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CDK12 regulates DNA repair genes by suppressing intronic polyadenylation.

ABSTRACT: Mutations that attenuate homologous recombination (HR)-mediated repair promote tumorigenesis and sensitize cells to chemotherapeutics that cause replication fork collapse, a phenotype known as 'BRCAness'1. BRCAness tumours arise from loss-of-function mutations in 22 genes1. Of these genes, all but one (CDK12) function directly in the HR repair pathway1. CDK12 phosphorylates serine 2 of the RNA polymerase II C-terminal domain heptapeptide repeat2-7, a modification that regulates transcription elongation, splicing, and cleavage and polyadenylation8,9. Genome-wide expression studies suggest that depletion of CDK12 abrogates the expression of several HR genes relatively specifically, thereby blunting HR repair3-7,10,11. This observation suggests that the mutational status of CDK12 may predict sensitivity to targeted treatments against BRCAness, such as PARP1 inhibitors, and that CDK12 inhibitors may induce sensitization of HR-competent tumours to these treatments6,7,10,11. Despite growing clinical interest, the mechanism by which CDK12 regulates HR genes remains unknown. Here we show that CDK12 globally suppresses intronic polyadenylation events in mouse embryonic stem cells, enabling the production of full-length gene products. Many HR genes harbour more intronic polyadenylation sites than other expressed genes, and these sites are particularly sensitive to loss of CDK12. The cumulative effect of these sites accounts for the enhanced sensitivity of HR gene expression to CDK12 loss, and we find that this mechanism is conserved in human tumours that contain loss-of-function CDK12 mutations. This work clarifies the function of CDK12 and underscores its potential both as a chemotherapeutic target and as a tumour biomarker.

SUBMITTER: Dubbury SJ

PROVIDER: S-EPMC6328294 | biostudies-literature | 2018 Dec

REPOSITORIES: biostudies-literature

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CDK12 regulates DNA repair genes by suppressing intronic polyadenylation.

Dubbury Sara J SJ Boutz Paul L PL Sharp Phillip A PA

Nature 20181128 7734

Mutations that attenuate homologous recombination (HR)-mediated repair promote tumorigenesis and sensitize cells to chemotherapeutics that cause replication fork collapse, a phenotype known as 'BRCAness'1. BRCAness tumours arise from loss-of-function mutations in 22 genes1. Of these genes, all but one (CDK12) function directly in the HR repair pathway1. CDK12 phosphorylates serine 2 of the RNA polymerase II C-terminal domain heptapeptide repeat2-7, a m ...[more]

PMID: 30487607

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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.

Dataset Information

CDK12 regulates DNA repair genes by suppressing intronic polyadenylation.

Publications

CDK12 regulates DNA repair genes by suppressing intronic polyadenylation.

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