Center for Cancer Research, NCI-Frederick, National Institutes of Health, Laboratory of Cellular and Developmental Signaling, Frederick, Maryland 21702.
Department of Biomedical Sciences, University of Sheffield, Sheffield S10 2TN, United Kingdom.
J Biol Chem. 2019 Oct 18;294(42):15418-15434. doi: 10.1074/jbc.RA119.008615. Epub 2019 Aug 29.
The primary cilium is a cellular sensor that detects light, chemicals, and movement and is important for morphogen and growth factor signaling. The small GTPase Rab11-Rab8 cascade is required for ciliogenesis. Rab11 traffics the guanine nucleotide exchange factor (GEF) Rabin8 to the centrosome to activate Rab8, needed for ciliary growth. Rabin8 also requires the transport particle protein complex (TRAPPC) proteins for centrosome recruitment during ciliogenesis. Here, using an MS-based approach for identifying Rabin8-interacting proteins, we identified C7orf43 (also known as microtubule-associated protein 11 (MAP11)) as being required for ciliation both in human cells and zebrafish embryos. We find that C7orf43 directly binds to Rabin8 and that C7orf43 knockdown diminishes Rabin8 preciliary centrosome accumulation. Interestingly, we found that C7orf43 co-sediments with TRAPPII complex subunits and directly interacts with TRAPPC proteins. Our findings establish that C7orf43 is a TRAPPII-specific complex component, referred to here as TRAPPC14. Additionally, we show that TRAPPC14 is dispensable for TRAPPII complex integrity but mediates Rabin8 association with the TRAPPII complex. Finally, we demonstrate that TRAPPC14 interacts with the distal appendage proteins Fas-binding factor 1 (FBF1) and centrosomal protein 83 (CEP83), which we show here are required for GFP-Rabin8 centrosomal accumulation, supporting a role for the TRAPPII complex in tethering preciliary vesicles to the mother centriole during ciliogenesis. In summary, our findings have revealed an uncharacterized TRAPPII-specific component, C7orf43/TRAPPC14, that regulates preciliary trafficking of Rabin8 and ciliogenesis and support previous findings that the TRAPPII complex functions as a membrane tether.
初级纤毛是一种细胞传感器，可探测光、化学物质和运动，对形态发生素和生长因子信号转导很重要。小 GTP 酶 Rab11-Rab8 级联反应对于纤毛发生是必需的。Rab11 将鸟嘌呤核苷酸交换因子（GEF）Rabin8 运输到中心体以激活 Rab8，Rab8 对于纤毛生长是必需的。Rabin8 在纤毛发生过程中还需要运输颗粒蛋白复合物（TRAPPC）蛋白向中心体募集。在这里，我们使用基于 MS 的方法来鉴定 Rabin8 相互作用蛋白，发现 C7orf43（也称为微管相关蛋白 11（MAP11））在人类细胞和斑马鱼胚胎中都是纤毛发生所必需的。我们发现 C7orf43 直接与 Rabin8 结合，并且 C7orf43 敲低会减少 Rabin8 纤毛前中心体的积累。有趣的是，我们发现 C7orf43 与 TRAPPII 复合物亚基共沉淀，并与 TRAPPC 蛋白直接相互作用。我们的研究结果表明 C7orf43 是 TRAPPII 特异性复合物的组成部分，在此称为 TRAPPC14。此外，我们表明 TRAPPC14 对于 TRAPPII 复合物的完整性不是必需的，但介导 Rabin8 与 TRAPPII 复合物的结合。最后，我们证明 TRAPPC14 与远端附属蛋白 Fas 结合因子 1（FBF1）和中心体蛋白 83（CEP83）相互作用，我们在这里表明它们对于 GFP-Rabin8 中心体的积累是必需的，支持了 TRAPPII 复合物在纤毛发生过程中充当将前纤毛小泡系泊到母中心粒的作用。总之，我们的发现揭示了一个未被描述的 TRAPPII 特异性成分 C7orf43/TRAPPC14，它调节 Rabin8 的前纤毛运输和纤毛发生，并支持之前的发现，即 TRAPPII 复合物作为膜系泊物发挥作用。
