miR390及其靶基因对水稻体内镉转运的分子调控机制研究

Heavy metal contamination of agricultural soils has become increasingly serious in China in recent years. To realize safe production of rice in heavy metal polluted soils, it is necessary to clarify the molecular and physiological mechanisms of heavy metal uptake, translocation and response in rice. miRNAs, an important new class of modulators of gene expression, play important roles in plant adaptations to environmental stresses. However, the knowledge of the specific role of miRNAs in heavy metal stress response is limited. In our previous study, miR390 and its target gene a receptor-like protein kinase (RLK) (Os02g10100) were identified to be Cd-responsive. In this study, Japonica rice Zhonghua 11 is used for test material. The temporal and tissue expression patterns, and subcellular localization of miR390 and RLK will be tested. Rice transgenic technology will be used to investigate their function. Furthermore, the downstream target gene of RLK will be identified. The specific regulatory pathway of miR390 and its target gene RLK in Cd translocation in rice will be investigated. These results will provide a new insight into understanding the molecular mechanism of heavy metal stress response in rice, and an effective strategy for the genetic improvement in rice heavy metal tolerance.

近年来我国耕地重金属污染形势相当严峻。探明水稻对重金属吸收、转运及耐性分子生理机制，对保障水稻安全生产意义重大。miRNA是一类新型的基因表达调控因子，在植物逆境胁迫应答过程中发挥重要作用。然而有关miRNA在重金属胁迫应答中的具体作用机制并不清楚。本项目以水稻中花11为材料，前期研究获知的与镉胁迫应答相关的miR390及其靶基因——富含亮氨酸的类受体蛋白激酶（Receptor-like protein kinase，RLK，Os02g10100）为研究对象，分析miR390及RLK的组织时序表达模式和亚细胞定位，采用转基因手段验证其功能，明确与重金属转运及解毒相关的RLK作用的下游底物，探究它们参与调控水稻对镉转运的具体分子生物学途径，掌握miR390对水稻镉胁迫应答的调控机制，为揭示水稻对重金属耐性分子机制提供理论依据，也为水稻重金属耐性遗传改良提供有效手段。

近年来我国耕地重金属污染形势相当严峻。大量农田受到重金属污染，不仅危害作物生长，也通过食物链危害人体健康。探明水稻对重金属吸收、转运及耐性分子生理机制，对保障水稻安全生产意义重大。有关miRNA及其靶基因在植物重金属胁迫应答中的具体作用机制并不清楚。本项目以水稻中花11为材料，前期研究获知的与镉胁迫应答相关的miR390及其靶基因——富含亮氨酸的类受体蛋白激酶（Receptor-like protein kinase，RLK，Os02g10100）为研究对象，分析miR390及RLK的组织时序表达模式。发现在苗期、分蘖期及穗分化期，RLK主要在叶片中表达。抽穗期、灌浆期及成熟期，RLK主要在稻穗及节中表达。我们采用转基因手段研究miR390及RLK在镉胁迫响应中的功能，发现miR390超量表达水稻和RLK RNAi水稻类似，均增加了对镉毒害的敏感性以及镉向地上部的转运。而RLK的过表达增强了水稻植株对镉胁迫的耐受性，并降低了水稻对镉的积累和转运。另外，我们以RLK的RNAi转基因和野生型水稻植株为材料，利用iTRAQ技术对镉胁迫相关差异蛋白进行分析，共鉴定出289个差异表达蛋白。发现RLK可能在镉胁迫下通过调节水稻的基础代谢、应激反应和氧化还原状态来调控水稻耐镉。利用qRT-PCR验证筛选出了RLK下游调节基因Peroxidase P7，发现其在镉胁迫下表达上升，过量表达Peroxidase P7增加了水稻对镉的耐受性。综上发现miR390-RLK-Peroxidase调控通路在水稻对镉胁迫的响应过程中发挥了重要作用。以上研究可为揭示水稻对重金属耐性分子机制提供理论依据，也为利用转基因技术进行水稻重金属耐性遗传改良提供有效手段。