SCF treatment (Fig. 1H). |
PMC10589262 | Identification of novel pathogenic roles of BLZF1/ATF6 in tumorigenesis of gastrointestinal stromal tumor showing Golgi-localized mutant KIT | The fate of membranous MT-KIT was tracked in a time-dependent manner to understand its role in GIST tumorigenesis. |
PMC10589262 | Identification of novel pathogenic roles of BLZF1/ATF6 in tumorigenesis of gastrointestinal stromal tumor showing Golgi-localized mutant KIT | GIST882 cells harboring the homozygous KIT mutation were used for analysis. |
PMC10589262 | Identification of novel pathogenic roles of BLZF1/ATF6 in tumorigenesis of gastrointestinal stromal tumor showing Golgi-localized mutant KIT | GIST882 cells were labeled with a fluorescent-dye-conjugated KIT antibody and examined over time using a confocal microscope. |
PMC10589262 | Identification of novel pathogenic roles of BLZF1/ATF6 in tumorigenesis of gastrointestinal stromal tumor showing Golgi-localized mutant KIT | Membranous MT-KIT was internalized within 15 min and disappeared within 1 h. Colocalization was not observed between MT-KIT and a Golgi complex marker, GM130, until MT-KIT disappeared, which excludes the possibility that Golgi retention of MT-KIT is mediated by the endosome-to-trans-Golgi retrieval pathway, a reported Golgi retention mechanism of the G-protein-coupled receptor (Supplementary Fig. S3A). |
PMC10589262 | Identification of novel pathogenic roles of BLZF1/ATF6 in tumorigenesis of gastrointestinal stromal tumor showing Golgi-localized mutant KIT | Next, the involvement of PM quality-control (PMQC) in the disappearance of MT-KIT was investigated. |
PMC10589262 | Identification of novel pathogenic roles of BLZF1/ATF6 in tumorigenesis of gastrointestinal stromal tumor showing Golgi-localized mutant KIT | The labeling experiment was repeated to evaluate the colocalization of MT-KIT with an early endosome marker, EEA1, or a lysosomal marker, LAMP1, involved in the PMQC processes . |
PMC10589262 | Identification of novel pathogenic roles of BLZF1/ATF6 in tumorigenesis of gastrointestinal stromal tumor showing Golgi-localized mutant KIT | Colocalization of membranous MT-KIT with EEA1 and LAMP1 was clearly observed 15 and 30 min after tracking, and disappeared after 1 h (Supplementary Fig. S3B, C). |
PMC10589262 | Identification of novel pathogenic roles of BLZF1/ATF6 in tumorigenesis of gastrointestinal stromal tumor showing Golgi-localized mutant KIT | To quantitatively validate the ICC results, western blot analysis of membranous KIT was performed in GISTs and CCs. |
PMC10589262 | Identification of novel pathogenic roles of BLZF1/ATF6 in tumorigenesis of gastrointestinal stromal tumor showing Golgi-localized mutant KIT | In GISTs, the remaining membranous MT-KIT levels was measured through biotin-labeled protein pulldown assay and western blot at the indicated time-points. |
PMC10589262 | Identification of novel pathogenic roles of BLZF1/ATF6 in tumorigenesis of gastrointestinal stromal tumor showing Golgi-localized mutant KIT | Bafilomycin A1 (BafA1), a lysosomal inhibitor, was used to validate lysosome-mediated protein degradation. |
PMC10589262 | Identification of novel pathogenic roles of BLZF1/ATF6 in tumorigenesis of gastrointestinal stromal tumor showing Golgi-localized mutant KIT | CCs were included as controls to determine the original stability of WT-KIT. |
PMC10589262 | Identification of novel pathogenic roles of BLZF1/ATF6 in tumorigenesis of gastrointestinal stromal tumor showing Golgi-localized mutant KIT | Since WT-KIT is mostly expressed in the PM, the remaining WT-KIT abundance was measured at the indicated time-points after cycloheximide (CHX) treatment. |
PMC10589262 | Identification of novel pathogenic roles of BLZF1/ATF6 in tumorigenesis of gastrointestinal stromal tumor showing Golgi-localized mutant KIT | After 4 h, >50% of the original WT-KIT level remained (Supplementary Fig. S4A), whereas in GIST882 cells, 8% remained. |
