棉花木质素代谢关键因子的遗传定位与基因功能解析

Verticillium wilt caused by the fungus Verticillium dahliae is the most devastating disease for cotton production in China. Characterization the resistance mechanism and the resistance genes are very important for cotton disease-resistance molecular breeding. In our previous study, lignin metabolism was found play vital roles in cotton response to V. dahliae through whole genome gene expression profile and enzyme activity assay. Moreover, we found that a transcriptional factor, GbERF1-like, could improve the cotton resistance to V. dahliae by regulating the expression level of GhHCT1 and activating the synthesis of lignin. Furthermore, the expression level of GhLac1, which involved in the synthesis of lignin, was found also influence the signal pathways of plant innate system. And overexpression of GhLac1 confer broad-spectrum resistance of cotton to fungus and insect, which inferred that the metabolism of lignin involved in the regulation of plant innate system. However, the mechanism in regulation of lignin synthesis and the key genes involved in lignin synthesis or synthesis regulation are still unclear. In this project, we want to measure the content of total lignin and monomer composition in roots and hypocotyls in cotton seedlings, respectively, and identify the key genetic loci and the key genes in lignin metabolism through genome-wide association study with the complete cotton population genome re-sequencing. Characterization of the key genes involved in lignin metabolism could provide gene resources for germplasm innovation of cotton resistance to V. dahliae.

黄萎病是影响棉花生产的最主要病害。鉴定棉花抗病机制和抗病基因是棉花抗病分子育种的基础。我们前期表达谱分析和酶活检测发现木质素代谢参与了棉花抗黄萎病反应。进一步研究发现GbERF1-like能激活木质素合成基因GhHCT1的表达来增强木质素合成并提高棉花对黄萎病的抗性。而参与木质素合成的基因GhLac1在棉花中的表达水平不仅影响木质素合成，还能影响抗病信号路径。超量表达GhLac1能提高棉花对病害和虫害的广谱抗性，表明木质素代谢参与了棉花免疫系统的调控。但木质素在棉花中的代谢调控机制与关键因子还不清楚。我们拟在已建立的棉花自然群体及已完成的基因组重测序基础上，结合棉花根与下胚轴木质素总含量、木质素单体以及胞壁其他成分的代谢分析，进行全基因组木质素代谢关联分析，鉴定参与木质素代谢或调控的关键遗传位点，解析关键代谢基因或调控基因。项目的完成预期为通过调控木质素代谢来创新棉花抗病种质提供基因资源。

项目主要研究了棉花木质素调控关键因子鉴定及其在抗黄萎病中的作用。通过四年研究，项目完成了棉花自然群体和MAGIC群体的木质素代谢组分析，完成棉花自然群体的全基因组重测序和SNP分型，完成棉花自然群体的茎秆强度测定与分析。开展了棉花自然群体苗期下胚轴重量（干重、鲜重）、细胞壁成分（H、G、S型木质素单体含量）及群体成株期的茎秆强度等性状的全基因组关联分析，定位到与木质素单体代谢相关的候选遗传区间8个，与可溶性酚代谢相关的候选遗传区间1个，与茎秆强度相关的候选遗传区间2个；通过SNP分析，在棉花 A04、A10、和D05染色体上检测到7个与总木质素含量显著相关的SNPs，并且有3个SNPs在多个环境中均检测到。目前正在对这些定位区间的基因进行功能分析，期望鉴定到控制棉花木质素代谢的关键因子。完成了候选基因GhMYB4、GhWRKY41和GhTINY2等在木质素代谢调控及与抗黄萎病关系的功能鉴定。分析发现GhMYB4直接绑定GhC4H-1/2、Gh4CL-4、GhCAD-3、GhLac1等木质素合成相关基因的启动子并抑制其表达负调控木质素合成；GhWRKY41与自身形成正反馈调节环直接激活苯丙烷代谢关键酶，正调控木质素合成与黄萎病的抗性；GhTINY2通过正调控SA的合成，抑制油菜素内酯的合成来激活植物免疫反应和抑制生长发育。棉花木质素代谢关键位点鉴定以及与棉花免疫和发育的互作仍需进一步鉴定。