Auto-inhibition of PRC2 by the broadly expressed long isoform of AEBP2.

Polycomb Repressive Complex 2 (PRC2) is an essential chromatin regulator responsible for mono-, di- and tri- methylating H3K27. Control of PRC2 activity is a critical process in development and disease. While PRC2 is inhibited in germinal cells, no inhibitory cofactor has been identified in somatic cells. Here we show that the alternative isoforms of its accessory subunit AEBP2, namely AEBP2 S (short) and AEBP2L (long), perform opposite functions in modulating PRC2 activity. While AEBP2S is predominantly expressed during early embryogenesis, AEBP2L is expressed throughout embryogenesis and adulthood. AEBP2L inhibits both DNA binding by PRC2 and its histone methyltransferase activity in vitro and impairs PRC2 binding to target genes in embryonic stem cells. In contrast, AEBP2S promotes the DNA-binding activity of PRC2 and is essential for de novo repression of target genes during the transition from naïve to primed pluripotency. Mechanistically, through high-resolution Cryo-EM and mutagenesis, we show that the recently evolved, negatively charged N-terminal region of AEBP2L inhibits PRC2. We propose a model in which the N-terminus of AEBP2L arose in vertebrates to restrain PRC2 activity in somatic cells. To investigate the ability of AEBP2 isoforms to bind chromatin in cells, we ectopically expressed human AEBP2 isoforms in generated by us Aebp2 KO mouse embryonic stem cells (E14), and performed ChIP-Rx of AEBP2 and SUZ12. The cell lines included in this dataset are: WT E14, Aebp2 KOs, Aebp2 KO+AEBP2L (AL) and Aebp2 KO+AEBP2S (AS). To determine the transcriptional activity of the AEBP2 promoters in mouse embryonic stem cells, we performed 3 replicates of RNA-Seq in WT E14 mESCs. To evaluate the genome-wide localisation of AEBP2, SUZ12, JARID2 and H3K27me3, we performed ChIP-Seq/Rx (proteins), or Cut&Run (H3K27me3) in our Aebp2 knock-out ESC line panel: WT E14, AEBP2L KO (LKO), AEBP2S KO (SKO) and complete Aebp2 KO (AEBP2KO). To determine the contributions of the AEBP2 isoforms to de novo gene repression, we performed 3 replicates of QuantSeq in ESCs and Epiblast-like cells (D0 and D2 samples, respectively) in the four KO cell lines (WT, LKO, SKO, Aebp2 KO).