ICK(也称为CILK1)是位于睫状尖端的丝裂原活化蛋白激酶样激酶。已知其缺乏会导致纤毛伸长并导致人类纤毛病。然而,关于ICK如何运输到睫状尖端知之甚少。我们在这里表明,ICK的C端非催化区域与IFT机械的步行内运输(IFT)-B复合物相互作用,并参与其运输到睫状尖端。此外,全内反射荧光显微镜显示ICK在纤毛内进行双向运动,类似于IFT粒子。ICK敲除细胞的分析表明,ICK缺乏严重损害了IFT颗粒和睫状G蛋白偶联受体的逆行运输。此外,我们发现在ICK敲除细胞中,纤毛蛋白在隆起的纤毛尖端积累,它似乎被撕下并作为细胞外囊泡释放到环境中。各种ICK构建体在ICK敲除细胞中的外源表达表明ICK的IFT依赖性转运,以及其在规范TDY基序的激酶活性和磷酸化,对于ICK功能至关重要。因此,我们明确表明,逆行睫状蛋白运输以及正常的睫状功能都需要运输到睫状尖端的ICK。
ICK (also known as CILK1) is a mitogen-activated protein kinase-like kinase localized at the ciliary tip. Its deficiency is known to result in the elongation of cilia and causes ciliopathies in humans. However, little is known about how
ICK is transported to the ciliary tip. We here show that the C-terminal noncatalytic region of ICK interacts with the intraflagellar transport (IFT)-B complex of the IFT machinery and participates in its transport to the ciliary tip. Furthermore, total internal reflection fluorescence microscopy demonstrated that ICK undergoes bidirectional movement within cilia, similarly to IFT particles. Analysis of
ICK knockout cells demonstrated that
ICK deficiency severely impairs the retrograde trafficking of IFT particles and ciliary G protein-coupled receptors. In addition, we found that in ICK knockout cells, ciliary proteins are accumulated at the bulged ciliary tip, which appeared to be torn off and released into the environment as an extracellular vesicle. The exogenous expression of various
ICK constructs in
ICK knockout cells indicated that the IFT-dependent transport of ICK, as well as its kinase activity and phosphorylation at the canonical TDY motif, is essential for
ICK function. Thus, we unequivocally show that ICK transported to the ciliary tip is required for retrograde ciliary protein trafficking and consequently for normal ciliary function.