水稻H3K36组蛋白甲基转移酶调节生殖发育的分子机制研究

Rice histone H3 lysine 36 (H3K36) methyltransferases (e.g. SDG708, SDG724, SDG725) are important epigenetic regulators involved in the regulation of plant growth and development. Yet, whether and how these multiple enzymes work independently and/or cooperatively remain obscure so far. Our preliminary analyses indicate that these different enzymes display varied degree of activity in H3K36 mono-, di- and tri-methylation and their combined knockdown synergistically impacts rice plant height, tiller number and grain yield. Within this project: 1) we will extend our genetic analyses by generating overexpression lines and combined mutants and analyzing their phenotypes into detail; 2) we will use diverse molecular biology methods to investigate functional relationships of the different H3K36-methyltransferases; 3) we will use RNA-seq to address transcriptome changes and correlations in the different single and combined knockdown mutants; 4) we will use ChIP-seq to unravel H3K36 methylation patterns and their correlation with transcriptome changes in the different mutants; 5) we will use yeast two hybrid and co-immunoprecipitation/Mass Spectrometry to identify and characterize new partners of the different H3K36-methyltransferases. Together, our work will provide important insight into molecular mechanisms about how multiple H3K36-methyltransferases synergistically regulate rice plant reproductive development, which has great interest in basic research as well as in rice high-yield grain breeding.

水稻组蛋白H3K36甲基转移酶与生长发育过程密切相关。我们通过分析分别缺失H3K36甲基转移酶SDG708、SDG724或SDG725的水稻单突变体和双突变体，发现各H3K36甲基转移酶在H3K36甲基化修饰中具有不同的酶活性并相互影响，表达下调影响水稻分蘖数、结实率和千粒重。为了研究H3K36甲基转移酶协同作用调控水稻生殖发育的分子机理，本项目将展开下述研究：1.进一步构建各H3K36甲基转移酶过表达和多突变材料，分析表型和统计重要农艺性状；2.蛋白质相互作用技术分析各H3K36甲基转移酶在水稻体内的相互关系；3.RNA测序分析各H3K36甲基转移酶缺失对转录组的影响；4.ChIP-seq分析各H3K36甲基转移酶的靶基因；5.利用酵母双杂交及质谱技术筛选和鉴定H3K36甲基转移酶的互作蛋白。这些研究对于揭示水稻组蛋白H3K36甲基转移酶调节水稻生殖发育的分子机理具有重要意义。

水稻组蛋白H3K36甲基转移酶与生长发育过程密切相关。本项目在在立项之初已发现水稻H3K36甲基转移酶SDG708、SDG724或SDG725在H3K36甲基化修饰中具有不同的酶活性并相互影响，并调节水稻分蘖数、结实率和千粒重等重要农艺性状。为了研究三个水稻H3K36甲基转移酶如何协同调控水稻生殖发育的分子机理，本项目不仅研究了各H3K36甲基转移酶过表达和多突变材料的表型特征，还对重要农艺性状进行了大田统计分析；同时，通过RNA和ChIP-seq 分析研究了受各H3K36甲基转移酶调控靶基因的特征。研究结果表明水稻SDG708，SDG724和SDG725甲基转移酶在H3K36me1/2/3修饰建立上具有底物偏好性，而且调控的靶基因也存在明显差异，有两个重要的成花素基因，Hd3a和RFT1，在708Ri-1和725Ri-1转基因材料中表达量都明显下降。这可能是影响水稻开花时间及产量性状的形成关键靶基因，Hd3a和RFT1表达量变化与水稻H3K36甲基转移酶相关突变体呈现的表型相一致。因此，这项研究揭示了SDG708、SDG724和SDG725改变H3K36me1/2/3的修饰状态，促进下游靶基因的表达，从而调控水稻水稻开花和生殖发育相关的多重表型的分子机制，也为探索开发“优质抗逆水稻新品种”提供重要的思路。