Structural Analysis of the Ash2L/Dpy-30 Complex Reveals a Heterogeneity in H3K4 Methylation. Structural Analysis of the Ash2L/Dpy-30 Complex Reveals a Heterogeneity in H3K4 Methylation

Dpy-30 is a regulatory subunit controlling the histone methyltransferase activity of the KMT2 enzymes in vivo. Paradoxically, in vitro methyltransferase assays revealed that Dpy-30 only modestly participates in the positive heterotypic allosteric regulation of these methyltransferases. Detailed genome-wide, molecular and structural studies reveal that an extensive network of interactions taking place at the interface between Dpy-30 and Ash2L are critical for the correct placement, genome-wide, of H3K4me2 and H3K4me3 but marginally contribute to the methyltransferase activity of KMT2 enzymes in vitro. Moreover, we show that H3K4me2 peaks persisting following the loss of Dpy-30 are found in regions of highly transcribed genes, highlighting an interplay between Complex of Proteins Associated with SET1 (COMPASS) kinetics and the cycling of RNA polymerase to control H3K4 methylation. Overall, our data suggest that Dpy-30 couples its modest positive heterotypic allosteric regulation of KMT2 methyltransferase activity with its ability to help the positioning of SET1/COMPASS to control epigenetic signaling. Overall design: 17 experiments: (4 genotypes x 2 ChIP antibodies x 2 replicates), 1 input

Dpy-30为体内调控KMT2家族酶组蛋白甲基转移酶活性的调控亚基。矛盾的是，体外甲基转移酶活性检测实验显示，Dpy-30仅对这类甲基转移酶发挥微弱的正向异型变构调控作用。
全基因组、分子及结构层面的细致研究表明，Dpy-30与Ash2L蛋白的互作界面所形成的广泛相互作用网络，对于全基因组范围内组蛋白H3第4位赖氨酸二甲基化（H3K4me2）与组蛋白H3第4位赖氨酸三甲基化（H3K4me3）的正确定位至关重要，但对KMT2家族酶的体外甲基转移酶活性仅具有微弱促进作用。
此外，本研究发现，Dpy-30缺失后仍持续存在的H3K4me2峰位于高转录基因区域，这凸显了SET1相关复合体（Complex of Proteins Associated with SET1, COMPASS）动力学与RNA聚合酶循环之间的相互作用，该相互作用可调控H3K4甲基化。
综上，本研究数据表明，Dpy-30将其对KMT2甲基转移酶活性的微弱正向异型变构调控，与其协助SET1/COMPASS复合体定位以调控表观遗传信号的功能相偶联。
整体实验设计：共17组实验，包含（4种基因型 × 2种染色质免疫沉淀（Chromatin Immunoprecipitation, ChIP）抗体 × 2次生物学重复），外加1组输入对照。