A Drosophila ESC-E(Z) protein complex is distinct from other polycomb group complexes and contains covalently modified ESC

The extra sex combs (ESC) and Enhancer of zeste [E(Z)] proteins, members of the Polycomb group (PcG) of transcriptional repressors, interact directly and are coassociated in fly embryos. We report that these two proteins are components of a 600-kDa complex in embryos. Using gel filtration and affini ty chromatography, we show that this complex is biochemically distinct from previously described complexes containing the PcG proteins Polyhomeotic, P olycomb, and Sex comb on midleg. In addition, we present evidence that ESC is phosphorylated in vivo and that this modified ESC is preferentially asso ciated in the complex with E(Z). Modified ESC accumulates between 2 and 6 h of embryogenesis, which is the developmental time when esc function is fir st required. We And that mutations in E(z) reduce the ratio of modified to unmodified ESC in vivo. We have also generated germ line transformants that express ESC proteins bearing site-directed mutations that disrupt ESC-E(Z) binding in vitro. These mutant ESC proteins fail to provide esc function, show reduced levels of modification in vivo, and are still assembled into c omplexes. Taken together, these results suggest that ESC phosphorylation no rmally occurs after assembly into ESC-E(Z) complexes and that it contribute s to the function or regulation of these complexes. We discuss how biochemi cally separable ESC-E(Z) and PC-PH complexes might work together to provide PcG repression.