Experiment type: Genome binding/occupancy profiling by high throughput sequencing.
Overall design: Genome-wide distribution of PRC2 subunits EZH2 and SUZ12 in mESC in presence or absence of R-loops.
R-loops are three-stranded nucleic acid structures that form naturally during transcription, especially over unmethylated CpG-rich promoters. In mESC, such promoters of developmental regulator genes are occupied by the Polycomb-repressor complexes PRC1 and PRC2. Here we have explored the possibility that R-loops form over Polycomb-repressed genes and play a role in their transcriptional silencing. Using single gene and genome-wide analyses, we show that R-loops form at a specific subset of PRC-target genes and contribute to Polycomb occupancy on chromatin. Removal of R-loops leads to an up-regulation of nascent and processed transcripts and the appearance of the elongating form of RNA polymerase II. In contrast, removal of PRC2 does not influence R-loop formation, transcriptional repression and PRC1 recruitment. We finally show that R-loops and PRC1 can repress Polycomb-target genes in the absence of PRC2. Our results uncover an unanticipated synergy between R-loops and PRC1 in Polycomb repression mechanisms.
Skourti-Stathaki K, Torlai Triglia E, Warburton M, Voigt P et al. R-Loops Enhance Polycomb Repression at a Subset of Developmental Regulator Genes. Mol Cell 2019 Mar 7;73(5):930-945.e4. PMID: 30709709