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Rouquier, S.

Publications and source records attributed to Rouquier, S..

2 recordsLinked to original sources

PAF1 facilitates RNA polymerase II ubiquitination by the Elongin A complex through phosphorylation by CDK12

The conserved Polymerase-Associated Factor 1 complex (PAF1C) regulates all stages of the RNA polymerase II (RNAPII) transcription cycle from the promoter to the 3 end formation site of mRNA encoding genes and has been linked to numerous transcription related processes. Here, we show that PAF1 interacts with Elongin A, a transcription elongation factor as well as a component of a cullin-RING ligase that targets stalled RNAPII for ubiquitination and proteasome-dependent degradation in response to DNA damage or other stresses. We show that, in absence of any induced stress, PAF1 physically interacts with the E3 ubiquitin ligase form of the Elongin A complex and facilitates ubiquitination of RNAPII. We demonstrate that this ubiquitination is dependent of the Ser2 phosphorylation of the RNAPII carboxy-terminal domain (CTD) by CDK12. Our findings highlight a novel unexpected role of PAF1-CDK12 in RNAPII transcription cycle, raising the possibility that the Elongin A ubiquitin ligase plays a role in normal transcription process, and suggest a transcription surveillance mechanism ready to degrade RNAPII if needed.

cell biology

Chromatin-associated MRN complex protects highly transcribing genes from genomic instability

The MRN-MDC1 complex plays a central role in the DNA damage response (DDR) and repair. Using Proteomics of Isolated Chromatin Fragments (PICh), we identified DDR factors, such as MDC1, among those that become highly associated with a genomic locus upon transcriptional activation. Purification of endogenous MDC1, in the absence of exogenous DNA damage, revealed its interaction with factors involved in gene expression and co-transcriptional RNA processing, in addition to DDR factors. ChIP-seq analysis showed that MDC1 interacting factors, MRE11 and NBS1 subunits of MRN, were co-localized throughout the genome and notably at TSSs and gene bodies of actively transcribing genes. Blockade of transcriptional elongation showed that binding of MRN was dependent on the RNAPII transcriptional complex rather than transcription per se. Depletion of MRN increased RNAPII abundance at TSSs and gene bodies of MRE11/NBS1-bound genes. Prolonged exposure of cells to either MRE11- or NBS1-depletion led to single nucleotide polymorphism formation across actively transcribing, MRE11 or NBS1 target genes. These data support a model by which association of the MRN complex with the transcriptional machinery constitutively scans active genes for transcription-induced DNA damage to preserve the integrity of the coding genome.

molecular biology