BRCA2, ATM, and CDK12 defects differentially shape prostate tumor driver genomics and clinical aggression
Ontology highlight
ABSTRACT:
PROVIDER: EGAS00001004800 | EGA |
REPOSITORIES: EGA
Similar Datasets
Project description:Cyclin-dependent kinases (CDKs) are key regulators of the cell cycle or transcription, and are emerging as promising therapeutic targets in cancer cells. The transcriptional kinase, CDK12, is especially intriguing, as it modulates transcription elongation by phosphorylating the carboxy-terminal domain (CTD) of RNA polymerase (Pol II) with subsequent effects on expression of DNA damage response (DDR) genes. Yet the exact mechanism by which CDK12 regulates this vital process remains unclear. Using the selective inhibitor of CDK12/13, THZ5317 and transient transcriptome sequencing (TT-seq), we sought to capture the dynamic transcriptional changes linked to CDK12 by analyzing the immediate, early effects on nascent RNA production after CDK12 depletion in neuroblastoma (NB) cells. CDK12-induced inhibition resulted in gene length-dependent defects in transcription elongation, leading to a disproportionate loss of 3' reads in long genes. Short transcripts, by contrast, including those of replication-dependent histone genes, underwent 3' UTR extension. The effect on long genes was accompanied by premature cleavage and polyadenylation (PCPA), followed by early termination and loss of gene expression. DDR gene transcripts were the most prominent among those terminated by PCPA. Phosphoproteomic analysis indicated that pre-mRNA processing factors, including those involved in CPA , are direct phospho-targets of CDK12 and that CDK depletion phenocopied their effects on transcription. Importantly, sensitivity to CDK12 inhibition was predicted by a lower proportion of U1 snRNP binding sites compared to polyadenylation sites (PAS) and hence a higher propensity to intronic poly(A) site selection - a characteristic observed in most DDR genes. These results support a model in which CDK12 regulates expression of DDR genes not only by regulating transcription elongation through its effects on Pol II CTD phosphorylation, but also through direct phosphorylation of pre-mRNA processing factors. These mechanistic insights may enable the development of new anti-cancer strategies targeting multiple vulnerable steps in CDK12-dependent regulation of DNA damage repair.
2018-04-18 | MSV000082283 | MassIVE
Project description:Mutations in the tumour suppressor BRCA2 are associated with predisposition to breast and ovarian cancers. BRCA2 has a central role in genome integrity by facilitating the repair of toxic DNA double-strand breaks (DSBs) by homologous recombination. BRCA2 acts by promoting RAD51 nucleofilament formation on resected single-stranded DNA, but how BRCA2 activity is regulated during HR is not fully understood. Here, we delineate a pathway where ATM and ATR kinases phosphorylate a highly conserved region in BRCA2 in response to DNA DSBs. These phosphorylations stimulate the binding of the protein phosphatase PP2A-B56 to BRCA2 through a conserved binding motif. We show that the phosphorylation-dependent formation of the BRCA2-PP2A-B56 complex is required for efficient RAD51 loading to sites of DNA damage and HR-mediated DNA repair. Moreover, we find that several cancer-associated mutations in BRCA2 deregulate the BRCA2-PP2A-B56 interaction and sensitize cells to PARP inhibition. Collectively, our work uncovers PP2A-B56 as a positive regulator of BRCA2 function in HR with clinical implications for BRCA2 and PP2A-B56 mutated cancers.
2021-07-26 | MSV000087884 | MassIVE
Project description:P-TEFb and CDK12 facilitate transcription elongation by RNA polymerase II and play prominent roles in cancer. Understanding their functional interplay could inform novel anti-cancer strategies. While inhibition of CDK12 downregulates unique sets of genes, eliciting genomic instability that is being exploited for novel therapies, little is known about the significance of transcriptional induction in CDK12-targeted cells. We show that inhibition of CDK12 in colon cancer-derived cells activates P-TEFb and induces genes of key cancer signaling pathways, including p53 and NF-kB. Mechanistically, cancer cells become exquisitely dependent on P-TEFb through activation of p53-dependent apoptosis and attenuation of NF-kB-dependent proliferation. Furthermore, we show that the DNA damage-responsive ATM kinase mediates these effects. While ATM is required for the synthetic lethality of CDK12 and P-TEFb co-targeting in p53-proficient cells, co-inhibition of ATM and CDK12 synergizes in decreasing viability of p53-deficient cells. Finally, pairwise targeting of CDK12, P-TEFb and transcription initiation kinase CDK7 stimulates p53-dependent apoptosis of cancer cell spheroids. We propose that the stimulation of Pol II pause release by P-TEFb at the signal-responsive genes underlies the dependence of CDK12-targeted cancer cells on P-TEFb. Together, our work provides a rationale for combinatorial targeting of CDK12 and P-TEFb or the induced oncogenic pathways in cancer.