PMC10589262 | Identification of novel pathogenic roles of BLZF1/ATF6 in tumorigenesis of gastrointestinal stromal tumor showing Golgi-localized mutant KIT | MT-KIT degradation was efficiently blocked by BafA1 treatment (Supplementary Fig. S4B). |
PMC10589262 | Identification of novel pathogenic roles of BLZF1/ATF6 in tumorigenesis of gastrointestinal stromal tumor showing Golgi-localized mutant KIT | These findings suggest that membranous MT-KIT is hardly involved in GIST tumorigenesis due to its low membranous protein level caused by Golgi retention and plasma membrane quality control (PMQC). |
PMC10589262 | Identification of novel pathogenic roles of BLZF1/ATF6 in tumorigenesis of gastrointestinal stromal tumor showing Golgi-localized mutant KIT | Because of low protein levels and rapid degradation of membranous MT-KIT, we hypothesized that the Golgi-localized MT-KIT is the primary contributor to sustained activation of downstream signaling in GISTs. |
PMC10589262 | Identification of novel pathogenic roles of BLZF1/ATF6 in tumorigenesis of gastrointestinal stromal tumor showing Golgi-localized mutant KIT | We first aimed to exclude the possibility that ER- or PM-localized MT-KIT initiates downstream signaling. |
PMC10589262 | Identification of novel pathogenic roles of BLZF1/ATF6 in tumorigenesis of gastrointestinal stromal tumor showing Golgi-localized mutant KIT | Brefeldin A (BFA) was used to block ER-to-Golgi trafficking, thereby inducing ER retention of MT-KIT. |
PMC10589262 | Identification of novel pathogenic roles of BLZF1/ATF6 in tumorigenesis of gastrointestinal stromal tumor showing Golgi-localized mutant KIT | ICC analysis showed that BFA led to ER retention of MT-KIT (Supplementary Fig. S5). |
PMC10589262 | Identification of novel pathogenic roles of BLZF1/ATF6 in tumorigenesis of gastrointestinal stromal tumor showing Golgi-localized mutant KIT | Western blot analysis showed that BFA dramatically reduced phospho-KIT, phospho-AKT, and phospho-ERK levels, indicating that ER-retained MT-KIT was insufficient to activate downstream signaling (Fig. 2A). |
PMC10589262 | Identification of novel pathogenic roles of BLZF1/ATF6 in tumorigenesis of gastrointestinal stromal tumor showing Golgi-localized mutant KIT | On the other hand, the levels of phospho-AKT and phospho-ERK were minimally affected by BFA treatment in CC cells, whereas KIT was retained in the ER (Supplementary Fig. S6A). |
PMC10589262 | Identification of novel pathogenic roles of BLZF1/ATF6 in tumorigenesis of gastrointestinal stromal tumor showing Golgi-localized mutant KIT | The involvement of membranous MT-KIT in downstream signaling was also investigated using 30N12, an inhibitor of trans-Golgi-to-PM trafficking . |
PMC10589262 | Identification of novel pathogenic roles of BLZF1/ATF6 in tumorigenesis of gastrointestinal stromal tumor showing Golgi-localized mutant KIT | To validate the effects of 30N12, western blot and ICC analysis of WT-KIT was performed in SCF-treated CC cells treated with or without 30N12. |
PMC10589262 | Identification of novel pathogenic roles of BLZF1/ATF6 in tumorigenesis of gastrointestinal stromal tumor showing Golgi-localized mutant KIT | Treatment with 30N12 efficiently induced WT-KIT retention in the Golgi complex, and reduced phospho-KIT, phospho-AKT, and phospho-ERK levels, that were upregulated by SCF treatment (Supplementary Fig. S6B, C). |
PMC10589262 | Identification of novel pathogenic roles of BLZF1/ATF6 in tumorigenesis of gastrointestinal stromal tumor showing Golgi-localized mutant KIT | However, in GIST cells, 30N12 treatment did not change the phosphorylation levels of effector molecules, indicating that PM-localized MT-KIT was also insufficient to activate downstream signaling (Fig. 2B).Fig. |
PMC10589262 | Identification of novel pathogenic roles of BLZF1/ATF6 in tumorigenesis of gastrointestinal stromal tumor showing Golgi-localized mutant KIT | 2Golgi complex is the main site where MT-KIT initiates downstream oncogenic signaling. |
