Ubiquitylation of MYC couples transcription elongation with double-strand break repair at active promoters [CHIP-seq_MYC-ER]

MYC family proteins are oncogenic transcription factors that can globally affect the function of RNA Polymerase II (RNAPII). The ability of MYC proteins to promote transcription elongation depends on their ubiquitination, but the underlying mechanism and its biological relevance are unknown. Here we show that MYC and the Polymerase II associated factor, PAF1c, interact directly and their function is mutually dependent, since the specific binding of MYC to active promoters depends on PAF1c and, conversely, PAF1c is required for MYC-dependent pause release. Upon binding, PAF1c is rapidly transferred from MYC onto RNAPII and this transfer is driven by the HUWE1 ubiquitin ligase. Both MYC and HUWE1 globally control histone H2B ubiquitylation, which promotes transcriptional elongation and alters chromatin structure for double-strand break repair. Consistently, MYC suppresses the accumulation of double-strand breaks at promoters in response to topoisomerase II inhibition. While depletion of PAF1c has only minor effects on MYC-dependent gene expression, MYC induces rampant transcription-dependent DNA damage in PAF1c-depleted cells. We propose that the HUWE1-dependent transfer of PAF1c from MYC onto RNAPII is critical for absorbing the topological stress accompanied with deregulated and oncogenic transcription. U2OS cells which stably express a MYC-ER system. Cells were either treated with ETOH OR 4-OHT (200 nM) for 10 min or 30 min Input samples for ChIPseq_MYCER rep1: For each of the conditions '0min'€ (= ethanol treatment only), '10min', '30min', two separate 'input' chromatin samples were prepared, designated as 'rep1A'€and 'rep1B'. All six resulting fastq files were subsequently merged. This merged input file was mapped to hg19 and randomly subsampled to different read numbers, resulting in the bedgraph files ChIPseq_MYCER_INPUT_CTR9_rep1.bedgraph, ChIPseq_MYCER_INPUT_MYC_rep1.bedgrap, ChIPseq_MYCER_INPUT_pS2_rep1.bedgraph, to be used as controls for the CTR9, MYC, and pS2 ChIPseq samples, respectively.