PA1控制水稻绒毡层降解与花药发育的机理研究

Rice (Oryza sativa) is one of the most important crops in our country and the model plant of monocotyledons research in the molecular biology, besides that rice mutants are the most important materials in gene function research. It will play an important role in revealing the rice development, especially the process of flower development by related rice mutant of flower development.. We isolated a mutant line pa1 in our T-DNA insertion mutant library, showing the pollen sterility. Histological examination of pa1 mutant showed that delayed degeneration of tapetum and defects in formation of male gametophytes were caused by inhibition of the tapetal PCD process. Successful complementary test verified that the loss-of-function the PA1 was the reason for male sterility of pa1. Preliminary studies revealed that PA1 was specifically expressed in panicle. These results implied that PA1 has particular significance for pollen development, rendering it great potential for its application in rice improvement. This project intends to carry out TUNEL experiment to analyze the differentially fragmented DNA sequences and a TEM analysis will help us to check the difference of tapetum subcellular structure; an extensive study on PA1 function utilizing multiple discipline approaches, including spatio-temporal distribution analysis, subcellular localization, RNA-seq and Y2H, aiming to elucidate the molecular mechanism underlying PA1 controlling pollen development and rice yield, to lay a solid theoretical base for the gene application in agricultural biotechnology.

水稻是重要粮食作物之一，水稻花粉发育与产量息息相关，且杂交育种也离不开对雄性不育系的利用。因此，研究花粉育性机理，对提高杂交育种及水稻产量具有重要意义。本研究通过筛选突变体库得到花粉败育突变体pa1；对pa1的细胞学分析显示造成花粉败育的原因是花药绒毡层PCD受到抑制和推迟；通过图位克隆成功获得了PA1基因，并发现PA1基因在水稻幼穗中具有特异性的高表达，这些结果暗示PA1在水稻花粉发育中发挥重要功能。因此，开展系统的PA1基因功能研究对进一步揭示花粉发育过程的分子机制具有重要意义。本项目拟采用TUNEL方法检测野生型和突变体DNA片段化差异及透射电镜分析绒毡层细胞中亚细胞结构差异，进一步确定绒毡层细胞PCD过程差异；通过时空表达分析、亚细胞定位、RNA-seq和酵母双杂等方法对PA1功能展开深入研究，以最终阐明PA1控制水稻花粉发育的分子机制，为其在水稻产量和生产应用奠定理论基础。

水稻花药发育是由一系列复杂的生物学过程组成，其中绒毡层细胞对花药发育起重要作用。绒毡层细胞需要经历一个细胞程序性死亡的过程来为花粉的成熟提供必需的营养物质。但是，人们对绒毡层细胞PCD过程以及这个过程是如何影响花粉发育的调控机理知之甚少。我们鉴定到一个花粉不育突变体post (pollen sterility)。研究发现，post突变体花药绒毡层细胞PCD过程受到抑制，使绒毡层降解受到抑制和推迟，导致花粉的败育。POST基因编码一个全新的蛋白，仅存在于被子植物中。POST主要在花药发育的第8和9时期的绒毡层和小孢子中表达。转录组结果表明，POST可能通过脂质代谢过程的参与绒毡层PCD。post突变体除了雄性不育外，其颖壳出现了明显增大，暗示了细胞死亡和细胞分裂存在着一定关联，这一发现为深层揭示绒毡层程序化细胞死亡过程提供了全新的思路。有意思的是，精细调控POST不仅可以保持其花粉育性，且能增大水稻颖壳，增加库的大小，进而提高籽粒产量。因此，POST不仅是一个新的调控花粉发育的关键因子，也为作物源库改良的潜在基因，为水稻产量改良提供了一种新的策略。