R-loops and PRC1 repress Polycomb-target genes in mouse embryonic stem cells

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: 3070970

Axial progenitors generate trunk neural crest cells at a high efficiency in vitro

Expression profiling by array. Gene expression profiling utilised total RNA extracted from ES cells (N=3); hPSC derived Cranial neural crest precursors (N=3); hPSC derived Cranial neural crest cells (N=3); hPSC derived Cranial neural crest cells after RA treatment to posteriorise (N=3); hPSC derived Neuromesodermal progenitors (N=3); hPSC derived Trunk neural crest progenitors (N=3); hPSC derived trunk neural crest cells (N=3)The in vitro generation of neural crest (NC) cells from human pluripotent stem cells (hPSCs) is a valuable approach to study human NC biology and isolate NC derivatives for disease modelling/regenerative medicine applications. However, conventional differentiation protocols induce only a modest yield of NC cells corresponding to the trunk level. Here we show that trunk NC cells and, their downstream derivatives, sympathoadrenal progenitors, can be produced at a high efficiency from hPSC-derived axial progenitors, the in vitro counterparts of the posteriorly-located drivers of embryonic axis elongation. Moreover, using transcriptome analysis, we define the molecular signatures associated with the emergence of human NC cells of distinct axial identities. Collectively, our findings indicate that a post-cranial NC state is achieved through two different routes: the birth of cardiac and vagal NC is facilitated by retinoic acid-induced posteriorisation of an anterior precursor whereas a trunk fate relies on a posterior axial progenitor intermediate.Frith TJ, Granata I, Stout E, Wind M, Thompson O, Stavish D, Heath PR, Hackland JO, Anastassiadis K, Gouti M, Briscoe J, Wilson V, Guarracino MR, Andrews PW, Tsakridis A, 2018, Axial progenitors generate trunk neural crest cells at a high efficiency in vitro, Gene Expression Omnibus, https://www.ncbi.nlm.nih.gov/geo/query/acc.cgi?acc=GSE10926