PMC10589262 | Identification of novel pathogenic roles of BLZF1/ATF6 in tumorigenesis of gastrointestinal stromal tumor showing Golgi-localized mutant KIT | A The status of downstream effector molecules in the KIT signaling pathway was analyzed by western blotting GIST cells with or without treatment with Brefeldin A (BFA, 5 µg/mL) for 4 h, a compound that blocks ER to Golgi trafficking. |
PMC10589262 | Identification of novel pathogenic roles of BLZF1/ATF6 in tumorigenesis of gastrointestinal stromal tumor showing Golgi-localized mutant KIT | B The same analysis was performed as in A using GIST cells with or without treatment for 18 h with 1 or 10 µM 30N12, a compound that blocks trans-Golgi trafficking to the plasma membrane (PM). |
PMC10589262 | Identification of novel pathogenic roles of BLZF1/ATF6 in tumorigenesis of gastrointestinal stromal tumor showing Golgi-localized mutant KIT | C Immunoprecipitation of GIST430 and GIST882 cell lysates was performed to evaluate the interaction of MT-KIT with P85 and GRB2, the most upstream molecules of the PI3K/AKT pathway and MAPK/ERK pathway, respectively. |
PMC10589262 | Identification of novel pathogenic roles of BLZF1/ATF6 in tumorigenesis of gastrointestinal stromal tumor showing Golgi-localized mutant KIT | D, E Confocal microscopic analysis of KIT, GRB2, P85, and GM130 in GIST cells transfected with HA-GRB2 or HA-P85 expression vectors was performed. |
PMC10589262 | Identification of novel pathogenic roles of BLZF1/ATF6 in tumorigenesis of gastrointestinal stromal tumor showing Golgi-localized mutant KIT | HA-GRB2 and HA-P85 were labeled with hemagglutinin (HA). |
PMC10589262 | Identification of novel pathogenic roles of BLZF1/ATF6 in tumorigenesis of gastrointestinal stromal tumor showing Golgi-localized mutant KIT | A The status of downstream effector molecules in the KIT signaling pathway was analyzed by western blotting GIST cells with or without treatment with Brefeldin A (BFA, 5 µg/mL) for 4 h, a compound that blocks ER to Golgi trafficking. |
PMC10589262 | Identification of novel pathogenic roles of BLZF1/ATF6 in tumorigenesis of gastrointestinal stromal tumor showing Golgi-localized mutant KIT | B The same analysis was performed as in A using GIST cells with or without treatment for 18 h with 1 or 10 µM 30N12, a compound that blocks trans-Golgi trafficking to the plasma membrane (PM). |
PMC10589262 | Identification of novel pathogenic roles of BLZF1/ATF6 in tumorigenesis of gastrointestinal stromal tumor showing Golgi-localized mutant KIT | C Immunoprecipitation of GIST430 and GIST882 cell lysates was performed to evaluate the interaction of MT-KIT with P85 and GRB2, the most upstream molecules of the PI3K/AKT pathway and MAPK/ERK pathway, respectively. |
PMC10589262 | Identification of novel pathogenic roles of BLZF1/ATF6 in tumorigenesis of gastrointestinal stromal tumor showing Golgi-localized mutant KIT | D, E Confocal microscopic analysis of KIT, GRB2, P85, and GM130 in GIST cells transfected with HA-GRB2 or HA-P85 expression vectors was performed. |
PMC10589262 | Identification of novel pathogenic roles of BLZF1/ATF6 in tumorigenesis of gastrointestinal stromal tumor showing Golgi-localized mutant KIT | HA-GRB2 and HA-P85 were labeled with hemagglutinin (HA). |
PMC10589262 | Identification of novel pathogenic roles of BLZF1/ATF6 in tumorigenesis of gastrointestinal stromal tumor showing Golgi-localized mutant KIT | To investigate whether Golgi-retained MT-KIT primarily contributes to downstream oncogenic signaling activation, we investigated the interaction of MT-KIT with P85 and GRB2, the upstream effectors of the PI3K/AKT and MAPK/ERK pathways, respectively. |
PMC10589262 | Identification of novel pathogenic roles of BLZF1/ATF6 in tumorigenesis of gastrointestinal stromal tumor showing Golgi-localized mutant KIT | Immunoprecipitation assays were performed using GIST cell lysates incubated with IgG (control) or KIT antibody. |
