以WRKY10/IKU通路为核心的控制早期胚乳生长和最终种子大小的遗传和表观机理研究

In flowering plants, the seed is produced by a double fertilization event, and is composed of the endosperm and the embryo, surrounded by maternal integuments. The endosperm controls the supply of maternal nutrients to the embryo and plays a central role in seed growth. Endosperm growth and consequently final seed size could be affected by the epigenetic regulation such as parental genome dosage imbalance. Paternal genome dosage excess results in bigger seeds while maternal genome dosage excess results in smaller seeds. The parental conflict theory hypothesized that effectors controlling endosperm growth could be imprinted and account for the impact of parental genome dosage imbalance on seed size. However, little is known about how the epigenetic regulations including imprinting genes affect seed development. .The iku mutants mimic the phenotype of maternal genome dosage excess, however, none of IKU genes are imprinted or clearly linked with epigenetic regulations. We have reported that the cytokinin degradation gene, CKX2 is regulated by IKU pathway and epigenetic regulations, however the cytokinin pathway accounts for only a part of small seed phenotype of iku mutant. In this project, we will further explore the cellular and molecular mechanisms of seed size control by IKU pathway. Our preliminary results showed that we identified a set of putative downstream targets of IKU pathway, in which the genes encoding the cell wall loosing and degradation enzymes are significantly enriched. These founding could explain the specific syncytial nuclear divisions found in early endosperm development, which are never reported before. Based on parental conflict theory, seed size control genes could be imprinting expressed. We design a novel strategy including three ecotypes instead two in traditional method to screen the maternal imprinting genes that specifically expressed in early development endosperm. We also design genome wide analysis to identify the cross talk between IKU pathway and epigenetic regulations. There is high conservation in both molecular level and morphology level during the early development stage of organisms. Thus our finding from early stage of endosperm development of Arabidopsis could be valuable for other plants including economically important crops.

胚乳像胎盘一样为胚的发育和萌发提供营养和合适的环境，是协调种子生长发育的中心，也是表观遗传调控的主要靶标。以WRKY10为代表的IKU通路调控早期胚乳发育和种子大小，但IKU基因未予预期一样与种子大小的表观调控联系起来。在前期工作的基础之上，本项目拟结合生物信息学方法，深入研究IKU通路调控胚乳生长和种子发育的细胞和分子生物学机理。除已知细胞分裂素途径外，初步分析表明IKU通路和表观调控还可能共同控制一个有上百个基因的网络，包括激活细胞壁降解途径以保持胚乳发育早期的同步核分裂和胚乳体积快速增长。本项目将深入探索表观遗传学与IKU基因网络互作的分子机理。为克服传统双亲法无法在发育早期分析基因印记的缺点，创新性引入第三亲本来鉴定胚乳特异性表达基因的印记表达现象。胚乳的早期发育像其它生物体早期发育一样非常保守，因此本项目关于拟南芥早期胚乳研究结果有望为其他作物遗传改造提供新的线索和理论支持。

种子大小是重要农艺性状之一，在包括拟南芥和各种作物中都克隆了一系列影响种子大小的关键基因。但是这些基因大部分都是通过母体组织如种皮，果皮或者颖壳影响种子最终大小，而种子（主要指双受精产物之一胚乳）对种子的生长发育以及与母体组织的营养信号交流等研究相对较少。以WRKY10为代表的IKU基因是少有的已知调控早期胚乳发育并影响最终种子大小的关键遗传信号通路。表观遗传也早就被报道通过调控胚乳的发育来影响最终种子的大小。通过执行本项目，我们发现IKU/WRKY10通路和表观遗传共同调控至少上百个具有不同功能的下游基因来控制种子发育和最终大小。尤其有意思的是，我们发现一个下游基因IDG10是移动蛋白，特异性的在胚乳中表达，但表达产物却被运送到母体组织种皮中，可能通过氧化还原状态调控种皮细胞壁成分进而影响种子的大小。通过该项目的执行，我们还开发了一种简单而全新的全基因组印记基因筛选和鉴定方法，该方法可以鉴定绝大多数传统确定的印记基因，远优于过去已发表的方法。基于该方法我们发现，虽然印记基因和种子大小调控有较为密切的关系，但是并不与亲本冲突学说一致，有力的支持了新的印记基因是胚乳表观特殊表观状态的副产物理论。