组蛋白H4上K20位点的甲基化是最普遍的表观遗传修饰之一。然而，H4K20甲基化的调控机制尚不完全明确。申请人此前发现了一类新的组蛋白H4K20去甲基化酶KDM9。KDM9家族成员包括线虫中的DPY-21及从果蝇到哺乳动物中保守的ROSBIN。在线虫中，DPY-21的失活导致X染色体上H4K20单甲基化的上调与基因表达的抑制，并引起染色质高级构象的变化。预实验结果表明ROSBIN在小鼠胚胎干细胞与多种胚胎组织中广泛表达。然而，ROSBIN在高等动物中的生物学功能目前尚属未知。申请人拟利用多种技术系统性地阐明小鼠ROSBIN对H4K20甲基化及基因表达的影响，并通过分析基因表达变化与染色质高级构象之间的关联，深入理解ROSBIN在基因调控中的功能和作用机制。本研究将为未来进一步研究ROSBIN的细胞生物学功能提供基础，并将推动对高等生物发育过程中表观遗传调控机制的全面理解。

H4K20 methylation is one of the most abundant chromatin modifications in eukaryotes and has been implicated in myriad of nuclear functions. However, the mechanisms that regulate different H4K20me states (H4K20me1/me2/me3) are not well understood. We have recently discovered a novel histone demethylase family, KDM9, that specifically converts H4K20me2 to H4K20me1. KDM9 demethylases include C. elegans protein DPY-21 and the ROSBIN proteins that are conserved from flies to mammals. Selective inactivation of DPY-21 in C. elegans eliminates H4K20me1 enrichment on X chromosomes，elevates X-linked gene expression and disrupts Mbp-scale higher-order chromosome structure. Our preliminary results have demonstrated that mouse ROSBIN is expressed in mouse ES cells and a wide range of embryonic tissues. However, the biological functions of ROSBIN in higher eukaryotes are currently unknown. In this project, we will examine the impacts of mouse ROSBIN on H4K20 methylation, gene expression and higher-order chromosome structure by employing a combination of biochemistry, cell biology and genomics approaches. Moreover, we will investigate the functional interplay between higher-order chromosome structure and H4K20 methylation during gene regulation. The proposed studies will provide key insights in understanding the cellular and physiological functions of ROSBIN and promote the understanding of epigenetic regulation during development in higher eukaryotes.