含有MALECTIN结构域的类受体蛋白激酶调控板栗淀粉积累的分子机理

板栗坚果中主要以积累淀粉为主，淀粉的生物合成直接决定其产量和品质特征。本研究通过不同组织和坚果发育期的时空表达分析，筛选确定响应ABA或蔗糖信号且与淀粉积累相关的类受体蛋白激酶CmMRLKs，挖掘候选互作候选互作蛋白，利用双分子荧光互补、Pull-Down 和等温滴定微量热技术，验证CmMRLK38/39与GAPDH在体内体外的互作；通过构建CmMRLK38/39 超表达和沉默载体转化板栗愈伤组织，经ABA和蔗糖处理，检测转基因愈伤和植株中CmMRLK38/39基因和淀粉合成相关基因的表达、CmMRLK38/39与GAPDH酶活性和淀粉含量等。确定CmMRLKs与GAPDH互作调控淀粉积累的分子机制。为调控板栗产量形成、品质调控和分子改良育种奠定理论基础。

Accumulation of starch determines chestnut seeds yield and quality. How starch is synthesized and accumulated in chestnut represents a core question. This research will study the temporal and spatial expression of genes in different organs. We will identify the receptor-like protein kinases containing the MALECTIN domain (MRLKs) that are in response to ABA and sugar signaling and shown to control starch accumulation in Castanea mollissim. Interactors of CmMRLKs will also be studied. Previously we have shown that CmMRLK38/39 interact with GAPDH through a yeast two hybrid assay. In this study, we will test whether this interaction occurs in vitro and in vivo by using BiFC, Pull-Down and iTC methods. Silencing and overexpression of CmMRLK38/39 mutants will be obtained via a callus transgenic system that established in our lab. Transcriptome profiling, to be specific the starch biosynthesis genes, in WT and CmMRLK38/39 mutants will be compared upon ABA and/or sugar treatment. Meanwhile, we will measure the enzymatic activity of GADPH and CmMRLKs, and quantify starch accumulation in WT and CmMRLK38/39 mutants. This could answer whether CmMRLKs interact with GADPH to regulate starch accumulation and shed light on genetic modification to improve chestnut yield and quality.