Centre for Chromosome Biology, School of Natural Sciences, National University of Ireland Galway, Galway H91 W2TY, Ireland.
Lunenfeld-Tanenbaum Research Institute, Sinai Health System, 600 University Avenue, Toronto, Ontario M5G 1X5, Canada.
J Cell Sci. 2019 Oct 3;132(19):jcs228486. doi: 10.1242/jcs.228486.
Centrin 2 is a small conserved calcium-binding protein that localizes to the centriolar distal lumen in human cells. It is required for efficient primary ciliogenesis and nucleotide excision repair (NER). Centrin 2 forms part of the xeroderma pigmentosum group C protein complex. To explore how centrin 2 contributes to these distinct processes, we mutated the four calcium-binding EF-hand domains of human centrin 2. Centrin 2 in which all four EF-hands had been mutated to ablate calcium binding (4DA mutant) was capable of supporting NER and was as effective as the wild-type protein in rescuing the UV sensitivity of centrin 2-null cells. However, we found that mutation of any of the EF-hand domains impaired primary ciliogenesis in human TERT-RPE1 cells to the same extent as deletion of centrin 2. Phenotypic analysis of the 4DA mutant revealed defects in centrosome localization, centriole satellite assembly, ciliary assembly and function and in interactions with POC5 and SFI1. These observations indicate that centrin 2 requires calcium-binding capacity for its primary ciliogenesis functions, but not for NER, and suggest that these functions require centrin 2 to be capable of forming complexes with partner proteins.This article has an associated First Person interview with the first author of the paper.
中心体蛋白 2 是一种小型保守的钙结合蛋白，在人类细胞中定位于中心体远端腔。它是有效的初级纤毛发生和核苷酸切除修复（NER）所必需的。中心体蛋白 2 是着色性干皮病组 C 蛋白复合物的一部分。为了探索中心体蛋白 2 如何促进这些不同的过程，我们突变了人中心体蛋白 2 的四个钙结合 EF 手结构域。所有四个 EF 手都被突变以消除钙结合的中心体蛋白 2（4DA 突变体）能够支持 NER，并且在拯救中心体蛋白 2 缺失细胞的 UV 敏感性方面与野生型蛋白一样有效。然而，我们发现 EF 手结构域的任何突变都以与中心体蛋白 2 缺失相同的程度损害了人 TERT-RPE1 细胞的初级纤毛发生。对 4DA 突变体的表型分析显示，在中心体定位、中心粒卫星组装、纤毛组装和功能以及与 POC5 和 SFI1 的相互作用方面存在缺陷。这些观察结果表明，中心体蛋白 2 对其初级纤毛发生功能需要钙结合能力，但不需要 NER，并表明这些功能需要中心体蛋白 2 能够与伴侣蛋白形成复合物。本文有一篇与论文第一作者的第一人称访谈。
