Lv Bo, Wan Lei, Taschner Michael, Cheng Xi, Lorentzen Esben, Huang Kaiyao
Key Laboratory of Algal Biology, Institute of Hydrobiology, Chinese Academy of Sciences, Wuhan, Hubei 430072, China.
University of Chinese Academy of Sciences, Beijing 100039, China.
J Cell Sci. 2017 May 1;130(9):1662-1674. doi: 10.1242/jcs.200758. Epub 2017 Mar 16.
Cilia are microtubule-based organelles and perform motile, sensing and signaling functions. The assembly and maintenance of cilia depend on intraflagellar transport (IFT). Besides ciliary localization, most IFT proteins accumulate at basal bodies. However, little is known about the molecular mechanism of basal body targeting of IFT proteins. We first identified the possible basal body-targeting sequence in IFT46 by expressing IFT46 truncation constructs in an mutant. The C-terminal sequence between residues 246-321, termed BBTS3, was sufficient to target YFP to basal bodies in the strain. Interestingly, BBTS3 is also responsible for the ciliary targeting of IFT46. BBTS3::YFP moves bidirectionally in flagella and interacts with other IFT complex B (IFT-B) proteins. Using IFT and motor mutants, we show that the basal body localization of IFT46 depends on IFT52, but not on IFT81, IFT88, IFT122, FLA10 or DHC1b. IFT52 interacts with IFT46 through residues L285 and L286 of IFT46 and recruits it to basal bodies. Ectopic expression of the C-terminal domain of IFT52 in the nucleus resulted in accumulation of IFT46 in nuclei. These data suggest that IFT52 and IFT46 can preassemble as a complex in the cytoplasm, which is then targeted to basal bodies.
纤毛是以微管为基础的细胞器，具有运动、传感和信号传导功能。纤毛的组装和维持依赖于鞭毛内运输（IFT）。除了在纤毛中的定位外，大多数IFT蛋白还聚集在基体上。然而，关于IFT蛋白靶向基体的分子机制知之甚少。我们首先通过在一个突变体中表达IFT46截短构建体，鉴定了IFT46中可能的基体靶向序列。246-321位残基之间的C端序列，称为BBTS3，足以将黄色荧光蛋白（YFP）靶向到该菌株的基体上。有趣的是，BBTS3也负责IFT46的纤毛靶向。BBTS3::YFP在鞭毛中双向移动，并与其他IFT复合体B（IFT-B）蛋白相互作用。利用IFT和运动突变体，我们发现IFT46的基体定位依赖于IFT52，而不依赖于IFT81、IFT88、IFT122、FLA10或动力蛋白重链1b（DHC1b）。IFT52通过IFT46的L285和L286残基与IFT46相互作用，并将其招募到基体上。IFT52 C端结构域在细胞核中的异位表达导致IFT46在细胞核中积累。这些数据表明，IFT52和IFT46可以在细胞质中预先组装成一个复合体，然后靶向基体。