PMC10589262 | Identification of novel pathogenic roles of BLZF1/ATF6 in tumorigenesis of gastrointestinal stromal tumor showing Golgi-localized mutant KIT | This analysis showed that the endogenous MT-KIT bound to both P85 and GRB2 (Fig. 2C). |
PMC10589262 | Identification of novel pathogenic roles of BLZF1/ATF6 in tumorigenesis of gastrointestinal stromal tumor showing Golgi-localized mutant KIT | The interaction of MT-KIT with GRB2 and P85 was further verified through ICC analysis. |
PMC10589262 | Identification of novel pathogenic roles of BLZF1/ATF6 in tumorigenesis of gastrointestinal stromal tumor showing Golgi-localized mutant KIT | Since there are no commercial antibodies for ICC analysis of P85 and GRB2, HA-tagged P85 and GRB2 expression vectors were constructed. |
PMC10589262 | Identification of novel pathogenic roles of BLZF1/ATF6 in tumorigenesis of gastrointestinal stromal tumor showing Golgi-localized mutant KIT | After vector transfection into GIST cells, cellular localization of MT-KIT, HA-P85, and HA-GRB2 were examined. |
PMC10589262 | Identification of novel pathogenic roles of BLZF1/ATF6 in tumorigenesis of gastrointestinal stromal tumor showing Golgi-localized mutant KIT | GRB2 and P85 clearly showed colocalization with MT-KIT and GM130 (Fig. 2D, E). |
PMC10589262 | Identification of novel pathogenic roles of BLZF1/ATF6 in tumorigenesis of gastrointestinal stromal tumor showing Golgi-localized mutant KIT | Quantification of immunofluorescence intensity further demonstrated that GRB2 and P85 colocalize with MT-KIT and GM130 at the Golgi complex (Supplementary Fig. S7). |
PMC10589262 | Identification of novel pathogenic roles of BLZF1/ATF6 in tumorigenesis of gastrointestinal stromal tumor showing Golgi-localized mutant KIT | The localization of GRB2 and P85 after SCF treatment in CC cells was further investigated to compare the PI3K/AKT and MAPK/ERK pathway activation mediated by WT-KIT and MT-KIT. |
PMC10589262 | Identification of novel pathogenic roles of BLZF1/ATF6 in tumorigenesis of gastrointestinal stromal tumor showing Golgi-localized mutant KIT | In contrast to GIST cells, ICC analysis showed that SCF treatment recruited P85 and GRB2 to the peri-PM region, where activated WT-KIT was localized (Supplementary Fig. S8). |
PMC10589262 | Identification of novel pathogenic roles of BLZF1/ATF6 in tumorigenesis of gastrointestinal stromal tumor showing Golgi-localized mutant KIT | These data showed that MT-KIT directly initiates downstream signals from the Golgi complex, whereas WT-KIT initiates signals from the PM. |
PMC10589262 | Identification of novel pathogenic roles of BLZF1/ATF6 in tumorigenesis of gastrointestinal stromal tumor showing Golgi-localized mutant KIT | Since the Golgi complex is a hub for constant protein trafficking, a complementary mechanism is necessary for the retention of MT-KIT. |
PMC10589262 | Identification of novel pathogenic roles of BLZF1/ATF6 in tumorigenesis of gastrointestinal stromal tumor showing Golgi-localized mutant KIT | Therefore, we aimed to identify a possible MT-KIT-binding partner from the Golgi-related proteins that could tether MT-KIT to the Golgi complex during trafficking. |
PMC10589262 | Identification of novel pathogenic roles of BLZF1/ATF6 in tumorigenesis of gastrointestinal stromal tumor showing Golgi-localized mutant KIT | Since there are only a few reported Golgi-related proteins [13–15], all possible candidates with commercially available antibodies were selected. |
PMC10589262 | Identification of novel pathogenic roles of BLZF1/ATF6 in tumorigenesis of gastrointestinal stromal tumor showing Golgi-localized mutant KIT | Western blot was performed on ten Golgi proteins (GBF1, GM130, Golgin97, TGN38, GRASP65, GRASP55, BLZF1, STX3, STX6, and GOLPH3) in small-cell lung cancer (SCLC), leukemia, colon cancer, and GIST cell lines expressing WT-KIT or MT-KIT. |
PMC10589262 | Identification of novel pathogenic roles of BLZF1/ATF6 in tumorigenesis of gastrointestinal stromal tumor showing Golgi-localized mutant KIT | All Golgi proteins showed variable levels in the cell lines, except for BLZF1 that selectively showed high protein levels in GIST 430 and GIST 882 cells (Fig. 3A). |
