SDG724及其互作蛋白IP71和ROC1L调控水稻基本营养期的分子机制

Rice heading date is an important agronomic trait for determining the cultivation area and harvest seasons, and is affected by genetic pathways that integrate endogenous and environmental signals. To date, most of the researches on the gene regulation network for heading date in rice are related to the photoperiod pathway, and the regulation mechanism of the endogenous pathway that affecting the basic vegetative growth stage remains unclear. The histone methylation transferase SDG724 plays an important role in regulating rice heading date, however, the mechanisms by which SDG724 targets specific genes and delivers methylation information need to be further elucidated. In the previous work, several SDG724 interaction proteins were obtained by yeast library screening. iP71 and rco1l knockout mutants all showed late heading date, which was similar to the phenotype of SDG724 mutant lvp1. Therefore, it was speculated that SDG724 may form protein complexes with IP71 and RCO1L to regulate rice heading date. Based on this speculation, the current application intends to analyze the interaction mode of the SDG724 protein complex, clarify the biochemical function of the complex subunits, explore the biological functions of the SDG724 protein complex in regulating the basic vegetative growth stage of rice, mining and utilize the downstream target genes co-regulated by the protein complex. Through the above research, the project aims to enrich the epigenetic regulation mechanism of rice heading date and lay a foundation for the creation of rice germplasm resources with long-term basic vegetative growth stage.

水稻抽穗期是决定栽培地区与种植季节的重要农艺性状，受整合内在因素和外部环境信号的遗传途径控制。目前研究较为透彻的抽穗期基因调控网络大多与光周期途径有关，影响基本营养生长期的内在途径调控机制尚不明确。组蛋白甲基化转移酶SDG724在调控水稻抽穗期中具有重要作用，但其靶定特异基因并传递甲基化信息的机制有待揭示。前期工作中，通过酵母文库筛选获得多个SDG724互作蛋白，其中ip71和rco1l敲除突变体均表现晚抽穗，与SDG724突变体lvp1表型相似，因此推测SDG724可能与IP71和RCO1L形成蛋白复合物共同调控水稻抽穗。申请人以此为切入点，解析SDG724蛋白复合物互作模式并明确复合物亚基生化功能，探究SDG724蛋白复合物在调控水稻基本营养生长期中的生物学功能，挖掘和利用蛋白复合物协同调控的下游靶基因，丰富水稻抽穗期的表观遗传调控机制，为培育长基本营养生长期的水稻种质资源奠定基础。

水稻的开花时期，农业上称为抽穗期，是决定品种栽培地区与种植季节的重要农艺性状。水稻抽穗期长短由品种的感光性、感温性和基本营养生长性来决定。目前对于影响基本营养生长期的内在途径调控机制尚不明确。本项目主要研究结果如下：（1）通过酵母筛库获得组蛋白甲基化转移酶SDG724的互作蛋白IP71和ROC1L，利用酵母双杂交、LUC、pull-down实验证明了SDG724与IP71和ROC1L在植物体内和体外都相互作用。进一步利用酵母双杂交试验证实SDG724 N端IxRN是水稻SDG724与IP71互作的关键基序；截短的SDG724与ROC1L不互作。利用LUC试验证实ROC1L能够增强SDG724与IP71的互作。ROC1L具有串联PHD结构域，能够非特异性结合H3多肽，并与IP71竞争性结合H3K36me3。（2）sdg724，roc1l和 ip71突变体在长日照和短日照下抽穗期延迟，光周期转移试验表明三者能够延长基本营养生长期并增强对光周期敏感程度。此外，双突变体sdg724 ip71或sdg724 roc1l的抽穗期均晚于单突变体sdg724，ip71或roc1l。（3）利用RNA-Seq和ChIP-Seq挖掘到一个受SDG724-IP71-ROC1L蛋白复合物共同调控的下游靶基因MADS64，该基因突变后同样表现出抽穗延迟的表型。通过本项目的研究，我们阐明了SDG724-IP71-ROC1L通过直接调控MADS64基因从而延长水稻基本营养生长期，创制了一系列长基本营养生长期的遗传材料，为培育广适性水稻品种提供理论依据和基因资源。