Ku蛋白调控microRNA加工成熟机制的研究

人类Ku蛋白是由Ku70和Ku80两条多肽链通过非共价键紧密结合形成的异二聚体，是一种进化保守蛋白。Ku蛋白在双链DNA断裂（DSBs)的非同源末端结合（NJEJ）修复、端粒结构的维持、肿瘤的发生发展等途径中发挥重要作用。但是，目前所有功能研究基本依赖于Ku蛋白的DNA结合特性。课题申请人带领课题组发现： 1）RNA-IP和深度测序技术鉴定出Ku蛋白与pri-miRNA及pre-miRNA结合； 2）Ku蛋白的表达水平直接影响上述microRNA的表达含量； 3）Ku蛋白能够与双链的RNA结合，包括pre-miRNA和miRNA二聚体； 4）Ku蛋白能够解旋miRNA二聚体，具有RNA解旋酶活性。 因此，本课题将围绕Ku蛋白的RNA结合作用和RNA解旋酶活性，深入开展Ku蛋白多层次多水平参与调控microRNA成熟加工的机制研究；揭示Ku蛋白调控microRNA加工成熟这一全新生物学功能。

Human Ku is a heterodimer of two polypeptides, Ku70 and Ku80, and is evolutionarily conserved. Ku is required for the non-homologous end joining (NHEJ) pathway of DNA repair, telomeric length maintenance， tumorigenesis and tumor development. Until now, all the basic functions research only depend on the DNA binding characteristics of Ku. However, we have sufficient evidence to prove that: 1) Ku bind with some pri-miRNA(primary miRNA) and pre-miRNA (precursor miRNA) sequences except telomerase and IRES thorough RNA-IP and deep sequencing experiments. These sequences contain typical stem ring double chain structure. 2) Ku protein expression level directly affects the binding microRNA expression contents. 3) Ku protein could bind double-stranded RNA, including pri-miRNA, pre-miRNA and miRNA dimmers. 4) Ku protein has RNA helicase activity. Therefore, depending on the RNA binding property and RNA helicase activity of Ku, we will carry out intensive research of the role of Ku in microRNA biogenesis at multilevel. We will explore the new unknown function of Ku protein in different subcellular distribution at different cell stress state, and reveal the biological significances of Ku regulation on microRNA biogenesis.