Dynamic Protein Interactions of the Polycomb Repressive Complex 2 during Differentiation of Pluripotent Cells

Abstract

Polycomb proteins assemble to form complexes with important roles in epigenetic regulation. The Polycomb Repressive Complex 2 (PRC2) modulates the di- and tri-methylation of lysine 27 on histone H3, each of which are associated with gene repression. Although three subunits, EZH1/2, SUZ12, and EED, form the catalytic core of PRC2, a wider group of proteins associate with low stoichiometry. This raises the question of whether dynamic variation of the PRC2 interactome results in alternative forms of the complex during differentiation. Here we compared the physical interactions of PRC2 in undifferentiated and differentiated states of NTERA2 pluripotent embryonic carcinoma cells. Label-free quantitative proteomics was used to assess endogenous immunoprecipitation of the EZH2 and SUZ12 subunits of PRC2. A high stringency data set reflecting the endogenous state of PRC2 was produced that included all previously reported core and associated PRC2 components, and several novel interacting proteins. Comparison of the interactomes obtained in undifferentiated and differentiated cells revealed candidate proteins that were enriched in complexes isolated from one of the two states. For example, SALL4 and ZNF281 associate with PRC2 in pluripotent cells, whereas PCL1 and SMAD3 preferentially associate with PRC2 in differentiating cells. Analysis of the mRNA and protein levels of these factors revealed that their association with PRC2 correlated with their cell state-specific expression. Taken together, we propose that dynamic changes to the PRC2 interactome during differentiation may contribute to directing its activity during cell fate transitions.

The N‐terminal methyltransferase homologs NRMT1 and NRMT2 exhibit novel regulation of activity through heterotrimer formation

“…In contrast to PRC2‐EZH2, PRC2‐EZH1 represses transcription by binding and compacting nucleosomes . The PRC2‐EZH2 and PRC2‐EZH1 complexes are thought to be primarily mutually exclusive, but data indicate EZH1 and EZH2 can interact with each other both in vitro and in vivo . It is still functionally unclear why a PRC2 complex that contains EZH2 would also contain EZH1, but as they accomplish the same goal by different mechanisms, there may be certain circumstances when synergistic activity would be beneficial.…”

Enhancer function regulated by combinations of transcription factors and cofactors

“…SALL4, ZNF281, and SMAD3 were identified as interacting partners of PRC2 (Oliviero et al, ). SALL4 recruits PRC2 to the SOX2 and SOX17 loci in embryonic lineages (Abboud et al, ).…”

Unravelling the biology of chromatin in health and cancer using proteomic approaches

“…Label free quantitative proteomics has been routinely used to study chromatin remodeling complexes and protein interaction networks. This includes the analysis of the Tip60, Ino80, and SRCAP complexes [45], histone deacetylase complexes [46], the polycomb complex [47, 48, 49], and the cardiac TBX5 interactome [50], for example. In addition, the stoichiometry of chromatin remodeling proteins has been analyzed using label free quantitative proteomics approaches [51].…”