TRIM28 is a transcriptional activator of the mutant TERT promoter in human bladder cancer
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ABSTRACT:
PROVIDER: PRJNA752104 | ENA |
REPOSITORIES: ENA
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Project description:Human telomerase reverse transcriptase (hTERT) is a rate-limiting catalytic subunit of the enzyme telomerase, which plays a central role in maintaining replicative immortality in normal stem cells and many cancer cells. Mostly silenced in normal somatic cells, hTERT is reactivated in cancer commonly through highly recurrent hot spot point mutations in the hTERT promoter. Bladder cancer has a 60-70% hTERT promoter mutation prevalence, commonly at -124 bp from the translation start site, and this is associated with increased hTERT expression and poor patient prognosis. Identifying targetable signaling pathways that drive hTERT transcription from the mutant promoter could lead to therapies with reduced effects on normal progenitors. Here we describe a new platform that probes endogenous hTERT regulation by allele-specific insertion of GFP at either wild-type or mutant promoter allele within human bladder cancer cells harboring a heterozygous mutation. Using cells expressing the GFP-hTERT fusion protein, we performed a pooled CRISPR-Cas9 Kinome KO genetic screen to identify regulators of mutant hTERT promoter activity. We identified TRIM28 as an activator and TRIM24 as a suppressor of hTERT expression. Similar to hTERT, TRIM28 expression stratifies patient outcome and inhibits promotes cell growth both in vitro and in vivo, supporting their clinical relevance. TRIM28 and TRIM24 get recruited to the mutated promoter through GABPA, where TRIM24 interacts with TRIM28 and inhibits its activity as a transcriptional activator. TRIM28 phosphorylation causes its release from TRIM24 and activates hTERT transcription. mTOR phosphorylates TRIM28 through the mTORC1 complex leading to hTERT expression, while mTORC1 inhibition with rapamycin analog Ridaforolimus suppresses the phosphorylation and promoter binding of TRIM28, resulting in reduced hTERT expression and lower cell viability. This study uses a bespoke platform to identify a novel therapeutically tractable regulatory pathway for cancers harboring mutant hTERT promoters.
2021-08-06 | GSE181461 | GEO
Project description:Point mutations within the TERT promoter are the most common recurrent somatic non-coding mutation identified across different cancer types, including glioblastoma, melanoma, hepatocellular carcinoma and bladder cancer. They are most abundant at C146T and C124T and more rare at A57C, with the latter originally described as a familial case but subsequently shown also to occur somatically. All three mutations create de novo ETS (E-twenty-six specific) binding sites and result in the reactivation of the TERT gene, allowing cancer cells to achieve replicative immortality. Here, we employed a systematic proteomics screen to identify transcription factors preferentially binding to the C146T, C124T and A57C mutations. While we confirmed binding of multiple ETS factors to the mutant C146T and C124T sequences, we identified E4F1 a an A57C-specific binder and ZNF148 as a TERT WT binder that is excluded from the TERT promoter by the C124T allele. Both proteins are activating transcription factors that bind specifically to the A57C and wildtype (at position 124) TERT promoter sequence in corresponding cell lines and upregulate TERT transcription and telomerase activity.
2023-11-04 | PXD037776 | Pride
Project description:Blocking telomerase is recognized as a key anti-cancer mechanism. Unlike in stem cells, levels of telomerase catalytic subunit TERT are limiting in reconstituting telomerase activity in somatic cells. However in some cancers, Tert is transcriptionally reactivated by mutations in its promoter. Given that Tert in stem cells is driven by WT Tert promoter, if we can selectively target Tert reactivation through mutant Tert promoters we can block telomerase activity specifically in cancer cells without toxicity in stem cells. Here we report the epigenetic regulation of Tert promoter comparing WT and mutant promoters. We showed that GABPA homodimerization through long-range interaction stabilizes Gabpa to drive Tert expression. Furthermore, BRD4 specifically activates the C250T mutant promoter via dual mechanism involving GABPA, thereby setting the stage for future therapeutics.
2017-02-27 | GSE77265 | GEO
Project description:A recent study by Castelo-Branco, P., et al. Methylation of the TERT promoter and risk stratification of childhood brain tumours: an integrative genomic and molecular study. Lancet Oncol 2013;14:534-542 found upstream of the transcription start site (UTSS) hypermethylation of TERT is associated with tumor progression and poor prognosis in paediatric brain tumours. They interpreted that the UTSS region of telomerase reverse transcriptase (TERT) gene is a potentially accessible biomarker for various cancers. This study, we aimed to explore the role of TERT in hepatocellular carcinoma (HCC) and to investigate whether the UTSS region of TERT promoter shows the same methylation pattern in HCC. We analyzed a methylation assay of TERT including the UTSS region in 125 paired HCC samples using Mass Array EpiTyper (Sequenom, SanDiego, CA, USA). To obtain the relationship between TERT promoter methylation status and TERT expression level, we analysed a validation group of 12 paired HCC samples and acquired the FPKM value of TERT gene.
2015-01-03 | E-GEOD-63863 | biostudies-arrayexpress
Project description:A recent study by Castelo-Branco, P., et al. Methylation of the TERT promoter and risk stratification of childhood brain tumours: an integrative genomic and molecular study. Lancet Oncol 2013;14:534-542 found upstream of the transcription start site (UTSS) hypermethylation of TERT is associated with tumor progression and poor prognosis in paediatric brain tumours. They interpreted that the UTSS region of telomerase reverse transcriptase (TERT) gene is a potentially accessible biomarker for various cancers. This study, we aimed to explore the role of TERT in hepatocellular carcinoma (HCC) and to investigate whether the UTSS region of TERT promoter shows the same methylation pattern in HCC.
