ChIP-seq and R-ChIP-seq following BET bromodomain protein loss in HeLa cells
Ontology highlight
ABSTRACT:
ORGANISM(S): Homo sapiens
PROVIDER: GSE141763 | GEO | 2019/12/11
REPOSITORIES: GEO
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Project description:Unperturbed pre-mRNA splicing is crucial for proper expression of eukaryotic genes, but also maintenance of genome integrity. Loss of splicing factors is associated to DNA damage arising through different mechanisms, among which is the accumulation of mutagenic DNA/RNA hybrids (R-loops). RNase H1 recognizes and degrades the RNA moiety within R-loops, and is a commonly used marker for R-loops. Ecdysoneless protein was previously shown to participate in pre-mRNA splicing through its association with the U5 snRNP. In this study we have performed in vivo Targeted DamID using the Dam-RNase H1 construct to map the localization of R-loops throughout the Drosophila genome in control and Ecd-deficient Drosophila wing imaginal discs.
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Project description:Purpose: Characterization of mechanism and vulnerability of BET protein dependency in GBM cells. Methods: ChIP-seq and RNA-seq were performed on GBM cells at different time points following treatment with dBET6, a novel BET protein degrader. The transcriptome responses of parental and JQ1-resistant U87 cells to JQ1 and dBET6 were compared as well. Result: This study reveals crucial functions of BET proteins and provides the rationale and therapeutic merits of targeted degradation of BET proteins by dBET6 in GBM.
2018-05-15 | GSE99171 | GEO
Project description:Purpose: Characterization of mechanism and vulnerability of BET protein dependency in GBM cells. Methods: ChIP-seq and RNA-seq were performed on GBM cells at different time points following treatment with dBET6, a novel BET protein degrader. The transcriptome responses of parental and JQ1-resistant U87 cells to JQ1 and dBET6 were compared as well. Result: This study reveals crucial functions of BET proteins and provides the rationale and therapeutic merits of targeted degradation of BET proteins by dBET6 in GBM.
2018-05-15 | GSE99175 | GEO
Project description:Purpose: Characterization of mechanism and vulnerability of BET protein dependency in GBM cells. Methods: ChIP-seq and RNA-seq were performed on GBM cells at different time points following treatment with dBET6, a novel BET protein degrader. The transcriptome responses of parental and JQ1-resistant U87 cells to JQ1 and dBET6 were compared as well. Result: This study reveals crucial functions of BET proteins and provides the rationale and therapeutic merits of targeted degradation of BET proteins by dBET6 in GBM.
2018-05-15 | GSE99181 | GEO
Project description:RNase H2 is a specialized enzyme that degrades RNA in RNA/DNA hybrids and deficiency of this enzyme causes a severe neuroinflammatory disease, Aicardi Goutières syndrome (AGS). However, the molecular mechanism underlying AGS is still unclear. Here, we show that RNase H2 is associated with a subset of genes, in a transcription-dependent manner where it interacts with RNA Polymerase II. RNase H2 depletion impairs transcription leading to accumulation of R-loops, structures that comprise RNA/DNA hybrids and a displaced DNA strand, mainly associated with short and intronless genes. Importantly, accumulated R-loops are processed by XPG and XPF endonucleases which leads to DNA damage and activation of the immune response, features associated with AGS. Consequently, we uncover a key role for RNase H2 in the transcription of human genes by maintaining R-loop homeostasis. Our results provide insight into the mechanistic contribution of R-loops to AGS pathogenesis.
