玉米X1基因可变剪接调控籽粒大小的分子机理研究

Kernel size and weight are the important factor of maize yield components, and related formation process involves complex regulatory network. Alternative splicing is an important mechanism of expression regulation, which is widespread in eukaryotic organisms. Recent studies have shown that alternative splicing may play an important role in kernel development, and exploiting the role of alternative splicing in the formation of kernel traits is one of the challenging frontiers. Based on the RNA sequencing data of maize population from our previous study, we determined associations between alternative splicing variations and kernel size-related traits at the genome-wide level. A key splicing event was identified and located in the X1 gene was significantly associated with grain length, 100-kernel volume, and 100-kernel weight. The ortholog of X1 gene (IDN2/RDM12) is involved in the RNA-mediated DNA methylation (RdDM) pathway in Arabidopsis and functions during seed development. On this basis, this study aims to determine the biological function of X1 gene via molecular genetic experiments, and to clarify the mechanism of X1 gene splicing controlling kernel development and its influence on kernel trait formation. The results of this study will not only provide an important reference for understanding the role of gene alternative splicing during the maize improvement on agronomic traits, but also provides theoretical support for the further genetic improvement in maize.

籽粒大小与粒重是玉米产量的重要构成因子，其形成过程涉及到复杂的调控网络。可变剪接是一种重要的表达调控机制，在真核生物体内广泛存在。近期研究表明可变剪接在籽粒发育过程中可能担当重要角色，探索可变剪接机制在籽粒性状形成中的作用是具有挑战的前沿问题之一。本研究前期利用玉米关联群体转录组测序数据，在全基因组水平上进行可变剪接变异与籽粒大小性状的关联分析，鉴定到X1基因的可变剪接与粒长、百粒体积及百粒重都存在极显著关联。X1直系同源基因IDN2/RDM12在拟南芥中参与RNA介导的DNA甲基化过程，在种子的发育期间发挥重要功能。在此基础上，本研究将进一步通过分子遗传实验确定X1基因的生物学功能，明确X1基因可变剪接影响籽粒发育与籽粒性状形成的作用机理。本研究结果将为理解基因可变剪接在玉米重要农艺性状形成过程中的作用机理提供重要参考，也将为玉米的遗传改良提供理论支撑。

籽粒大小与粒重是玉米产量的重要构成因子，其形成过程涉及到复杂的调控网络，挖掘控制粒重的相关基因揭示其功能具有重要意义。项目组利用玉米关联群体转录组测序数据，在全基因组水平上进行可变剪接变异与籽粒大小性状的关联分析，发现X1基因的可变剪接与粒长、百粒体积及百粒重都存在极显著关联。在此基础上，项目组通过遗传学、分子生物学和生物信息学等技术手段，对X1基因的功能开展深入研究。首先构建X1转基因过表达材料，表型分析发现，过表达X1基因会导致玉米籽粒粒重显著降低，粒长变短。相反，X1基因突变体的籽粒粒长和粒重较野生型都显著性增加，表明X1对籽粒大小及粒重存在负调控作用。对X1基因5'UTR区的剪接变异进行分析，发现T03转录本的UTR序列与T01转录本相比会显著性降低目标基因的蛋白表达水平，造成含有不同转录本的材料在粒型和粒重等方面出现差异。对X1基因的表达模式分析发现，其主要在授粉后12天的籽粒中高表达。亚细胞定位结果表明，X1蛋白在细胞质和细胞核表达。X1直系同源基因IDN2/RDM12在拟南芥中参与RNA介导的DNA甲基化过程，在种子的发育期间发挥重要功能。酵母双杂交实验发现，X1蛋白与FDM同源基因ZM03G05120存在互作。综合上述结果，本项目克隆了一个影响玉米籽粒大小及粒重的负调控因子X1，并初步解析了X1基因调控粒型与粒重的分子机理，为玉米产量性状遗传改良提供重要的理论依据。