2015-01-03 | GSE63863 | GEO
Project description:Hotspot mutations in the core promoter region of the telomerase reverse transcriptase (TERT) gene have been well established to associate with aggressive clinical characteristics, radioiodine refractory, tumor recurrence and mortality in thyroid cancer. Several E-twenty-six (ETS) transcription factors were reported to selectively bound to the mutant TERT promoter and activated TERT expression. In this study we aimed to investigate whether TERT promoter mutations confer sensitivity to ETS inhibitor YK-4-279 in thyroid cancer cells and whether this inhibitor could be served as a potential therapeutic agent for thyroid cancer. In vitro assays showed that YK-4-279 treatment sharply suppressed cell viability, colony formation, migration and invasion, as well as induced cell cycle arrest and apoptosis in a panel of thyroid cancer cells. The cell viability after YK-4-279 treatment were similar between cell lines harboring mutant and wild-type TERT promoter. Furthermore, YK-4-279 treatment reduced both luciferase activity and mRNA expression of TERT independent of TERT promoter mutation status. Data from RNA-seq further revealed that YK-4-279 significantly affected biological processes including DNA replication and cell cycle. Reduced DNA helicase activity and decreased expression of several helicase genes were observed after YK-4-279 treatment. Moreover, YK-4-279 significantly inhibited tumor growth and induced apoptosis in a xenograft mice model. Thus, ETS inhibitor YK-4-279 suppressed TERT expression and conferred anti-tumor activity in a TERT promoter mutation-independent manner, and it could be a potential agent for the treatment of advanced thyroid cancers.
2021-04-06 | GSE171473 | GEO
Project description:Analysis of total PolII and Ser2-phosphorylated PolII genomic occupancy in control-transfected (WT) or Trim28-knockdown (Trim28-KD) embryonic stem cells. Embryonic stem cells were transfected with luciferase siRNA (WT) or Trim28 siRNA (Trim28 KD) in duplicate experiments. 48 hrs after transfection, cells were crosslinked with formaldehyde, collected, lysed, and chromatin was fragmented with sonication. Total PolII or Ser2-phosphorylated PolII genomic occupancy in WT or Trim28 KD cells were determined by ChIP-seq.
2014-07-28 | E-GEOD-48253 | biostudies-arrayexpress
Project description:Using microcell-mediated chromosome transfer (MMCT) into the mouse melanoma cell line, B16F10, we have previously found that human chromosome 5 carries a gene, or genes, that can negatively regulate TERT expression. To identify the gene responsible for the regulation of TERT transcription, we performed cDNA microarray analysis using parental B16F10 cells, telomerase negative B16F10 microcell hybrids with a human chromosome 5 (B16F10MH5), and its revertant clones (MH5R) with reactivated telomerase. Here we report the identification of PITX1, whose restoration leads to the downregulation of mouse tert (mtert) transcription, as a TERT suppressor gene. Additionally, both human TERT (hTERT) and mouse TERT (mtert) promoter activity can be suppressed by PITX1. We showed that three and one binding sites, respectively, within the hTERT and mtert promoters that express a unique conserved region are responsible for the transcriptional activation of TERT. Furthermore, we showed that PITX1 binds to the TERT promoter both in vitro and in vivo. Thus, PITX1 suppresses TERT transcription through direct binding to the TERT promoter, which ultimately regulates telomerase activity.
2010-09-10 | GSE23216 | GEO
Project description:We assessed methylation landscape near the human TERT promoter in various normal samples and tumor samples, to assess the unique hypermethylation phenotype that exist in human cancers
2018-09-27 | GSE120511 | GEO
Project description:Using microcell-mediated chromosome transfer (MMCT) into the mouse melanoma cell line, B16F10, we have previously found that human chromosome 5 carries a gene, or genes, that can negatively regulate TERT expression. To identify the gene responsible for the regulation of TERT transcription, we performed cDNA microarray analysis using parental B16F10 cells, telomerase negative B16F10 microcell hybrids with a human chromosome 5 (B16F10MH5), and its revertant clones (MH5R) with reactivated telomerase. Here we report the identification of PITX1, whose restoration leads to the downregulation of mouse tert (mtert) transcription, as a TERT suppressor gene. Additionally, both human TERT (hTERT) and mouse TERT (mtert) promoter activity can be suppressed by PITX1. We showed that three and one binding sites, respectively, within the hTERT and mtert promoters that express a unique conserved region are responsible for the transcriptional activation of TERT. Furthermore, we showed that PITX1 binds to the TERT promoter both in vitro and in vivo. Thus, PITX1 suppresses TERT transcription through direct binding to the TERT promoter, which ultimately regulates telomerase activity. We transferred intact human chromosome 5 into mouse melanoma B16F10 cells by microcell fusion. The microcell hybrids (MH5) exhibited suppression of telomerase, we also obtained revertant clones (MH5R) in which telomerase is reactivated. To identify the differentially expressed genes on human chromosome 5, we performed expression microarray analysis using these two clones and parental B16F10 cells.
2010-09-10 | E-GEOD-23216 | biostudies-arrayexpress