玉米ZmbHLHA转录因子调控淀粉合成机制的研究

Starch content and composition directly determine the productivity and quality of maize. Considerable research has been conducted to better understand the function of these genes which determine the maize content and composition, but the regulating mechanism of these function genes, especially the regulating mechanism of transcription factor for starch synthesis gene not well understood. A maize basic helix-loop-helix transcription factor, ZmbHLHA, was isolated based on the transcriptome analysis and Genome Wide Association Studies in previous study. Over expression of this gene can lead to the difference of starch content and structure in transgenic rice and maize. In addition, mutants of ZmbHLHA of maize plants were also obtained. Further study also demonstrated that ZmbHLHA also regulates the maize starch synthesis. In this project, We will also character the biological function of ZmbHLHA in starch synthesis, through the characterization of enzyme activities of starch-synthesizing relating enzymes, starch content and starch-synthesizing relating physiological indexes in ZmbHLHA transgenic maize plants and mutant. This project will also reveal the molecular functional mechanism of ZmbHLHA by screening the target gene and interaction protein, interacting assay in vitro and ZmbHLHA with the interacting protein on the transcriptional activity of target gene. Moreover, the expression level of starch-synthesizing related genes will be analyzed in ZmbHLHA transgenic maize plants and mutant, The results of this project will not only enrich our knowledge of the regulating mechanism in maize starch synthesis, but also supply theory for the breeding new maize varieties.

淀粉含量和成分直接决定着玉米产量和品质，决定玉米淀粉含量和成分的基因已大量研究，然而这些功能基因的作用机制尤其是转录因子对淀粉合成基因的调控机制还不清楚。项目组基于转录组和基因组关联分析结果克隆了一个玉米转录因子ZmbHLHA基因，过量表达该基因可以改变水稻和玉米种子淀粉含量和结构，对玉米ZmbHLHA突变体分析同样发现对淀粉合成的调控功能。本研究在此基础上拟通过ZmbHLHA过表达与突变体玉米种子发育过程中淀粉合成相关酶活、淀粉含量的动态差异及淀粉结构等生理生化指标分析，明确ZmbHLHA调控淀粉合成的生理生化作用特征；通过ZmbHLHA靶基因及互作蛋白的筛选、互作蛋白对靶基因转录活性的分析和ZmbHLHA调控淀粉合成途径的分析，明晰其调控基因与作用网络，从而解析ZmbHLHA调控淀粉合成的机制。本项目研究结果不仅为解析淀粉合成调控机理提供新证据，也为高产优质玉米新品种选育奠定理论基础。

淀粉含量和成分直接决定着玉米产量和品质，决定玉米淀粉含量和成分的基因已大量研究，然而这些功能基因的作用机制尤其是转录因子对淀粉合成基因的调控机制还不清楚。通过GWAS和转录组数据，我们从玉米中克隆了一个bHLH家族基因，ZmbHLHA。ZmbHLHA定位于细胞核且具有转录活性，组织表达模式分子其主要在胚乳中表达。过表达玉米植株籽粒表现出淀粉含量增加、可溶性糖和还原性糖含量下降；通过扫描电镜发现转基因玉米籽粒淀粉结构出现了异常。此外，转基因植株中淀粉合成相关基因的表达量明显提高。酵母单杂交显示ZmbHLHA可以直接结合到淀粉合成基因ZmAGPS1a, ZmAGPL1, ZmGBSSI, ZmSSIIa, ZmISA1 和ZmISA3 的启动子上，进一步研究发现在烟草细胞中ZmbHLHA可以激活 ZmGBSSI and ZmISA1的表达。总之，这些结果证明ZmbHLHA是淀粉合成中的核心调控因子，有助于改良玉米胚乳淀粉含量的应用研究。