Variant Pcgf-PRC1 Regulates Susceptibility of PRC2 Mediated H3K27me3 to Safeguard B Cell Fate of Hematopoietic Stem/Progenitor Cells

Polycomb repressive complex (PRC) resides in two major complexes PRC1 and PRC2. They cooperate with each other to coordinate proper developmental process by silencing target genes; PRC1 posits H2AK119ub1 and PRC2 catalyzes trimethylation of H3K27 (H3K27me3). The PRC1 component BMI1/PCGF4 has long been recognized to be essential in the maintenance of normal and malignant hematopoietic stem cells (HSCs). Recently, diversity of PRC1 has been noticed and PRC1 is now classified into six alternative complexes depending on PCGF proteins. In embolic stem cells, PRC1 which contains PCGF1 (PCGF1-PRC1) has been demonstrated to serve upstream of the BMI1/PCGF4-PRC1. However, the impact of BCOR, which is a component of PCGF1-PRC1 on hematopoiesis is clearly different from BMI1/PCGF4; the mice deficient for BCOR exhibited normal HSC activities and BCOR rather prevented leukemic transformation of HSCs, suggesting the previously unappreciated gene control mechanisms of PCGF1-PRC1. To tackle this issue, we focused on the roles of PCGF1 in hematopoiesis.

Loss of Pcgf1 in hematopoietic stem cells led to severe reduction of B lineage cells with an expansion of myeloid progenitors due to defects in lymphoid-primed multipotent progenitor (LMPP) cells. To explore the molecular mechanisms, we have established Id3-overexpressing hematopoietic progenitor cells (IdHPs) which correspond to LMPP-like cells (Ikawa et al. 2015) from bone marrow of ERT2-Cre Pcgf1 flox mice. The ChIP-seq analysis of normal IdHPs identified 1274 genes whose promoters were associated with PCGF1 peaks and 37% of them exhibited enrichment of H3K27me3 and binding of SUZ12 (PRC2). Deletion of Pcgf1 destabilized H3K27me3 levels, resulting in re-activation of genes associated with PCGF1 and SUZ12 peaks, whereas the chromatin occupancy of SUZ12 was not affected. Intriguingly, proteomic analysis demonstrated that PCGF1 interacts with key factors responsible for the organization of nucleosomes and PCGF1 loss triggered a decline of nucleosome-densities in promoters of genes occupied by PCGF1 and SUZ12 peaks. Since enzymatic activity of PRC2 is dependent on nucleosome-densities, PCGF1 is likely to regulate the susceptibility of H3K27me3 by PRC2 through determination of the nucleosome-densities. Furthermore, genes which were downregulated by PCGF1-nucleosome-H3K27me3 axis entailed many myeloid-related genes and knock down of one of those myeloid genes partially restored the B cell differentiation potential of Pcgf1-KO hematopoietic stem/progenitor cells (HSPCs), supporting the biological significance of the PCGF1-nucleosome-H3K27me3 axis. Collectively, these results indicate PCGF1 determines cellular fate of HSPCs through stabilization of nucleosomal organization.