ObSH3基因参与的水稻落粒性遗传调控网络解析

The key step in the evolution during domestication of rice is the elimination of seed dispersal. Compared with its ancestor wild rice, cultivated rice has eliminated the seed shattering trait with partially developed abscission layer. In our previous study, the seed shattering gene ObSH3 was cloned from African wild rice accession W1411 with shattering phenotype, which controls the asymmetric development of abscission layer. In this project we will further identify the protein interaction relationship between ObSH3 and the other dispersal genes using yeast two hybridization system, co-immunoprecipitation (Co-IP), bimolecular fluorescence complementation (BiFC) and other technologies. In combination with transcriptome analysis, reverse transcriptional fluorescence quantitative PCR, chromatin immunoprecipitation-Seq (ChIP-Seq) and electrophoretic mobility shift assays (EMSA) to analysis the downstream genes regulated by ObSH3, further mapping the seed shattering genetic regulation network which ObSH3 gene mainly involved. Besides, the different variation combinations of single mutant, double mutant and triple mutant of ObSH3, SH4 and qSH1 were constructed under the same background of W1411 by gene editing to study their effects on seed shattering. This study will give new clues to reveal the genetic regulation mechanism of seed shattering, and provide the theory foundation for breeding new varieties of rice which possess the modest seed shattering and suitable for mechanized harvesting in agricultural production.

落粒性的丧失是水稻驯化过程中的关键一步。与野生稻相比，栽培稻的离层发育不完整，导致其落粒性降低。申请人前期从非洲野生稻W1411中成功克隆了一个落粒性基因ObSH3，发现该基因控制离层的不对称发育。本项目将在此基础上进一步利用酵母双杂交、免疫共沉淀（Co-IP）以及双分子荧光互补（BiFC）等技术重点研究ObSH3基因与已克隆的落粒基因之间的蛋白互作关系，同时结合转录组分析、反转录荧光定量PCR、结合位点分析（ChIP-Seq）以及凝胶迁移（EMSA）分离ObSH3蛋白所调控的下游基因，解析ObSH3基因参与的水稻落粒性遗传调控网络。同时在同一背景下通过基因编辑构建ObSH3、SH4、qSH1等落粒基因的单突、双突和三突等变异组合，并比较它们对落粒性的影响。该研究不仅为揭示水稻落粒性遗传调控机制提供新线索，同时也为生产上培育落粒性适中、适于机械化收割的水稻新品种提供理论基础。

落粒性是水稻驯化过程中最重要的性状之一，离层（Abscission zone）是控制水稻落粒性的重要部位。与野生稻相比，栽培稻的离层发育不完整，导致其落粒性降低。我们的研究发现ObSH3基因是短舌野生稻W1411中的一个关键落粒基因。进一步的研究表明，ObSH3基因对水稻离层的发育具有重要作用，且能够正向调控水稻另外两个主效落粒基因SH4和qSH1的表达。酵母双杂交结果表明ObSH3基因与已克隆的落粒基因qSH1存在蛋白互作关系。根据上述结果，初步绘制了ObSH3基因参与的水稻落粒性遗传调控网络。通过构建3个基因的不同突变体组合，并对其落粒性进行考察，结果表明ObSH3/sh4/qsh1 变异组合的收获指数最高，为生产上培育落粒性适中、适于机械化收割的水稻品种提供了理论参考。