Understanding the role of accessory components in regulation of the Polycomb Repressive Complex 2 in mESCs and cancer

Abstract

Polycomb Repressive Complex 2 is a chromatin regulatory complex composed of core subunits EZH1/2, EED, SUZ12 and RBBP4/7, around which several accessory components assemble in a mutually exclusive manner to form structurally distinct PRC2.1 and PRC2.2. The primary function of the complex is mono-, di- and trimethylation of histone H3 at lysine K27, marks associated with active gene bodies, intergenic regions, and repressed gene promoters, respectively. While recent proteomic studies have greatly expanded our knowledge on the function of the complex, it remains to be elucidated how exactly the accessory components contribute to Polycomb-mediated transcriptional regulation. For example, it remains unclear whether the accessory components contribute to the enzymatic activity of the complex, or its targeting to chromatin. Moreover, their potential role in cancer development is mostly unknown. This PhD thesis addresses these questions using mouse embryonic stem cells (mESCs) and cancer cells as models. Firstly, through generation and analysis of PCL1-3/JARID2 quadruple knockout mESCs cell line I establish that PRC2.1 and PRC2.2 act synergistically to localise core PRC2 enzymatic activity towards its target sites. Secondly, using a PRC2-focused CRISPR tiling screen, I find novel regions of potential functional dependency within AEBP2 and PALI1 proteins. I also identify EED as a potential therapeutic target in malignant rhabdoid tumours (MRTs). Finally, I elucidate the functional divergences of two isoforms of AEBP2 (AEBP2-S and AEBP2-L) in regulating Polycomb function in mESCs. In summary, I believe that these results contribute to our understanding of Polycomb function and the specific roles of accessory components in mESCs and cancer.