Drosha/DGCR8复合物调控microRNA加工成熟过程的结构与功能研究

MicroRNA（miRNA）作为真核生物基因表达调控的重要途径一直是研究热点。其在转录水平、转录后修饰、翻译水平等各方面参与调节一些重要基因的表达。目前已在多种癌症中发现相关miRNA、以及miRNA加工成熟过程关键蛋白因子的异常表达。因此，深入了解miRNA表达调控的机制对于疾病诊疗、药物靶向研发等具有重要意义。.由基因组上特定区域转录生成的初级miRNA（pri-miRNA）需经过核内与胞质两步切割反应才能形成成熟miRNA。其中，核酶Drosha与伴侣蛋白DGCR8组成的microprocessor复合物催化完成第一步加工反应，然而其内在的机理并不明确。本项目拟采用X-射线晶体学手段，解析Drosha/DGCR8与pri-miRNA复合物三维结构，并结合生化功能实验，阐明Drosha/DGCR8蛋白复合物与底物miRNA相互作用的关键因素，从而揭示miRNA早期加工成熟的分子机制。

MicroRNA (miRNA) offers an efficient way for gene regulation in eukaryotes. It is widely studied for its important roles in expression repression of many genes that may cause diseases. MicroRNAs regulate gene expression not only at the transcriptional level, but also at post-transcriptional and translational levels. In various diseases especially for cancers, dysregulation of certain types of miRNAs has been identified, as well as key protein components that function in miRNA processing regulation. Therefore, it is quite worthful to dissect the mechanism of how miRNA biogenesis is regulated and this will provide useful information for diagnosis and drug discovery..Pri-miRNA is primary transcribed from certain genomic regions. Generally, it will undergo two steps of processing to produce the mature miRNA, which occur in the nucleus and cytoplasm, respectively. The first processing step is catalyzed by an RNase III enzyme Drosha in complex with its partner DGCR8, which is called the microprocessor complex. However, the details of this site-specific processing reaction are still largely unknown. This project aims to investigate the mechanism of how Drosha/DGCR8 complex specifically recognizes and processes pri-miRNAs. Using X-ray crystallography, we plan to solve the structure of Drosha/DGCR8-miRNA complex. These findings will allow us to identify the key domains and RNA motifs that determine the complex formation and pri-miRNA recognition. Moreover, biochemical and functional studies will be performed to clarify the action mode of Drosha, thus lead us to gain insights into the molecular mechanism of pri-miRNA processing.