AP2家族转录因子SH6调控水稻落粒的分子机制研究

Research on molecular mechanisms of shattering is a great subject to direct modern breeding of rice adapting to the mechanized harvesting.To date,only 5 genes（qSH1,SH4/SHA1,SHAT1,OsCPL1 and SH5) for rice seed shattering were cloned, of which SH4 and qSH1 were key for regulating shattering in rice domestication. We cloned a SH6 for difficult seed shattering encoding a AP2 family transcription factor with dominant inheritance, and was testified by complementary functions. qRT-PCR results showed that SH6 and SH4 may be in the same regulatory route, and SH6 maybe suppress the expression of SH4. Based on the good research bases, three important problems will be resolved in the project. (1) Clear the basic function of SH6 by methods such as RNAi, In situ hybridization, and qRT-PCR et al. (2) Clarify the regulating relationship of SH6 and SH4 and interaction protein of SH6, and therefore how to regulate the formation of abscission layer leading to seed shattering by inheritance analysis of double mutant, ChIP, EMSA Yeast two-hybrid, BiFC et al. (3) Analyze whether existing a common site in SH6 between indica and japonica rice for controlling seed shattering. Consequently, the project will be of great value for explicating the molecular mechanisms of regulating shattering by SH6 and for breeding novel rice varieties to adapt to rice mechanized harvesting.

落粒分子机制研究对指导适合机械化生产的水稻现代化育种有重大意义。目前仅克隆了5个水稻落粒基因qSH1、SH4/SHA1、SHAT1、OsCPL1和SH5，其中qSH1和SH4在水稻驯化中最为关键。我们图位克隆了一个新的显性控制水稻难落粒的AP2家族转录因子SH6，并得到功能互补。qRT-PCR表明，SH6和SH4可能是同一通路的基因，且SH6可能抑制SH4的表达。本项目拟在此良好的研究基础上，重点解决三个问题，（1）以RNAi、原位杂交、qRT-PCR等方法明确SH6的基本功能；（2）以双突变体遗传验证、ChIP、EMSA、酵母双杂交、BiFC等实验手段明确SH6与SH4调控关系及SH6的互作蛋白，从而解析水稻离层分化而导致落粒的分子机制。（3）分析SH6是否在籼、粳亚种间存在共同位点的进化差异。因而本研究对明晰水稻落粒性分子机理及适合机械化收获的新品种培育具有十分重要的意义。

随着农村劳动力向二、三产业转移，水稻传统生产的劳力短缺及成本上升矛盾非常突出。水稻机械化收获迫在眉睫。然而，目前适合机收品种很少。尤其籼稻区机收落粒损失率达8%-10%。可见适合机收的水稻新品种选育势在必行。本项目已明确显性难落粒基因SH6的生物学功能，为定位于细胞核的编码AP2结构域蛋白，具有转录自激活活性，在根和12cm穗和离层中高表达，SH6的靶基因为细胞骨架结合蛋白RMD，可能通过RMD 调控水稻穗部枝梗细胞壁成分调控离层从而调控水稻落粒性，为不同于SH4-SHAT1-qSH1调控落粒的新途径，但还需进一步完善。SH6 基因的 cds 序列80bp 处保守序列“GCAGCA”的缺失与否可能是籼、粳稻品种遗传分化的一个关键位点，据该位点的缺失与否基本可区分籼粳两大类品种（系），当该序列存在时，多数为粳稻品种，表现为难落粒；当缺失时，多数为籼稻品种，且表现为易落粒。此外，本项目已构建籼型恢复系西恢18为背景的SH6难落粒单片段代换系Z983，以易落粒不育系110A与Z983配组的组合110A/ Z983的产量（8333.20 kg.Hm-2）比重庆生产对照品种渝香203（7671.30 kg.Hm-2）增产显著，达8.63个百分点，难落粒性显著提高，其落粒拉力（0.95N）显著高于渝香203的0.61N和110A/西恢18的0.85N。表明以显性SH6改造籼稻品种的易落粒性切实可行，在适合机械化收获品种培育上潜力巨大。