Genomic context- and H2AK119 ubiquitination-dependent inheritance of human Polycomb silencing.

Polycomb repressive complexes 1 and 2 (PRC1 and 2) are required for heritable repression of developmental genes. The cis- and trans-acting factors that contribute to epigenetic inheritance of mammalian Polycomb repression are not fully understood. Here, we show that, in human cells, ectopically induced Polycomb silencing at initially active developmental genes, but not near ubiquitously expressed housekeeping genes, is inherited for many cell divisions. Unexpectedly, silencing is heritable in cells with mutations in the H3K27me3 binding pocket of the Embryonic Ectoderm Development (EED) subunit of PRC2, which are known to disrupt H3K27me3 recognition and lead to loss of H3K27me3. This mode of inheritance is less stable and requires intact PRC2 and recognition of H2AK119ub1 by PRC1. Our findings suggest that maintenance of Polycomb silencing is sensitive to local genomic context and can be mediated by PRC1-dependent H2AK119ub1 and PRC2 independently of H3K27me3 recognition.

SENP3 and USP7 regulate Polycomb-rixosome interactions and silencing functions.

The rixosome and PRC1 silencing complexes are associated with deSUMOylating and deubiquitinating enzymes, SENP3 and USP7, respectively. How deSUMOylation and deubiquitylation contribute to rixosome- and Polycomb-mediated silencing is not fully understood. Here, we show that the enzymatic activities of SENP3 and USP7 are required for silencing of Polycomb target genes. SENP3 deSUMOylates several rixosome subunits, and this activity is required for association of the rixosome with PRC1. USP7 associates with canonical PRC1 (cPRC1) and deubiquitinates the chromodomain subunits CBX2 and CBX4, and inhibition of USP activity results in disassembly of cPRC1. Finally, both SENP3 and USP7 are required for Polycomb- and rixosome-dependent silencing at an ectopic reporter locus. These findings demonstrate that SUMOylation and ubiquitination regulate the assembly and activities of the rixosome and Polycomb complexes and raise the possibility that these modifications provide regulatory mechanisms that may be utilized during development or in response to environmental challenges.

Rixosomal RNA degradation contributes to silencing of Polycomb target genes.

Polycomb repressive complexes 1 and 2 (PRC1 and PRC2) are histone-modifying and -binding complexes that mediate the formation of facultative heterochromatin and are required for silencing of developmental genes and maintenance of cell fate1-3. Multiple pathways of RNA decay work together to establish and maintain heterochromatin in fission yeast, including a recently identified role for a conserved RNA-degradation complex known as the rixosome or RIX1 complex4-6. Whether RNA degradation also has a role in the stability of mammalian heterochromatin remains unknown. Here we show that the rixosome contributes to silencing of many Polycomb targets in human cells. The rixosome associates with human PRC complexes and is enriched at promoters of Polycomb target genes. Depletion of either the rixosome or Polycomb results in accumulation of paused and elongating RNA polymerase at Polycomb target genes. We identify point mutations in the RING1B subunit of PRC1 that disrupt the interaction between PRC1 and the rixosome and result in diminished silencing, suggesting that direct recruitment of the rixosome to chromatin is required for silencing. Finally, we show that the RNA endonuclease and kinase activities of the rixosome and the downstream XRN2 exoribonuclease, which degrades RNAs with 5' monophosphate groups generated by the rixosome, are required for silencing. Our findings suggest that rixosomal degradation of nascent RNA is conserved from fission yeast to human, with a primary role in RNA degradation at facultative heterochromatin in human cells.

DNA Demethylation Dynamics in the Human Prenatal Germline.

Global DNA demethylation in humans is a fundamental process that occurs in pre-implantation embryos and reversion to naive ground state pluripotent stem cells (PSCs). However, the extent of DNA methylation reprogramming in human germline cells is unknown. Here, we performed whole-genome bisulfite sequencing (WGBS) and RNA-sequencing (RNA-seq) of human prenatal germline cells from 53 to 137 days of development. We discovered that the transcriptome and methylome of human germline is distinct from both human PSCs and the inner cell mass (ICM) of human blastocysts. Using this resource to monitor the outcome of global DNA demethylation with reversion of primed PSCs to the naive ground state, we uncovered hotspots of ultralow methylation at transposons that are protected from demethylation in the germline and ICM. Taken together, the human germline serves as a valuable in vivo tool for monitoring the epigenome of cells that have emerged from a global DNA demethylation event.