微管结合蛋白WDR62调节有丝分裂纺锤体极微管负端动态性的功能及机制

有丝分裂纺锤体将姐妹染色体平均分配到两个子细胞，其正确组装对于有丝分裂正常进行至关重要。纺锤体组装是一个受各类微管结合蛋白精密调控的过程。微管结合蛋白WDR62定位于纺锤体极，其突变频率在原发性小头症相关基因中排名第二。WDR62突变可导致纺锤体异常，有丝分裂紊乱，并最终引起小头症发生。但WDR62微管结合特性及其在纺锤体组装过程中的具体机制仍未知。本项目拟通过多学科交叉手段研究WDR62调节纺锤体极微管负端动态性的功能和机制。具体来说，通过基于质谱的蛋白质组学方法寻找WDR62上下游分子；结合基因编辑和活细胞成像技术阐明WDR62及其互作蛋白调控纺锤体装配的细节；借助全内反射荧光显微镜观察WDR62体内体外微管结合行为，并体外重组WDR62及其结合蛋白对微管动态性的调控过程。通过对WDR62生化特性与细胞功能的综合阐述，我们将揭示纺锤体组装调控的新机制并为研究其他微管结合蛋白提供新思路。

The bipolar spindle plays an essential role in ensuring faithful chromosome segregation during mitosis. Spindle assembly is a tightly regulated process, which is orchestrated by a variety of microtubule associated proteins (MAPs). WDR62 is a MAP localizing to spindle poles, where microtubule minus ends are enriched. Mutations in WDR62 are the second most common cause of autosomal recessive primary microcephaly (MCPH). Genetic disruptions of WDR62 lead to defects in spindle assembly and aberrant mitotic progression, which may underlie microcephaly. However, the microtubule binding property of WDR62 is yet obscure and how this activity correlates with its cellular functions remains elusive. Here, we aim to understand the precise roles and mechanisms of WDR62 mediated regulation of microtubule minus-end dynamics at spindle poles using multidisciplinary approaches. Specifically, we start this project with identifying binding partners of WDR62 using Mass spectrometry-based proteomics analysis. Subsequently, by combining CRISPR/Cas9 mediated gene editing and live-cell imaging, we will dissect the contributions of WDR62 and its binding partners to the regulation of spindle dynamics. Finally, we will apply in vitro reconstitution analyses to examine precisely how WDR62 and its binding partners modulate microtubule dynamics using purified components. With a comprehensive study of the biochemical properties and cellular functions of WDR62, this project will uncover mechanisms underlying WDR62 mediated regulation of spindle organization and set a new paradigm for dissecting the roles of other spindle associated MAPs.