2023-09-17 | GSE236803 | GEO
Project description:Maintenance of genomic stability depends on the DNA damage response (DDR), a biological barrier in early stages of cancer development. Failure of this response results in genomic instability and high predisposition toward lymphoma, as seen in patients with ataxia-telangiectasia mutated (ATM) dysfunction. ATM activates multiple cell cycle checkpoints and DNA repair following DNA damage, but its influence on posttranscriptional gene expression has not been examined on a global level. We show that ionizing radiation (IR) modulates the dynamic association of the RNA-binding protein HuR with target mRNAs in an ATM-dependent manner, potentially coordinating the genotoxic response as an RNA operon. Pharmacologic ATM inhibition and use of ATM-null cells revealed a critical role for ATM in this process. Numerous mRNAs encoding cancer-related proteins were differentially associated with HuR depending on the functional state of ATM, in turn affecting expression of encoded proteins. The findings presented here reveal a previously unidentified role of ATM in controlling gene expression post-transcriptionally. Dysregulation of this DDR RNA operon is likely relevant to lymphoma development in ataxia-telangiectasia individuals. These novel RNA regulatory modules and genetic networks provide critical insight into the function of ATM in oncogenesis. B-lymphocyte cell lines GM02184 (wild type, ATM +/+) and GM03332 (AT, ATM -/-) were either untreated or exposed to 1 Gy of IR. 6 h later cells were harvested and used for immunoprecipitation (IP) in the presence of HuR antibody (Santa Cruz Biotech.). RNA from IP material was extracted and used for microarray analysis.
2011-01-05 | E-GEOD-25848 | biostudies-arrayexpress
Project description:Biallelic loss of cyclin-dependent kinase 12 (CDK12) defines a unique molecular subtype of metastatic castration resistant prostate cancer (mCRPC). It remains unclear, however, whether CDK12 loss per se is sufficient to drive prostate cancer development—either alone, or in the context of other genetic alterations—and whether CDK12-mutant tumors exhibit sensitivity to specific pharmacotherapies. Here, we demonstrate that tissue-specific Cdk12 ablation is sufficient to induce preneoplastic lesions in the mouse prostate. Allograft-based CRISPR screening demonstrated that Cdk12 loss is positively associated with p53 inactivation, but negatively associated with Pten inactivation—similar to what is seen in human mCRPC. Consistent with this, ablation of Cdk12 in prostate organoids with concurrent p53 loss promotes their proliferation and ability to form tumors in mice, while Cdk12 knockout in the Pten-null prostate cancer mouse model abrogates tumor growth. Bigenic CDK12 and p53 loss allografts represent a new syngeneic model for the study of prostate cancer. Cdk12-null organoids (either with or without p53 co-ablation) and patient-derived xenografts from tumors with CDK12 inactivation are highly sensitive to inhibition or degradation of its paralog kinase, CDK13. Together, these data identify CDK12 as a bona fide tumor suppressor gene with impact on tumor progression and paralog-based synthetic lethality is a promising strategy for treating CDK12 mutant mCRPC.
2024-04-17 | GSE254388 | GEO
Project description:Biallelic loss of cyclin-dependent kinase 12 (CDK12) defines a unique molecular subtype of metastatic castration resistant prostate cancer (mCRPC). It remains unclear, however, whether CDK12 loss per se is sufficient to drive prostate cancer development—either alone, or in the context of other genetic alterations—and whether CDK12-mutant tumors exhibit sensitivity to specific pharmacotherapies. Here, we demonstrate that tissue-specific Cdk12 ablation is sufficient to induce preneoplastic lesions in the mouse prostate. Allograft-based CRISPR screening demonstrated that Cdk12 loss is positively associated with p53 inactivation, but negatively associated with Pten inactivation—similar to what is seen in human mCRPC. Consistent with this, ablation of Cdk12 in prostate organoids with concurrent p53 loss promotes their proliferation and ability to form tumors in mice, while Cdk12 knockout in the Pten-null prostate cancer mouse model abrogates tumor growth. Bigenic CDK12 and p53 loss allografts represent a new syngeneic model for the study of prostate cancer. Cdk12-null organoids (either with or without p53 co-ablation) and patient-derived xenografts from tumors with CDK12 inactivation are highly sensitive to inhibition or degradation of its paralog kinase, CDK13. Together, these data identify CDK12 as a bona fide tumor suppressor gene with impact on tumor progression and paralog-based synthetic lethality is a promising strategy for treating CDK12 mutant mCRPC.
