水稻冠根形成基因的克隆与功能分析

Roots are essential organs for exploiting soil resources such as water and mineral nutrients, and supporting plant growth as well as crop productivity. The rice root system is mainly composed of postembryonic shoot-borne roots termed crown roots, and its numbers play the crucial role in capturing water and mineral nutrients. However, the molecular mechanism of crown root formation in rice remains unclear. The project is proposed to map-based clone a gene for crown root number (CRN) by using the segregation population derived from chromosomal substitution segment lines in rice, and to analyze its biological functions with the integrated approaches in genetics, physiology and molecular biology. Other main activities proposed in this project also include identification and validation of gene regulatory network and signaling pathways that the CRN gene involved in controlling crown root formation, and mining of the natural occurring alleles with its functions in use-efficiency of water and nutrients (such as nitrogen and phosphate) through association mapping with the core collection of rice germplasm. These results will facilitate better understanding of the molecular mechanism of root formation and root architecture in rice, and establish a practical and theoretical foundation for improving water and nutrient use-efficient rice varieties.

根是植物吸收土壤中水分和养分、固定植株、影响植物生长和产量潜力的主要营养器官。冠根是水稻根系的主要组成部分，其数目决定了水稻对土壤水分和养分的吸收利用效率。目前，冠根形成的分子机理尚不清晰。本项目拟利用水稻染色体片段代换系及其衍生群体，采用图位克隆的方法分离克隆一个影响水稻冠根数的基因（CRN），通过遗传学、生理学和分子生物学等技术手段，研究其生物学功能。在表达水平和蛋白水平上分析和验证CRN的互作基因，剖析其参与的分子调控网络。利用水稻核心种质测序及关联分析，鉴定CRN自然发生的等位变异及其对水分和养分(如氮、磷)吸收利用的影响，发掘优异等位基因。预期研究结果将有利于更好地了解水稻根系形成的分子调控机理，为培育水分和养分高效利用的水稻新品种奠定理论和实践基础。

根系形态是影响水分和养分吸收利用效率和产量形成的关键因素。本项目利用水稻染色体片段代换系和突变体等群体开展了水稻根系（冠根）形成相关基因的定位与克隆，分析其自然发生的等位变异及其功能，剖析调控水稻冠根形成的分子机理。项目分离克隆到一个冠根相关基因CR6。该基因编码含一个AP2结构域的转录因子。组织表达谱分析显示，CR6在幼苗、叶片以及穗部的表达水平较低，在成熟叶鞘和茎秆中表达未检测到，而在种子发芽胚和根系中高度表达。在亚洲栽培稻品种中，CR6的启动子和编码区存在较大的变异。利用启动子区的58个SNP对CR6进行单倍型分析，发现栽培稻中CR6存在至少10种单倍型，籼稻的CR6启动子区核苷酸多样性仅为野生稻的十分之一，表明在水稻驯化过程中该基因受到了较强烈的选择。转基因CR6超表达、抑制表达以及T-DNA突变体等试验结果表明，CR6负向调控水稻根长和冠根数目。超量表达CR6抑制根系生长，根长变短、冠根数减少；而下调CR6表达，冠根数目增多。超表达株系的生物量降低、每穗粒数减少、产量降低。转录组分析显示在CR6超表达株系中生长素合成和转运等相关基因的表达受到显著的抑制。与之相一致的是，CR6启动子上存在生长素响应元件，可以与生长素早期响应因子ARF25蛋白互作。CR6蛋白可以与生长素转运因子OsPINi结合，从而抑制其表达；下调OsPINi基因的表达引起水稻根长变短、冠根数减少。综合这些结果表明，CR6通过参与生长素途径调节根系中生长素的运输，负调控水稻根系和地上部植株的生长，最终影响单株产量。本研究结果为深入理解水稻冠根发生和发育的分子遗传机理提供新的见解。