植物生殖发育相关的长链非编码RNA功能与调控机制

Sexual reproduction is one of the most essential biological processes and occurs in both animals and plants. It is more crucial for crop plants because of its applications in agriculture. Identification and functional study of reproduction-related genes are of importance in understanding the reproductive processes. We previously performed genome-wide screening and functional study of noncoding RNAs from a model plant species, rice (Oryza sativa), and identified a small noncoding RNA could positively regulate plant embryo development and rice grain yield (Nat Biotechnol, 2013), suggesting that noncoding RNAs are important for crop plant agronomic trait improvement. We have also performed strand-specific whole transcriptome sequencing and systematically identified long noncoding RNAs (lncRNAs) from rice and revealed that a number of rice lncRNAs are highly tissue specific, and a large portion of these RNAs are specifically expressed in conjunction with reproduction-related processes, particularly in anthers and pistils. Based on these previous findings, in this project, we endeavor to further investigate the functions and regulatory mechanisms of the lncRNAs which are specifically expressed in sexual reproduction-related processes. Followings are our main focuses: (1) Systematic screening of potential lncRNAs associated with plant reproductive development; (2) In Vivo investigation of the functions of the aberrantly expressed lncRNAs particularly in anthers (male reproductive organ) and pistils (female reproductive organ). (3) The molecular mechanisms of the lncRNAs mediated regulation networks, especially the RNA binding proteins associated with anthers, pistils and embryo development. The accomplishment of the project could not only reveal the novel function of lncRNAs in plant sexual reproduction, but also provide new resources for the genetic modification of crop plant agronomic traits.

非编码RNA调控生殖发育的生物学功能是当前生命科学研究的重要问题之一。我们前期以水稻为模式材料，获得调控水稻胚胎发育发育和产量的小分子非编码RNA （Nat Biotechnol，2013），揭示了非编码RNA在植物生殖发育中的重要作用，可以作为一类新的遗传资源用于植物农艺性状改良。我们进一步对水稻生殖发育相关长链非编码RNA(lncRNA)进行大规模筛选与鉴定，发现一批在水稻的雌雄配子体中特异性表达lncRNA。本项目拟在此基础上开展以下方面研究：一是筛选鉴定生殖阶段特异性表达lncRNA，分析其基因结构等信息；二是解析生殖发育相关重要lncRNA功能及规律；三是研究这些lncRNA介导生殖调控作用的分子机制及调控网络，本项目的立项和实施，从不同于蛋白质编码基因的角度阐明植物生殖发育过程的规律，阐明lncRNA参与植物雌雄配子体发育等生殖过程的功能，吻合重大研究计划指南方向一的科学目标。

生殖发育是植物最重要的生命活动过程之一，也是作物遗传育种工作中的重要环节；挖掘生殖阶段重要的遗传资源，阐明其功能和机制的解析将为改良水稻产量、育性等农艺性状提供借鉴和指导。本项目重点围绕模式植物水稻生殖发育相关长链非编码RNA功能与调控机制这一关键科学问题，从三个不同方向和层次展开研究：一是筛选鉴定生殖阶段特异性表达ncRNA，特别是lncRNA,探明其时空表达模式；二是解析部分生殖阶段特异性表达ncRNA的功能及对水稻开花和育性的影响；三是研究这些ncRNA介导的调控通路在水稻生殖发育中的作用机制。项目主要进展如下：（1）lncRNA-phasiRNA-靶基因调控网络鉴定及RNA修饰在水稻雄配子发育中的作用研究。水稻等禾本科植物中，一大批lncRNA在生殖阶段特异性表达，并且是生殖阶段特异性表达一类小分子非编码RNA phasiRNA的前体。但是哪类lncRNA产生phasiRNA？下游靶基因是什么？和生殖发育的关系等等问题都是未清楚的。我们通过转录组、小RNA组、降解组学测序和分析，结合实验验证和突变株构建表型分析，揭示了lncRNA-phasiRNA新的调控通路。（2）雌蕊特异性表达的lncRNA的功能机制。我们的前期研究发现，lncRNA XLOC_057324 为一个在雌蕊特异性表达的lncRNA 分子。通过功能解析，揭示了结实率降低主要是受精后胚乳发育缺陷所致，进一步研究发现该lncRNA影响了胚乳的细胞化过程；通过lncRNA结合蛋白的鉴定及作用机制分析，步阐明该lncRNA通过调控DNA解旋酶，影响了三倍体胚乳细胞核的发育和细胞。（3）阐明了生殖阶段表达的lncRNA ALEX对逆境应答的机制，揭示其参与茉莉酸通路介导的应答反应。（4）除了长链非编码RNA， 我们还发现小分子非编码RNA miR528在调控花粉粒后期发育中的功能。miR408在水稻生殖发育中也起了重要的调控作用，miR408介导的OsUCL还影响了花粉管在柱头上的延伸，最终影响了水稻的结实率。该项目已发表论文SCI收录文章10篇（其中8篇为第一标注，2篇为第2标注），包括Nature Communications、PNAS等；申请专利3项；获得广东省自然科学奖一项；对照项目合同书，本项目研究目标已达到。