水稻品种间氧化胁迫耐受性差异的分子机理解析

Oxidative stress is one of the common secondary stresses of various environmental stresses which can dramatically reduce crop yield. It is expected to increase the wide adaptability of crops by improving their oxidative stress tolerance. However, the molecular mechanisms underlying difference in oxidative stress tolerance among rice varieties remain elusive. In our previous study, using the advanced backcross segregation population, we successfully cloned a critical gene RT1 which determines the difference in oxidative stress tolerance among rice varieties. Based on the comprehensive analysis of genetic complementation lines, loss-of-function mutant, overexpression transgenic lines, and near isogenic line, we confirmed that RT1 can effectively improves the oxidative stress tolerance and tolerance to several other stresses in rice. On these basis, we intend to further investigate the molecular mechanisms underlying RT1-mediated antioxidant response in this project, including: to elucidate the mechanism in which RT1 responses to upstream oxidative stress signaling, to dissect the biological function of RT1-interacting protein RIP1, to illustrate the anti-oxidative transcriptional regulation network downstream of RT1, to analysis the regulation of RT1 on different stress signal pathways, and to assess the potential application of RT1 in rice production.

氧化胁迫是不同逆境胁迫导致作物大规模减产的共同次生胁迫之一，改善作物氧化胁迫耐受性有望提高作物的广泛适应性。然而，迄今有关水稻品种间氧化胁迫耐受性差异关键调控基因的鉴定还没有相关报道。在前期研究中，我们利用高世代回交群体克隆到调控水稻品种间氧化胁迫耐受性差异的关键基因RT1。基于遗传互补株系、功能丧失突变体、过表达转基因株系、以及近等基因系的综合分析，我们证实RT1能有效的改善水稻的氧化胁迫耐受性及多个逆境胁迫的耐受性。本项目在此基础上，拟进一步深入分析RT1调控水稻氧化胁迫耐受性的分子机制，包括阐明RT1对上游氧化胁迫信号的响应机制，解析RT1互作蛋白RIP1的生物学功能，明确RT1下游抗氧化胁迫的转录调控网络，分析RT1对不同胁迫信号途径的调控，及评估RT1在水稻生产上的应用潜力。

水稻品种对氧化胁迫的耐受性存在显著差异，调控水稻品种间氧化胁迫耐受性差异的主效基因的鉴定有望应用于水稻胁迫耐受性的改良，从而提高水稻品种对不同环境的广泛适应性。本研究前期鉴定到一个调控品种间氧化胁迫耐受性差异的主效QTL位点ROS Tolerance 1 (RT1)。为了进一步研究RT1介导的氧化胁迫耐受性的分子机制，本项目中我们利用酵母双杂交系统鉴定到了RT1的互作蛋白RIP1，并通过荧光素酶互补成像和免疫共沉淀等实验证实了RT1和RIP1存在相互作用。与RT1类似，RIP1能够在转录水平上响应外界的氧化胁迫刺激，并提高水稻对氧化胁迫的耐受性。此外，基于转录组数据分析我们鉴定到RT1-RIP1下游靶基因RRG1，并通过酵母单杂交、瞬时转录激活、以及基于染色质免疫沉淀(ChIP)的实时定量PCR等实验证实RT1能够直接结合在RRG1的启动子区，从而调控后者的表达。而且RIP1增强了RT1对下游靶基因的激活能力。遗传分析证实过表达RRG1能够明显提高水稻对氧化胁迫的耐受性。此外，RT1在品种间存在基因型分化，我们通过瞬时转录激活实验证实RT1的不同基因型对下游基因的激活功能存在差异，其中粳稻型RT1 激活下游靶基因的活性显著高于aus型的RT1。由此，本研究揭示了不同水稻品种间氧化胁迫耐受性差异的一种分子机制，同时近等基因系的表型分析进一步证实粳稻型RT1能有效提高水稻品种对不同胁迫的耐受性，表明这一位点有望应用于水稻品种广泛适应性的改良。