Structural And Functional Research Of Methylated Histone Binding Modules

Histone post-translational modifications (PTMs) is a principle component of epigenetic regulations. N-terminal tails of histones are the most accessible regions that are subjected to various PTMs such as methylation, acetylation, phosphorylation and ubiquitination and so on. PTMs are believed to function in a combinatorial pattern referred to as the "histone code". The major function of PTMs is to create sites for the recruitment of proteins with specific binding modules which subsequently stimulate downstream biological processes. In the past decades, a wealth of conserved protein domains have been identified through biochemical and biophysical assays, which specifically bind histone PTMs, including bromodomain, PHD domain, WD40repeat and domains of Royal-superfamily and so on. On account of the variety of PTMs, PTM-binding modules are involed in nearly all biological processes. Currently, numerous of PTM-binding modules have been reported to be implicated in different human disease. More complex than other PTMs, histone methylation is recognized by much more different binding modules due to the diversity of methyl lysine signals. This dissertation focuses on the members of Royal-superfamily that specifically binding methylated lysine residues.In Chapter2, we solved the crystal structures of the tandem Tudor domain of Saccharomyces cerevisiae Sgf29and their complexes with H3K4me2and H3K4me3peptides, respectively, and show that Sgf29selectively binds H3K4me2/3marks. The tandem Tudor domains in Sgf29tightly pack against each other face-to-face. The H3A1and K4me3binding pockets created by each of the tandem Tudor domains and the fixed distance between these two pockets are the structural determinants in conferring Sgf29the ability of selectively recognizing H3K4me2/3, not other histone lysine sites. Our in vitro and in vivo functional assays show that Sgf29recognizes methylated H3K4to recruit the SAGA complex to target genes, underscoring the importance of Sgf29in gene regulation.Chapter3presents our work on chromodomain of heterchromatin protein HP1γ (Cbx3). HP1proteins have been identified to be involved multiple biological processes including gene transcription and DNA repair. We determined the crystal structures of the human Cbx3chromodomain in complex with dimethylated histone H1K26and trimethylated G9aK185peptides, respectively. Our in vitro assay indicates Cbx3chromodomain can bind to both H1K26me2and G9aK185me3with comparable binding affinities compared to H3K9me3. The complex structures unveil that the Cbx3chromodomain specifically binds methylated histone H1K26and G9aK185through a conserved mechanism.In Chapter4, we reported the crystal structures of human MPP8(hMPP8) chromodomain both in apo-form and in complex with the trimethylated histone H3lysine9(H3K9me3) peptide (residue1-15). Consistent with the high sequence homology of MPP8with Polycomb and HP1chromodomains, the complex structure of hMPP8-H3K9me3uncovers the detailed molecular mechanism of recruitment of MPP8chromodomain by HK9me3as well as its unexpected homodimerization. Based on the homodimerization, we build two models showing that the simultaneous binding of two histone tails to hMpp8homodimer either from the same nucleosome or from two separated nucleosomes.