Ubiquitination of DNA Damage-Stalled RNAPII Promotes Transcription-Coupled Repair

Transcription-coupled nucleotide excision repair (TC-NER) is initiated by the stalling of elongating RNA polymerase II (RNAPIIo) at DNA lesions. The ubiquitination of RNAPIIo in response to DNA damage is an evolutionarily conserved event, but its function in mammals is unknown. Here, we identified a single DNA damage-induced ubiquitination site in RNAPII at RPB1-K1268, which regulates transcription recovery and DNA damage resistance. Mechanistically, RPB1-K1268 ubiquitination stimulates the association of the core-TFIIH complex with stalled RNAPIIo through a transfer mechanism that also involves UVSSA-K414 ubiquitination. We developed a strand-specific ChIP-seq method, which revealed RPB1-K1268 ubiquitination is important for repair and the resolution of transcriptional bottlenecks at DNA lesions. Finally, RPB1-K1268R knockin mice displayed a short life-span, premature aging, and neurodegeneration. Our results reveal RNAPII ubiquitination provides a two-tier protection mechanism by activating TC-NER and, in parallel, the processing of DNA damage-stalled RNAPIIo, which together prevent prolonged transcription arrest and protect against neurodegeneration. [Display omitted] •RNAPII RPB1-K1268 ubiquitination is essential for transcription recovery and DNA repair•Sequential ubiquitination of RNAPII and UVSSA coordinates the recruitment of TFIIH•Strand-specific ChIPseq enables mapping of RNAPII and reveals genome-wide repair kinetics•RNAPII ubiquitination protects against neurodegeneration phenotype in Cockayne syndrome Ubiquitination of RNA polymerase promotes recruitment of TFIIH and initiation of transcription-coupled repair, and dysregulation of this process in mice has neurological consequences akin to Cockayne syndrome.