The Ciliopathy Protein CC2D2A Associates with NINL and Functions in RAB8-MICAL3-Regulated Vesicle Trafficking.
The Ciliopathy Protein CC2D2A Associates with NINL and Functions in RAB8-MICAL3-Regulated Vesicle Trafficking.
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DOI：
10.1371/journal.pgen.1005575
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发表时间：
2015-10
期刊：
影响因子：
4.5
通讯作者：
van Wijk E
中科院分区：
文献类型：
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作者：
Bachmann-Gagescu R;Dona M;Hetterschijt L;Tonnaer E;Peters T;de Vrieze E;Mans DA;van Beersum SE;Phelps IG;Arts HH;Keunen JE;Ueffing M;Roepman R;Boldt K;Doherty D;Moens CB;Neuhauss SC;Kremer H;van Wijk E
纤毛病是一组由初级纤毛功能障碍引起的人类疾病，初级纤毛是一种普遍存在的微管细胞器，参与细胞外信号向细胞的转导。该功能需要受体和通道在睫状膜中的集中，这是通过复杂的运输机制（部分由小 GTP 酶 RAB8 控制）以及通过位于睫状室入口处的过渡区的分选来实现的。过渡区基因 CC2D2A 的突变会导致相关的 Joubert 和 Meckel 综合征，这是两种典型的以中枢神经系统畸形为特征的纤毛病，并导致各种模型中多种蛋白质的纤毛定位丧失。 CC2D2A 和其他过渡区蛋白控制蛋白质进入纤毛的精确机制以及它们如何与传入货物的囊泡运输联系起来仍然很大程度上未知。在这项工作中，我们将中心体蛋白 NINL 确定为 CC2D2A 的物理相互作用伙伴。 NINL 与 CC2D2A 部分共定位于纤毛基部，斑马鱼中 ninl 敲低会导致光感受器外节丢失、视蛋白错误定位和囊泡积累，类似于 cc2d2a-/- 表型。此外，cc2d2a-/-胚胎中的部分ninl敲低增强了突变体的视网膜表型，表明体内存在遗传相互作用，在来自Joubert综合症队列的患者中发现了一个例证。与斑马鱼 cc2d2a 突变体类似，ninl 突变体显示出改变的 Rab8a 定位。对 NINL 相关相互作用组的进一步探索发现了 MICAL3，一种已知与 Rab8 相互作用并在囊泡对接和融合中发挥重要作用的蛋白质。总之，这些数据支持 CC2D2A 与 NINL 结合为纤毛定向货物囊泡提供对接点的模型，表明过渡区蛋白可以控制纤毛区室的蛋白质含量的机制。纤毛病是一组由初级纤毛功能障碍引起的疾病，初级纤毛是参与信号转导的普遍细胞器。 CC2D2A 突变会导致两种纤毛病：Joubert 综合征和 Meckel 综合征，并导致纤毛蛋白定位丧失。位于纤毛过渡区的 CC2D2A 控制纤毛蛋白组成的机制及其与传入货物的囊泡运输的联系仍然很大程度上未知。在这里，我们确定了一系列将 CC2D2A 与由小 GTPase RAB8 控制的囊泡运输联系起来的物理相互作用，提出了一种新模型，其中 CC2D2A 为睫状室入口处的纤毛结合囊泡提供了特定的对接点。我们首先将 NINL 确定为 CC2D2A 的物理和遗传相互作用伴侣，表明两种蛋白在纤毛入口处共定位，并证明 Ninl 或 Cc2d2a 的缺失会导致斑马鱼相似的视网膜表型，包括 Rab8 的错误定位。我们进一步鉴定出 MICAL3（一种已知结合 RAB8 的蛋白质）作为另一个 NINL 相互作用伙伴，从而将 CC2D2A 与 RAB8A 控制的运输联系起来。最后，我们描述了一个患有 Joubert 综合征的个体，其中 CC2D2A 和 NINL 突变的组合导致表型增强，说明了检测到的相互作用对疾病的影响。
Ciliopathies are a group of human disorders caused by dysfunction of primary cilia, ubiquitous microtubule-based organelles involved in transduction of extra-cellular signals to the cell. This function requires the concentration of receptors and channels in the ciliary membrane, which is achieved by complex trafficking mechanisms, in part controlled by the small GTPase RAB8, and by sorting at the transition zone located at the entrance of the ciliary compartment. Mutations in the transition zone gene CC2D2A cause the related Joubert and Meckel syndromes, two typical ciliopathies characterized by central nervous system malformations, and result in loss of ciliary localization of multiple proteins in various models. The precise mechanisms by which CC2D2A and other transition zone proteins control protein entrance into the cilium and how they are linked to vesicular trafficking of incoming cargo remain largely unknown. In this work, we identify the centrosomal protein NINL as a physical interaction partner of CC2D2A. NINL partially co-localizes with CC2D2A at the base of cilia and ninl knockdown in zebrafish leads to photoreceptor outer segment loss, mislocalization of opsins and vesicle accumulation, similar to cc2d2a-/- phenotypes. Moreover, partial ninl knockdown in cc2d2a-/- embryos enhances the retinal phenotype of the mutants, indicating a genetic interaction in vivo, for which an illustration is found in patients from a Joubert Syndrome cohort. Similar to zebrafish cc2d2a mutants, ninl morphants display altered Rab8a localization. Further exploration of the NINL-associated interactome identifies MICAL3, a protein known to interact with Rab8 and to play an important role in vesicle docking and fusion. Together, these data support a model where CC2D2A associates with NINL to provide a docking point for cilia-directed cargo vesicles, suggesting a mechanism by which transition zone proteins can control the protein content of the ciliary compartment. Ciliopathies are a group of disorders caused by dysfunction of primary cilia, ubiquitous organelles involved in signal transduction. Mutations in CC2D2A cause two ciliopathies, Joubert and Meckel syndromes, and result in loss of ciliary protein localization. The mechanism by which CC2D2A, located at the ciliary transition zone, controls ciliary protein composition and its link to vesicular trafficking of incoming cargo remain largely unknown. Here, we identify a series of physical interactions linking CC2D2A to vesicular trafficking controlled by the small GTPase RAB8, suggesting a new model, whereby CC2D2A provides a specific docking point for ciliary-bound vesicles at the entrance to the ciliary compartment. We first identify NINL as a physical and genetic interaction partner of CC2D2A, show that both proteins co-localize at the entrance to the cilium and demonstrate that absence of Ninl or Cc2d2a result in similar retinal phenotypes in zebrafish, including mislocalization of Rab8. We further identify MICAL3, a protein known to bind RAB8, as another NINL interaction partner, thus linking CC2D2A to RAB8A-controlled trafficking. Finally, we describe an individual with Joubert syndrome, in whom combined CC2D2A and NINL mutations result in an enhanced phenotype, illustrating the impact of the detected interaction on the disease.