水稻落粒性调控基因SSH1的克隆与功能分析

Seed shattering is an important agronomic trait affecting crop yield, and precision regulation of seed shattering will be helpful for production management and reduction of yield loss in modern agriculture. Some important QTLs/genes associating with seed shattering have recently been identified in rice, however, their regulatory pathways still remain unknown. To identify new regulators of seed shattering, a wild rice introgression line with the seed shattering habit was mutagenized by ethyl methanesulfonate (EMS) to generate a library, in which a mutant with dramatic reduction of seed shattering, referred to as suppression of shattering 1 (ssh1), was identified. Further genetic analysis revealed that the non-shattering of ssh1 was controlled by a single recessive gene. On this basis, the aims of the proposed study are to isolate the SSH1 gene regulating seed shattering using map-based cloning approach, identify the interaction genes and genome-wide binding sites of SSH1 through an integrated strategy that combined bioinformatics and molecular biology analysis, and then construct the genetic network involving in the development of abscission zone and seed shattering. The results of the study will not only shed light on the genetic mechanism underlying seed shattering in rice, but also have great potential in crop improvement for avoiding yield loss.

落粒性是影响作物产量的重要农艺性状，其精准调控有助于现代农业的生产管理和减少产量损失。虽然近年来克隆多个与水稻落粒性相关的重要QTL/基因，但是其复杂的调控网络仍不清楚。为进一步揭示水稻落粒性的遗传调控，本项目前期利用EMS诱变自然落粒的野生稻渗入系构建突变体库，并筛选到一个落粒性下降的突变体ssh1（suppression of shattering 1）。遗传分析表明该突变表型受一对隐性核基因控制。本项目拟在此基础上，采用图位克隆策略分离水稻落粒性调控基因SSH1，并验证其功能；进一步结合生物信息学和多种分子生物学分析方法，鉴定SSH1的互作基因和全基因组的结合位点，初步绘制其参与水稻落粒性遗传调控网络。研究结果不仅有助于揭示水稻落粒性调控的分子机制，而且将为水稻品种落粒性的分子选育提供重要参考。

水稻落粒性是影响产量的重要性状之一。本研究利用EMS诱变野生稻落粒渗入系构建突变体库，并从中筛选到一个落粒性降低的突变体ssh1（suppression of shattering 1）。通过组织学分析发现，野生型在颖花和果柄连接处可形成完整的离层结构，而突变体ssh1离层发育异常且维管束变粗。遗传分析结果表明，突变体ssh1落粒性降低的表型受一对隐性核基因控制。利用MutMap定位策略，将SSH1基因定位于第7染色体短臂。定位区间序列和基因表达分析发现，在突变体ssh1中LOC_Os07g13170（SNB基因）第9内含子3′末端存在一个由C到A的单碱基突变（SNV6）。通过遗传转化实验证明LOC_Os07g13170即SSH1基因。SSH1 mRNA和蛋白表达水平分析发现，单碱基突变SNV6可导致mRNA异常剪接，全长转录本含量减少，功能蛋白丰度降低，进而导致离层发育异常和落粒性下降。进一步研究发现，SSH1可通过绑定qSH1和SH5的启动子正向调控二者表达，进而降低木质素合成相关基因的表达，最终抑制离区木质素沉积，影响水稻落粒性。产量相关性状调查发现，除落粒性降低外，突变体ssh1还表现为籽粒增长，千粒重增加。进一步以优良籼稻品种93-11为遗传背景构建ssh1突变基因近等基因系，其粒长和千粒重显著增加，表明ssh1突变基因在水稻育种中具有一定的应用潜力。综上所述，AP2转录因子SSH1/SNB在水稻小穗发育过程中扮演了十分重要的角色，具有影响离层发育和颖花发育的多效性。