Polycomb group (PcG) proteins maintain transcriptional repression during development, likely by creating repressive chromatin states. The Extra Sex Combs (ESC) and Enhancer of Zeste [E(Z)] proteins are partners in an essential PcG complex, but its full composition and biochemical activities are not known. A SET domain in E(Z) suggests this complex might methylate histones. We purified an ESC-E(Z) complex from Drosophila embryos and found four major subunits: ESC, E(Z), NURF-55, and the PcG repressor, SU(Z)12. A recombinant complex reconstituted from these four subunits methylates lysine-27 of histone H3. Mutations in the E(Z) SET domain disrupt methyltransferase activity in vitro and HOX gene repression in vivo. These results identify E(Z) as a PcG protein with enzymatic activity and implicate histone methylation in PcG-mediated silencing.
Bibliographical noteFunding Information:
We thank Rick Jones, Gunter Reuter, Pierre Spierer, Gary Struhl, and Chiann-mun Chen for fly stocks and antibodies. We acknowledge the Molecular Biology Core Facilities at the Dana-Farber Cancer Institute and the HHMI Biopolymer-Keck Foundation Biotechnology Resource Laboratory at Yale University for protein sequencing. We thank LeeAnn Higgins of the University of Minnesota Mass Spectrometry Consortium for help with processing mass spectrometry samples. J.M. thanks Margarethe Bakala and Peter Ahnesorg for initial analysis and sequencing of novel E(z) alleles, Dirk Beuchle for help with generating transgenic flies, and Gary Struhl for discussions. J.M. also thanks Christiane Nüsslein-Volhard for support and encouragement. N.J.F. thanks members of the Kingston lab for assistance in growing Sf9 cells. J.A.S. thanks Tom Hays, Ann Rougvie, and Meg Titus for advice and discussions. This work was supported by NIH grants to J.A.S. and R.E.K. and funding from Hoechst AG to R.E.K. M.B.O. is an Associate Investigator for the Howard Hughes Medical Institute. N.J.F. is supported by the Cancer Research Fund of the Damon Runyon-Walter Winchell Foundation (DRG-1589).
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