植物PcG催化亚基CLF与SWN基因组结合特异性的机制研究

PcG复合体主要通过催化目标基因位点的组蛋白H3第27位赖氨酸三甲基化（H3K27me3）从而抑制基因表达，维持细胞属性。CLF和SWN是植物PcG复合体中高度同源的催化亚基，参与决定了不同器官的发育以及生长阶段的转换。而遗传学研究表明了CLF和SWN可以特异性地调控植物的生长发育过程，表明两者存在功能的差异，但具体的分子机制还未完全清楚。为了探索CLF-PcG和SWN-PcG复合体之间的功能区隔与联系，本课题拟通过染色质免疫共沉淀偶联高通量测序的方法研究CLF和SWN全基因组的结合差异以及催化H3K27me3的异同，从而阐明CLF和SWN在全基因组范围内的相互作用模式。拟通过免疫共沉淀偶联质谱的方法筛选CLF和SWN特异结合蛋白，研究差异蛋白对CLF和SWN基因组结合的调控，以期解析CLF和SWN基因组结合特异性的分子机制，完善PcG复合体的调控网络，为作物改良提供潜在的理论依据。

Polycomb-group proteins (PcG) are highly conserved chromatin-associated factors involved in the regulation of crucial developmental programs in multicellular organisms. Polycomb repressive complex 2 (PRC2) is the major regulatory complex of PcG that regulates gene expression by catalyzing trimethylation of histone H3 lysine 27 (H3K27me3), a repressive histone marker ultimately leading to transcriptional repression. In Arabidopsis thaliana, CURLY LEAF (CLF) and SWINGER (SWN) are two major H3K27 methyltransferases and core components of PRC2. CLF and SWN function to safeguard the appropriate expression of key developmental regulators throughout the plant life cycle. The earlier genetic analyses demonstrate that CLF and SWN have unique, shared, and redundant functions. Recent researches have advanced our knowledge of the biological roles and the regulation of the activities of CLF and SWN, but the molecular mechanisms underlying their differential functions at the genome-wide scale are poorly understood. In this study, we seek to investigate the molecular mechanisms underlying CLF and SWN’s differential roles at the genome-wide scale. To this end, we will perform chromatin immunoprecipitation followed by high-throughput sequencing (ChIP-seq) to identify CLF and SWN’s global targets and examine to what degree their genome-wide occupancy will be affected when CLF or SWN is lost. Then immunoprecipitation coupled with mass spectrometry (IP-MS) will be carried out to screen CLF/SWN-interacting partners and identify CLF/SWN-differential interacting proteins (CSDIPs). To answer whether those CSDIPs contribute to the differential functions between CLF and SWN, we will check the occupancy of CLF and SWN without the presence of CSDIPs, respectively. We will then examine whether the loss-of-function of CSDIPs enhances morphological defects of clf and swn mutants. Taken together, we aim at investigating the genome-wide interplay between CLF and SWN through the combination of global approaches, including ChIP-seq, RNA-seq, and IP-MS, which will provide insight into the understanding of PcG-mediated regulation in plants.