MKS-NPHP module proteins control ciliary shedding at the transition zone

Ciliary shedding occurs from unicellular organisms to metazoans. Although required during the cell cycle and during neurogenesis, the process remains poorly understood. In all cellular models, this phenomenon occurs distal to the transition zone (TZ), suggesting conserved molecular mechanisms. The TZ module proteins (Meckel Gruber syndrome [MKS]/Nephronophtysis [NPHP]/Centrosomal protein of 290 kDa [CEP290]/Retinitis pigmentosa GTPase regulator-Interacting Protein 1-Like Protein [RPGRIP1L]) are known to cooperate to establish TZ formation and function. To determine whether they control deciliation, we studied the function of 5 of them (Transmembrane protein 107 [TMEM107], Transmembrane protein 216 [TMEM216], CEP290, RPGRIP1L, and NPHP4) in Paramecium. All proteins are recruited to the TZ of growing cilia and localize with 9-fold symmetry at the level of the most distal part of the TZ. We demonstrate that depletion of the MKS2/TMEM216 and TMEM107 proteins induces constant deciliation of some cilia, while depletion of either NPHP4, CEP290, or RPGRIP1L prevents Ca2+/EtOH deciliation. Our results constitute the first evidence for a role of conserved TZ proteins in deciliation and open new directions for understanding motile cilia physiology.

纤毛脱落（ciliary shedding）可发生于单细胞生物乃至后生动物（metazoans）中。尽管该过程在细胞周期及神经发生阶段不可或缺，但其具体调控机制仍尚未完全阐明。在所有已报道的细胞模型中，该现象均发生于过渡区（transition zone, TZ）的远端，提示其分子机制具有进化保守性。已知过渡区模块蛋白包括Meckel Gruber综合征（MKS）、肾消耗病（NPHP）家族蛋白、290kDa中心体蛋白（CEP290）以及色素性视网膜炎GTP酶调节因子相互作用蛋白1样蛋白（RPGRIP1L），这些蛋白协同参与过渡区的形成与功能维持。为探明这些保守蛋白是否调控纤毛脱落过程，我们在草履虫（Paramecium）中对其中5种蛋白——跨膜蛋白107（TMEM107）、跨膜蛋白216（TMEM216）、CEP290、RPGRIP1L及NPHP4——的功能开展了系统性研究。实验结果显示，所有靶蛋白均被招募至正在生长的纤毛过渡区，并以9倍对称的方式定位于过渡区的最远端区域。我们证实，敲低MKS2/TMEM216与TMEM107蛋白会导致部分纤毛持续发生脱落；而敲低NPHP4、CEP290或RPGRIP1L中的任意一种，则可阻断Ca²+/乙醇诱导的纤毛脱落。本研究首次为保守型过渡区蛋白在纤毛脱落过程中发挥功能性作用提供了直接实验证据，为理解运动性纤毛的生理机制开辟了全新研究方向。