新中心体蛋白MPHOSPH9在纤毛形成过程中的功能分析

Primary cilium is an important structure for sensing signals and regulating physiological functions of cells, but the formation and regulation mechanisms are still largely unknown. MPHOSPH9 is a new centrosomal protein with few reports about its function. Our results showed that MPHOSPH9 was localized to the distal and proximal ends of both mother and daughter centrioles, and was removed from the distal end of the mother centriole during ciliogenesis induced by serum starvation. Knockdown MPHOSPH9 induced ciliogenesis even in non serum-starved RPE-1 cells. Furthermore, our preliminary data indicated that MPHOSPH9 could interact with CEP97, an important regulator in ciliogenesis. In this project, we will further investigate the relationship between MPHOSPH9 and CEP97, and the molecular mechanisms underlying their regulation of ciliogenesis. Besides, we will try to screen other MPHOSPH9-interacting proteins and reveal the protein network that regulate ciliogenesis. Also, we would like to uncover the function of MPHOSPH9 in embryonic development by creating the MPHOSPH9 knockout mice. We hope that this project will contribute to understand better the formation and regulation of cilia, and explore the molecular mechanisms of human genetic diseases caused by ciliogenesis defects.

纤毛是感知外界信号调控细胞生理功能的重要结构，但其自身的形成和调节机制还很不清楚。MPHOSPH9为一个功能几乎未知的新中心体蛋白。我们前期结果表明MPHOSPH9定位于母、子中心粒的远端和近端，且在血清饥饿诱导细胞生长纤毛时从母中心粒远端消失。用RNA干扰敲低其表达量则导致非血清饥饿状态下的RPE-1细胞形成纤毛。进一步的结果显示MPHOSPH9与调控纤毛生长的CEP97相互作用。本项目将研究MPHOSPH9与CEP97的关系，探讨它们对纤毛形成的调节机制；并进一步寻找与MPHOSPH9相互作用的其它可能的蛋白，以揭示在纤毛形成过程中特别是起始阶段的蛋白调控网络。此外，还将建立MPHOSPH9基因敲除小鼠，从个体水平阐明其通过调控纤毛形成影响胚胎发育的机制。本项目的实施将对纤毛发生的机制有更深入地了解，并进一步揭示纤毛生长缺陷相关的遗传性疾病的分子机制。

本项目主要研究了MPP9调控初级纤毛生长的机制。初级纤毛从母中心粒（基体）远端延长生长，介导多种信号通路的转导，因此初级纤毛在个体发育和疾病发生过程中起着十分重要的作用。CP110-CEP97复合体是初级纤毛生长的负调控因子，关于其调控初级纤毛生长的机制还不是十分清楚。本项目研究发现KIF24招募MPP9定位于母中心粒远端，MPP9在母中心粒远端形成环状结构；MPP9通过结合CEP97招募CP110-CEP97复合体定位于母中心粒远端。无论是在处于增殖期的细胞还是小鼠肾脏中，MPP9的缺失都会导致初级纤毛形成异常。在初级纤毛生长起始阶段，MPP9会被TTBK2（Tau Tubulin Kinase 2）磷酸化，随后磷酸化的MPP9通过泛素-蛋白酶体系统降解，MPP9的降解进一步促进CP110-CEP97复合体从母中心粒远端移除，最终使初级纤毛异常增多。因此，MPP9通过调节CP110-CEP97复合体在母中心粒远端的定位，实现对初级纤毛生长的调控作用。