RESUMO
The Drosophila Ash1 protein is a trithorax-group (trxG) regulator that antagonizes Polycomb repression at HOX genes. Ash1 di-methylates lysine 36 in histone H3 (H3K36me2) but how this activity is controlled and at which genes it functions is not well understood. We show that Ash1 protein purified from Drosophila exists in a complex with MRG15 and Caf1 that we named AMC. In Drosophila and human AMC, MRG15 binds a conserved FxLP motif near the Ash1 SET domain and stimulates H3K36 di-methylation on nucleosomes. Drosophila MRG15-null and ash1 catalytic mutants show remarkably specific trxG phenotypes: stochastic loss of HOX gene expression and homeotic transformations in adults. In mutants lacking AMC, H3K36me2 bulk levels appear undiminished but H3K36me2 is reduced in the chromatin of HOX and other AMC-regulated genes. AMC therefore appears to act on top of the H3K36me2/me3 landscape generated by the major H3K36 methyltransferases NSD and Set2. Our analyses suggest that H3K36 di-methylation at HOX genes is the crucial physiological function of AMC and the mechanism by which the complex antagonizes Polycomb repression at these genes.
Assuntos
Proteínas Cromossômicas não Histona/metabolismo , Metilação de DNA/genética , Proteínas de Ligação a DNA/metabolismo , Proteínas de Drosophila/metabolismo , Histona-Lisina N-Metiltransferase/metabolismo , Histonas/metabolismo , Fatores de Transcrição/metabolismo , Animais , Western Blotting , Proteínas de Ligação a DNA/genética , Drosophila/metabolismo , Proteínas de Drosophila/genética , Perfilação da Expressão Gênica , Genes Homeobox/genética , Humanos , Lisina/metabolismo , Espectrometria de Massas , Proteína 4 de Ligação ao Retinoblastoma/metabolismo , Fatores de Transcrição/genéticaRESUMO
Human ASH1L is the catalytic subunit of the conserved histone methyltransferase (HMTase) complex AMC that dimethylates lysine 36 in histone H3 (H3K36me2) to promote gene transcription in mammals and flies. Unlike AMC, ASH1L alone shows poor catalytic activity, because access to its substrate binding pocket is blocked by an autoinhibitory loop (AI loop) from the postSET domain. We report the crystal structure of the minimal catalytic active AMC complex containing ASH1L and its partner subunit MRG15. The structure reveals how binding of the MRG domain of MRG15 to a conserved FxLP motif in ASH1L results in the displacement of the AI loop to permit substrates to access the catalytic pocket of the ASH1L SET domain. Together, ASH1L activation by MRG15 therefore represents a delicate regulatory mechanism for how a cofactor activates an SET domain HMTase by releasing autoinhibition.