Polycomb group RING finger proteins 3/5 activate transcription via an interaction with the pluripotency factor Tex10...

Polycomb group (PcG) proteins are epigenetic transcriptional repressors that orchestrate numerous developmental processes and have been implicated in the maintenance of embryonic stem (ES) cell state. More recent evidence suggests that a subset of PcG proteins engages in transcriptional activation in some cellular contexts, but how this property is exerted remains largely unknown. Here, we generated ES cells with single or combined disruption of polycomb group RING finger protein 3 (Pcgf3) and Pcgf5 with the CRISPR-Cas9 technique. We report that although these mutant cells maintained their self-renewal and colony-forming capacity, they displayed severe defects in mesoderm differentiation in vitro and in vivo Using RNA-seq to analyze transcriptional profiles of ES cells with single or combined Pcgf3/5 deficiencies, we found that in contrast to the canonical role of the related polycomb repressive complex 1 (PRC1) in gene repression, Pcgf3/5 mainly function as transcriptional activators driving expression of many genes involved in mesoderm differentiation. Proteomic approaches and promoter occupancy analyses helped to establish an extended Pcgf3/5 interactome and identified several novel Pcgf3/5 interactors. These included testis-expressed 10 (Tex10), which may directly contribute to transcriptional activation via the transcriptional co-activator p300. Furthermore, Pcgf3/5 deletion in ES cells substantially reduced the occupancy of Tex10 and p300 at target genes. Finally, we demonstrated that Pcgf3/5 are essential for regulating global levels of the histone modifier H2AK119ub1 in ES cells. Our findings establish Pcgf3/5 as transcriptional activators that interact with Tex10 and p300 in ES cells and point to redundant activity of Pcgf3/5 in pluripotency maintenance.