2024-04-17 | GSE254387 | GEO
Project description:DNA Double Strand Breaks (DSBs) are harmful lesions that require rapid detection and repair in order to avoid toxic genomic rearrangements. DSBs elicit the so called DNA Damage Response (DDR), largely relying on ataxia telangiectasia mutated (ATM) and DNA Protein Kinase (DNAPK), two members of the PI3K-like kinase family, whose respective functions during the sequential steps of the DDR remains controversial. Using the DIvA cell line, expressing the AsiSI restriction enzyme, we have investigated the role of ATM and DNAPK in several aspects of the DDR upon induction of multiple clean DSBs throughout the human genome. High resolution mapping revealed that they are activated and spread in cis on a confined region surrounding all DSBs, independently of the pathway used for repair. However, a thorough analysis of repair kinetics, H2AX domain establishment and H2AX foci structure and dynamics revealed non overlapping functions for the two kinases once recruited at DSBs. Our results suggest that ATM is not solely acting on chromatin marks but also on chromatin organisation in order to ensure repair accuracy and survival.
2015-09-30 | E-MTAB-2992 | biostudies-arrayexpress
Project description:Maintenance of genomic stability depends on the DNA damage response (DDR), a biological barrier in early stages of cancer development. Failure of this response results in genomic instability and high predisposition toward lymphoma, as seen in patients with ataxia-telangiectasia mutated (ATM) dysfunction. ATM activates multiple cell cycle checkpoints and DNA repair following DNA damage, but its influence on posttranscriptional gene expression has not been examined on a global level. We show that ionizing radiation (IR) modulates the dynamic association of the RNA-binding protein HuR with target mRNAs in an ATM-dependent manner, potentially coordinating the genotoxic response as an RNA operon. Pharmacologic ATM inhibition and use of ATM-null cells revealed a critical role for ATM in this process. Numerous mRNAs encoding cancer-related proteins were differentially associated with HuR depending on the functional state of ATM, in turn affecting expression of encoded proteins. The findings presented here reveal a previously unidentified role of ATM in controlling gene expression post-transcriptionally. Dysregulation of this DDR RNA operon is likely relevant to lymphoma development in ataxia-telangiectasia individuals. These novel RNA regulatory modules and genetic networks provide critical insight into the function of ATM in oncogenesis.
2011-01-05 | GSE25848 | GEO
Project description:Genome instability is a potential limitation to the research and therapeutic application of induced pluripotent stem cells (iPSCs). Observed genomic variations reflect the combined activities of DNA damage, cellular DNA damage response (DDR), and selection pressure in culture. To understand the contribution of DDR on the distribution of copy number variations (CNVs) in iPSCs, we mapped CNVs of iPSCs with mutations in the central DDR gene ATM onto genome organization landscapes defined by genome-wide replication timing profiles. We show that following reprogramming the early and late replicating genome is differentially affected by CNVs in ATM deficient iPSCs relative to wild type iPSCs. Specifically, the early replicating regions had increased CNV losses during retroviral reprogramming. This differential CNV distribution was not present after later passage or after episomal reprogramming. Comparison of different reprogramming methods in the setting of defective DNA damage response reveals unique vulnerability of early replicating open chromatin to retroviral vectors. 4 cell lines, all in duplicates
2016-03-29 | E-GEOD-78717 | biostudies-arrayexpress
Project description:Genome instability is a potential limitation to the research and therapeutic application of induced pluripotent stem cells (iPSCs). Observed genomic variations reflect the combined activities of DNA damage, cellular DNA damage response (DDR), and selection pressure in culture. To understand the contribution of DDR on the distribution of copy number variations (CNVs) in iPSCs, we mapped CNVs of iPSCs with mutations in the central DDR gene ATM onto genome organization landscapes defined by genome-wide replication timing profiles. We show that following reprogramming the early and late replicating genome is differentially affected by CNVs in ATM deficient iPSCs relative to wild type iPSCs. Specifically, the early replicating regions had increased CNV losses during retroviral reprogramming. This differential CNV distribution was not present after later passage or after episomal reprogramming. Comparison of different reprogramming methods in the setting of defective DNA damage response reveals unique vulnerability of early replicating open chromatin to retroviral vectors. We isolated RNA from Ataxia-telangiectasia (A-T) patient fibroblast derived iPS cells and A-T patient fibroblasts for hybridization to the Affymetrix gene expression microarrays.
2016-03-29 | E-GEOD-78716 | biostudies-arrayexpress