5-甲酰四氢叶酸在玉米籽粒中积累的遗传学基础与分子机制研究

Folates are a group of human essential B vitamins. Folate deficiency can cause serious diseases, such as neural tube malformation, cancer, and hypertension. Thus the accumulation of folates in the edible parts (kernels) in crops is related to human health. 5-Formyltetrahydrofolate (5-F-THF) is the major derivatives of foaltes in maize kernels. In the preliminary work, the folate recombinant inbred lines mapping population had been used to investigate the genetic basic of the accumulation of 5-F-THF in maize kernels. Two quantitative trait loci (QTLs) A and B were obtained. ZmGFT1 had been identified in QTL B by the genome-wide association study. The enzyme activity experiment showed that ZmGFT1 could convert 5-methyltetrahydratefolate into 5-F-THF directly, and the overexpression of ZmGFT1 indicated that ZmGFT1 protein, which contained the excellent allele variation, significantly promoted the production of 5-F-THF in kernels. In addition, the candidate genes in QTL A with the higher contribution were around 2.0 Mb on chromosome 5. On this basis, the proposed project focuses on the regulatory mechanism of folate metabolism in corn kernels, and aims to figure out the molecular mechanism of candidate genes in these QTLs. Through the comprehensive utilization of methods in biochemistry, molecular biology, genetics, and transcriptomics, the molecular function of the candidate genes on the accumulation of 5-F-THF in maize kernels will be explored, and the theoretical basis and technical support on crop breeding by the molecular design to improve the accumulation of folates in crop seeds will also be provided through the implementation of this project.

叶酸是人体必需的B族维生素，摄入不足会造成神经管畸形、癌症、高血压等严重疾病，因此，叶酸在作物籽粒的积累关系到人体的营养健康。玉米籽粒中叶酸的主要形式是5-甲酰四氢叶酸。前期工作中，我们利用重组自交系鉴定了影响5-甲酰四氢叶酸在籽粒积累的主效QTL位点A和B，精细定位了位点B中的目标基因ZmGFT1，ZmGFT1可以直接将5-甲基四氢叶酸转化为5-甲酰四氢叶酸，且含有优良等位变异的ZmGFT1蛋白具有更高的活性，同时贡献率更高的位点A目标基因已经定位在5号染色体的2.0Mb附近。本项目拟在此基础上精细定位并克隆影响5-甲酰四氢叶酸积累的目标基因，继续完善ZmGFT1的功能分析，并综合利用生物化学、分子生物学、遗传学和转录组学等不同研究手段探讨基因影响5-甲酰四氢叶酸在籽粒中积累的遗传学基础和分子机制，为利用分子设计育种提高叶酸在玉米等作物籽粒中的积累水平提供理论依据和技术支撑。

叶酸是人体必需的B族维生素，摄入不足会造成神经管畸形、癌症、高血压等严重疾病，因此，叶酸在作物籽粒的积累关系到人体的营养健康。玉米籽粒中叶酸的主要形式是5-甲酰四氢叶酸。前期工作中，我们利用重组自交系鉴定了影响叶酸在籽粒积累的QTL位点A和B，精细定位了位点B中的目标基因ZmGFT1（ZmCTM），这个基因可以直接将5-甲基四氢叶酸（5-methyltetrahydrofolate, 5-M-THF）转化为更稳定的叶酸衍生物形式，且含有优良等位变异的蛋白具有更高的活性，同时贡献率更高的位点A目标基因已经定位在5号染色体的2.0Mb附近。通过本项目的实施，建立作物叶酸等系列B族维生素高通量检测平台，筛选获得高B族维生素玉米新材料；获得两个控制玉米籽粒5甲酰四氢叶酸（5-formyltetrahydrofolate, 5-F-THF）变异的QTL位点（q5-F-THFa和q5-F-THFb），开发高叶酸基因型的CAPS标记，用于高叶酸玉米的育种；利用生物化学、分子生物学、遗传学和转录组学等不同研究手段探讨基因影响叶酸在籽粒中积累的遗传学基础和分子机制，构建玉米籽粒叶酸转录调控网络，发现发育后期叶酸的积累主要受到氨基酸戊糖嘌呤代谢的共同调控，其中甲硫氨酸合酶2基因ZmMETS2可能是受调控的关键基因之一；基因编辑发现位于q5-F-THFb的基因ZmCTM突变可以增加叶酸含量，突变甲硫氨酸合酶2基因ZmMETS2导致甲硫氨酸含量的增加；证实了高叶酸玉米是可以有效的替代补充片剂来缓解小鼠孕期叶酸的缺乏症；同时还开展叶酸等维生素的其他相关工作。第一资助发表SCI文章1篇，中文核心2篇；第二资助发表SCI论文2篇，中文核心2篇；第三资助发表SCI论文2篇；第四资助发表SCI论文1篇；第五资助发表SCI论文2篇；中文核心1篇；授权发明专利2项；培养博士研究生3名；主持人作为第三完成人获得大北农科技奖创新奖1项。这些工作为深入解析玉米籽粒叶酸积累的分子机制及遗传改良提供了调控关键基因等重要资源，也通过创制高叶酸或高甲硫氨酸玉米新材料为营养强化农业提供材料支撑。