纤毛/鞭毛是一类存在于原生动物和脊椎动物细胞表面的细胞器，除执行运动功能外，还执行感受和分泌功能，后二者主要依赖于纤毛中特有的膜蛋白，目前已知很多重要的纤毛膜蛋白都被N-糖基化修饰，但N-糖基化对纤毛膜蛋白的运输及功能调节中的作用还不清楚。本项目首先利用分离纯化得到的鞭毛和鞭毛膜来鉴定鞭毛膜蛋白的糖基化位点，筛选糖基化鞭毛膜蛋白缺失突变体，分析其与鞭毛相关的表型。然后突变糖基化位点，分析N-糖基化对鞭毛膜蛋白运输和功能的影响。其次利用与糖基化蛋白结合的Concanavalin A或荧光蛋白融合技术，分析糖基化膜蛋白在鞭毛中的运动规律，以及与Intraflagellar transport的关系。最后，利用N-糖基化通路的突变体和抑制剂，研究N-糖基化对纤毛膜蛋白的运输以及对纤毛的感受等功能的影响。这些研究将为揭示N-糖基化在维持初级纤毛的正常结构和功能中的作用，以及与纤毛病的关系打下基础。

Cilia/flagella are widely distributed in protist and vertebrates, functioning as motile, sensory and secretory organelles. The last two categories of functions are dependent on the ciliary membrane proteins. It has been demonstrated that some of them are N-glycosylated; however, the roles of N-glycosylation on trafficking and regulation the function of flagellar/ciliary membrane proteins are unknown. To address these questions, first, we will isolate the pure flagella and flagellar membrane, and use them as materials to identify the N-glycosylation sites of these flagellar membrane proteins, then isolate their insertional mutants and analyze their flagellar phenotypes. In addition, we will mutate the N-glycosylation sites to determine their roles in ciliary trafficking and in regulation their functions in cilia. Second, we will use the fluorescence labeled Con A or fuse GFP with the glycosylated flagellar membrane proteins to study their dynamics in flagella. The role of Intraflagellar Transport (IFT) in their dynamics will be determined by using a temperature sensitive mutants of anterograde motor. Finally, we will take advantage of inhibitor and mutants of N-glycosylation pathway to pinpoint their roles in modulation of sensory and motile functions of cilia and in regulation the distribution in flagellar membrane proteins by using the quantitative proteomic techniques. These studies will speed our understanding the roles of N-glycosylation in maintaining the structure and function of primary cilia.