PMC10589262 | Identification of novel pathogenic roles of BLZF1/ATF6 in tumorigenesis of gastrointestinal stromal tumor showing Golgi-localized mutant KIT | BLZF1 expression was also similarly high in imatinib-resistant GIST430-V654A and GIST48 cells (Supplementary Fig. S9). |
PMC10589262 | Identification of novel pathogenic roles of BLZF1/ATF6 in tumorigenesis of gastrointestinal stromal tumor showing Golgi-localized mutant KIT | Moreover, immunoprecipitation with KIT antibody revealed that MT-KIT specifically bound to BLZF1 (Fig. 3B). |
PMC10589262 | Identification of novel pathogenic roles of BLZF1/ATF6 in tumorigenesis of gastrointestinal stromal tumor showing Golgi-localized mutant KIT | The colocalization of MT-KIT and BLZF1 in the Golgi complex of GIST cells was validated by ICC analysis (Fig. 3C, D). |
PMC10589262 | Identification of novel pathogenic roles of BLZF1/ATF6 in tumorigenesis of gastrointestinal stromal tumor showing Golgi-localized mutant KIT | On the other hand, fluorescence signal of BLZF was barely detected in the Golgi complex of CC and leukemia cell lines (DLD-1 and HMC-1) (Supplementary Fig. S10). |
PMC10589262 | Identification of novel pathogenic roles of BLZF1/ATF6 in tumorigenesis of gastrointestinal stromal tumor showing Golgi-localized mutant KIT | BLZF1 knockdown with short hairpin RNA (shRNA) dramatically downregulated MT-KIT expression, as demonstrated by ICC and western blot analyses (Fig. 3C–E). |
PMC10589262 | Identification of novel pathogenic roles of BLZF1/ATF6 in tumorigenesis of gastrointestinal stromal tumor showing Golgi-localized mutant KIT | BLZF1 knockdown strongly inhibited cell growth in both imatinib-sensitive GIST430 and GIST 882 cells, and in imatinib-resistant GIST430-V654A and GIST48 cells (Fig. 3F and Supplementary Fig. S11). |
PMC10589262 | Identification of novel pathogenic roles of BLZF1/ATF6 in tumorigenesis of gastrointestinal stromal tumor showing Golgi-localized mutant KIT | These findings collectively suggest that BLZF1 is required for Golgi retention and stable expression of MT-KIT, and consequently for GIST cell growth. |
PMC10589262 | Identification of novel pathogenic roles of BLZF1/ATF6 in tumorigenesis of gastrointestinal stromal tumor showing Golgi-localized mutant KIT | Fig. 3BLZF1 is a novel KIT binding partner, which is indispensable for tethering MT-KIT in the Golgi complex. |
PMC10589262 | Identification of novel pathogenic roles of BLZF1/ATF6 in tumorigenesis of gastrointestinal stromal tumor showing Golgi-localized mutant KIT | A Western botting was performed using antibodies against ten Golgi-related proteins in small cell lung carcinoma (SCLC), leukemia, CC, and GIST cell lines expressing WT-KIT or MT-KIT. |
PMC10589262 | Identification of novel pathogenic roles of BLZF1/ATF6 in tumorigenesis of gastrointestinal stromal tumor showing Golgi-localized mutant KIT | B Immunoprecipitation analysis was performed in GIST cells using a KIT antibody and binding between KIT and Golgi-related proteins was evaluated by western blotting. |
PMC10589262 | Identification of novel pathogenic roles of BLZF1/ATF6 in tumorigenesis of gastrointestinal stromal tumor showing Golgi-localized mutant KIT | C, D Confocal microscopic analysis of GM130, BLZF1, and KIT in GIST cells with or without BLZF1 shRNA treatment. |
PMC10589262 | Identification of novel pathogenic roles of BLZF1/ATF6 in tumorigenesis of gastrointestinal stromal tumor showing Golgi-localized mutant KIT | E Western blotting analysis of KIT and BLZF1 in GIST cells with or without BLZF1 shRNA treatment. |
PMC10589262 | Identification of novel pathogenic roles of BLZF1/ATF6 in tumorigenesis of gastrointestinal stromal tumor showing Golgi-localized mutant KIT | F A proliferation assay was performed in GIST cells with or without BLZF1 shRNA treatment. |
