新型GhDRP1(Drought Response Protein1) 调控棉花应答干旱的分子网络解析及育种利用评价

MAPK cascade plays important roles in responding to abiotic stresses for plants. In our previous study,23 association loci associated with seven drought-related traits were obtained based on GWAS（Genome-wide association study）method, and 50 differential expressed genes were selected as candidate ones combined with RNA-Seq data ,among of which one gene had a significant association with plant height trait under drought condition, named GhDRP1，which encods a MPK, then we created the overexpression(OE) and silencing (SI) GhDRP1 cotton materials, and found that OE plants were sensitive to drought stress, but resistance to drought treatment when down-regulated its expression. Yeast-two hybrid experiments found that GhDRP1 interacts with GhHT1( High leaf temperature 1)，and GhHT1interacts with GhOST1（Stomatal opening factor 1）,indicating that GhDRP1 plays key roles during cotton responding to drought stress. Based on these results, this project will further reveal the molecular mechanism in which GhDRP1 involved and how to respond to drought stress in cotton, and explore its drought resistance breeding value combined with these cotton germplsams. This study will broaden the acknowledge of molecular mechanism for cotton responding to drought stress, especially theoretical basis about MAPK cascade regulating drought resistance in cotton.

MAPK级联途径在调控植物应对非生物胁迫过程中起着重要作用。在前期研究中，基于全基因组耐旱性状关联分析，获得7个耐旱相关性状共23个关联位点。结合转录组分析，发现50个候选基因在干旱条件下显著差异表达。其中一个与株高极显著关联的干旱应答基因，命名为GhDRP1,该基因编码MPK蛋白。我们创制了过量和干扰表达该基因的转基因棉花材料，发现过表达GhDRP1对干旱敏感，而下调表达则显著提高耐旱性。酵母双杂发现，GhDRP1和GhHT1( High leaf temperature 1)互作，GhHT1与GhOST1（Stomatal opening factor 1）互作，表明GhDRP1在棉花抗旱中扮演重要角色。本项目将进一步深入揭示GhDRP1参与棉花应答干旱胁迫的分子机理，探究其抗旱育种价值。通过本研究可以拓宽对棉花应答干旱胁迫分子机理的认识，尤其是MAPK级联调控调节棉花抗旱的理论基础。

在前期研究中，基于全基因组耐旱性状关联分析，获得7个耐旱相关性状共23个关联位点，筛选到50个候选基因，包含NAC、WRKY、锌指蛋白等基因和脂质代谢相关的基因，如脂转移蛋白LTP4、GDSL等。其中一个与相对株高性状极显著关联的干旱应答基因，命名为GhDRP1,该基因编码MPK蛋白。创制了过量和干扰表达该基因的转基因棉花材料，发现干扰GhDRP1植株雌雄不育，无法产生后代；过表达GhDRP1对干旱敏感，而VIGS下调表达该基因的幼苗则提高了耐旱性。进一步利用酵母双杂交、Split-LUC和pull down实验筛选和验证了GhDRP1与MAPKKK蛋白GhHT1相互作用并体外相互磷酸化，进一步利用转录组、共表达分析和磷酸化蛋白组学分析发现二者的下游互作蛋白为GhWRKY40a, 而GhWRKY40a的下游靶标基因为GhABF2,并对这些基因参与棉花抗旱的生物功能进行了鉴定，本研究揭示了GhDRP1-GhHT1-GhWRKY40a-GhABF2模块调节棉花耐旱性的分子机理。同时，本项目也对另一个干旱关联基因GhLTP4的抗旱功能和分子机理进行了研究。通过本研究可以拓宽对棉花应答干旱胁迫分子机理的认识，尤其是对MAPK调控调节棉花抗旱的理论基础。