Follit John A, San Agustin Jovenal T, Jonassen Julie A, Huang Tingting, Rivera-Perez Jaime A, Tremblay Kimberly D, Pazour Gregory J
Program in Molecular Medicine, University of Massachusetts Medical School, Biotech II, Worcester, Massachusetts, United States of America.
Department of Microbiology and Physiological Systems, University of Massachusetts Medical School, Worcester, Massachusetts, United States of America.
PLoS Genet. 2014 Feb 20;10(2):e1004170. doi: 10.1371/journal.pgen.1004170. eCollection 2014 Feb.
The primary cilium is a sensory organelle, defects in which cause a wide range of human diseases including retinal degeneration, polycystic kidney disease and birth defects. The sensory functions of cilia require specific receptors to be targeted to the ciliary subdomain of the plasma membrane. Arf4 has been proposed to sort cargo destined for the cilium at the Golgi complex and deemed a key regulator of ciliary protein trafficking. In this work, we show that Arf4 binds to the ciliary targeting sequence (CTS) of fibrocystin. Knockdown of Arf4 indicates that it is not absolutely required for trafficking of the fibrocystin CTS to cilia as steady-state CTS levels are unaffected. However, we did observe a delay in delivery of newly synthesized CTS from the Golgi complex to the cilium when Arf4 was reduced. Arf4 mutant mice are embryonic lethal and die at mid-gestation shortly after node formation. Nodal cilia appeared normal and functioned properly to break left-right symmetry in Arf4 mutant embryos. At this stage of development Arf4 expression is highest in the visceral endoderm but we did not detect cilia on these cells. In the visceral endoderm, the lack of Arf4 caused defects in cell structure and apical protein localization. This work suggests that while Arf4 is not required for ciliary assembly, it is important for the efficient transport of fibrocystin to cilia, and also plays critical roles in non-ciliary processes.
初级纤毛是一种感觉细胞器，其缺陷会导致多种人类疾病，包括视网膜变性、多囊肾病和出生缺陷。纤毛的感觉功能需要特定的受体靶向质膜的纤毛亚结构域。有人提出Arf4在高尔基体复合体对运往纤毛的货物进行分类，并被认为是纤毛蛋白运输的关键调节因子。在这项研究中，我们发现Arf4与纤维囊素的纤毛靶向序列（CTS）结合。敲低Arf4表明，由于稳态CTS水平不受影响，它对于纤维囊素CTS运输到纤毛不是绝对必需的。然而，当Arf4减少时，我们确实观察到新合成的CTS从高尔基体复合体运输到纤毛的过程出现延迟。Arf4突变小鼠胚胎致死，在节点形成后不久于妊娠中期死亡。在Arf4突变胚胎中，节点纤毛看起来正常且功能正常，能够打破左右对称。在这个发育阶段，Arf4在内脏内胚层中的表达最高，但我们在这些细胞上未检测到纤毛。在内脏内胚层中，Arf4的缺失导致细胞结构和顶端蛋白定位缺陷。这项研究表明，虽然Arf4不是纤毛组装所必需的，但它对于纤维囊素向纤毛的有效运输很重要，并且在非纤毛过程中也起着关键作用。