PMC10589262 | Identification of novel pathogenic roles of BLZF1/ATF6 in tumorigenesis of gastrointestinal stromal tumor showing Golgi-localized mutant KIT | Cell growth was measured by MTT analysis 72 h after BLZF1 shRNA treatment. |
PMC10589262 | Identification of novel pathogenic roles of BLZF1/ATF6 in tumorigenesis of gastrointestinal stromal tumor showing Golgi-localized mutant KIT | Error bars in F represents the SD of the mean of three independent experiments. |
PMC10589262 | Identification of novel pathogenic roles of BLZF1/ATF6 in tumorigenesis of gastrointestinal stromal tumor showing Golgi-localized mutant KIT | One-way ANOVA with a post-hoc test was performed to compare multiple means (*p < 0.05, **p < 0.01, ***p < 0.001). |
PMC10589262 | Identification of novel pathogenic roles of BLZF1/ATF6 in tumorigenesis of gastrointestinal stromal tumor showing Golgi-localized mutant KIT | A Western botting was performed using antibodies against ten Golgi-related proteins in small cell lung carcinoma (SCLC), leukemia, CC, and GIST cell lines expressing WT-KIT or MT-KIT. |
PMC10589262 | Identification of novel pathogenic roles of BLZF1/ATF6 in tumorigenesis of gastrointestinal stromal tumor showing Golgi-localized mutant KIT | B Immunoprecipitation analysis was performed in GIST cells using a KIT antibody and binding between KIT and Golgi-related proteins was evaluated by western blotting. |
PMC10589262 | Identification of novel pathogenic roles of BLZF1/ATF6 in tumorigenesis of gastrointestinal stromal tumor showing Golgi-localized mutant KIT | C, D Confocal microscopic analysis of GM130, BLZF1, and KIT in GIST cells with or without BLZF1 shRNA treatment. |
PMC10589262 | Identification of novel pathogenic roles of BLZF1/ATF6 in tumorigenesis of gastrointestinal stromal tumor showing Golgi-localized mutant KIT | E Western blotting analysis of KIT and BLZF1 in GIST cells with or without BLZF1 shRNA treatment. |
PMC10589262 | Identification of novel pathogenic roles of BLZF1/ATF6 in tumorigenesis of gastrointestinal stromal tumor showing Golgi-localized mutant KIT | F A proliferation assay was performed in GIST cells with or without BLZF1 shRNA treatment. |
PMC10589262 | Identification of novel pathogenic roles of BLZF1/ATF6 in tumorigenesis of gastrointestinal stromal tumor showing Golgi-localized mutant KIT | Cell growth was measured by MTT analysis 72 h after BLZF1 shRNA treatment. |
PMC10589262 | Identification of novel pathogenic roles of BLZF1/ATF6 in tumorigenesis of gastrointestinal stromal tumor showing Golgi-localized mutant KIT | Error bars in F represents the SD of the mean of three independent experiments. |
PMC10589262 | Identification of novel pathogenic roles of BLZF1/ATF6 in tumorigenesis of gastrointestinal stromal tumor showing Golgi-localized mutant KIT | One-way ANOVA with a post-hoc test was performed to compare multiple means (*p < 0.05, **p < 0.01, ***p < 0.001). |
PMC10589262 | Identification of novel pathogenic roles of BLZF1/ATF6 in tumorigenesis of gastrointestinal stromal tumor showing Golgi-localized mutant KIT | It remains unclear how MT-KIT bypasses ERQC to reach the Golgi complex. |
PMC10589262 | Identification of novel pathogenic roles of BLZF1/ATF6 in tumorigenesis of gastrointestinal stromal tumor showing Golgi-localized mutant KIT | As shown in Fig. 1E, MT-KIT in GISTs was primarily in the post-ER form, which indicates that MT-KIT is folded enough to avoid ER accumulation and stress induction. |
PMC10589262 | Identification of novel pathogenic roles of BLZF1/ATF6 in tumorigenesis of gastrointestinal stromal tumor showing Golgi-localized mutant KIT | Since GIST cells are constantly exposed to nutrient deficiency and hypoxia that cause ER stress. |
PMC10589262 | Identification of novel pathogenic roles of BLZF1/ATF6 in tumorigenesis of gastrointestinal stromal tumor showing Golgi-localized mutant KIT | Therefore, we hypothesized that GISTs might have acquired a UPR-related intrinsic mechanism to relieve ER stress during tumorigenesis. |
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