2022-02-04 | GSE146970 | GEO
Project description:During transcription the nascent RNA can invade the DNA template, forming extended RNA-DNA duplexes (R-loops). Here we employ ChIP-seq in strains expressing or lacking RNase H to map targets of RNase H activity throughout budding yeast genome. In wild-type strains, R-loops were readily detected over the 35S rDNA region transcribed by Pol I and over the 5S rDNA transcribed by Pol III. In strains lacking RNase H activity, R-loops were elevated over other Pol III genes notably tRNAs, SCR1 and U6 snRNA, and were also associated with the cDNAs of endogenous TY1 retrotransposons, which showed increased rates of mobility to the 5?-flanking regions of tRNA genes. Unexpectedly, R-loops were also associated with mitochondrial genes in the absence of RNase H1, but not of RNase H2. Finally, R-loops were detected on highly expressed protein-coding genes in the wild-type, notably over the second exon of spliced ribosomal protein genes. ChIP-seq of RNA-DNA hybrids using antibody S9.6
2014-11-03 | E-GEOD-53420 | biostudies-arrayexpress
Project description:PURPOSE:MYC-amplified medulloblastomas are highly lethal tumors. Bromodomain and extraterminal (BET) bromodomain inhibition has recently been shown to suppress MYC-associated transcriptional activity in other cancers. The compound JQ1 inhibits BET bromodomain-containing proteins, including BRD4. Here, we investigate BET bromodomain targeting for the treatment of MYC-amplified medulloblastoma. EXPERIMENTAL DESIGN:We evaluated the effects of genetic and pharmacologic inhibition of BET bromodomains on proliferation, cell cycle, and apoptosis in established and newly generated patient- and genetically engineered mouse model (GEMM)-derived medulloblastoma cell lines and xenografts that harbored amplifications of MYC or MYCN. We also assessed the effect of JQ1 on MYC expression and global MYC-associated transcriptional activity. We assessed the in vivo efficacy of JQ1 in orthotopic xenografts established in immunocompromised mice. RESULTS:Treatment of MYC-amplified medulloblastoma cells with JQ1 decreased cell viability associated with arrest at G1 and apoptosis. We observed downregulation of MYC expression and confirmed the inhibition of MYC-associated transcriptional targets. The exogenous expression of MYC from a retroviral promoter reduced the effect of JQ1 on cell viability, suggesting that attenuated levels of MYC contribute to the functional effects of JQ1. JQ1 significantly prolonged the survival of orthotopic xenograft models of MYC-amplified medulloblastoma (P < 0.001). Xenografts harvested from mice after five doses of JQ1 had reduced the expression of MYC mRNA and a reduced proliferative index. CONCLUSION:JQ1 suppresses MYC expression and MYC-associated transcriptional activity in medulloblastomas, resulting in an overall decrease in medulloblastoma cell viability. These preclinical findings highlight the promise of BET bromodomain inhibitors as novel agents for MYC-amplified medulloblastoma.
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Project description:Cancer cells are often hypersensitive to the targeting of transcriptional regulators, which may reflect the deregulated gene expression programs that underlie malignant transformation. One of the most prominent transcriptional vulnerabilities in human cancer to emerge in recent years is the bromodomain and extraterminal (BET) family of proteins, which are coactivators that link acetylated transcription factors and histones to the activation of RNA polymerase II. Despite unclear mechanisms underlying the gene specificity of BET protein function, small molecules targeting these regulators preferentially suppress the transcription of cancer-promoting genes. As a consequence, BET inhibitors elicit anticancer activity in numerous malignant contexts at doses that can be tolerated by normal tissues, a finding supported by animal studies and by phase I clinical trials in human cancer patients. In this review, we will discuss the remarkable, and often perplexing, therapeutic effects of BET bromodomain inhibition in cancer.
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Project description:There is increasing interest in inhibitors targeting BET (bromodomain and extra-terminal) proteins because of the association between this family of proteins and cancer progression. BET inhibitors were initially shown to have efficacy in hematologic malignancies; however, a number of studies have now shown that BET inhibitors can also block progression of non-hematologic malignancies. In this Review, we summarize the efficacy of BET inhibitors in select solid tumors; evaluate the role of BET proteins in mediating resistance to current targeted therapies; and consider potential toxicities of BET inhibitors. We also evaluate recently characterized mechanisms of resistance to BET inhibitors; summarize ongoing clinical trials with these inhibitors; and discuss potential future roles of BET inhibitors in patients with solid tumors.
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Project description:Interleukin (IL) 17-producing T helper (T(H)17) cells have been selected through evolution for their ability to control fungal and bacterial infections. It is also firmly established that their aberrant generation and activation results in autoimmune conditions. Using a characterized potent and selective small molecule inhibitor, we show that the bromodomain and extra-terminal domain (BET) family of chromatin adaptors plays fundamental and selective roles in human and murine T(H)17 differentiation from naive CD4(+) T cells, as well as in the activation of previously differentiated T(H)17 cells. We provide evidence that BET controls T(H)17 differentiation in a bromodomain-dependent manner through a mechanism that includes the direct regulation of multiple effector T(H)17-associated cytokines, including IL17, IL21, and GMCSF. We also demonstrate that BET family members Brd2 and Brd4 associate with the Il17 locus in T(H)17 cells, and that this association requires bromodomains. We recapitulate the critical role of BET bromodomains in T(H)17 differentiation in vivo and show that therapeutic dosing of the BET inhibitor is efficacious in mouse models of autoimmunity. Our results identify the BET family of proteins as a fundamental link between chromatin signaling and T(H)17 biology, and support the notion of BET inhibition as a point of therapeutic intervention in autoimmune conditions.
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