Bengueddach Hakim, Lemullois Michel, Aubusson-Fleury Anne, Koll France
Institute for Integrative Biology of the Cell (I2BC), CEA, CNRS, Université Paris Sud, Université Paris-Saclay, 1 Avenue de la Terrasse, 91198 Gif sur Yvette, France.
Cilia. 2017 Mar 30;6:6. doi: 10.1186/s13630-017-0050-z. eCollection 2017.
The development of a ciliary axoneme requires the correct docking of the basal body at cytoplasmic vesicles or plasma membrane. In the multiciliated cell three conserved proteins, FOR20, Centrin 2, and Centrin 3 participate in this process, FOR20 and Centrin 2 being involved in the assembly of the transition zone. We investigated the function of two other evolutionary conserved proteins, OFD1 and VFL3, likely involved in this process.
In , a single gene encodes OFD1, while four genes encode four isoforms of VFL3, grouped into two families, VFL3-A and VFL3-B. Depletion of OFD1 and the sole VFL3-A family impairs basal body docking. Loss of OFD1 yields a defective assembly of the basal body distal part. Like FOR20, OFD1 is recruited early during basal body assembly and localizes at the transition zone between axoneme and membrane at the level of the microtubule doublets. While the recruitment of OFD1 and Centrin 2 proceed independently, the localizations of OFD1 and FOR20 at the basal body are interdependent. In contrast, in VFL3-A depleted cells, the unanchored basal bodies harbor a fully organized distal part but display an abnormal distribution of their associated rootlets which mark their rotational asymmetry. VFL3-A, which is required for the recruitment of Centrin 3, is transiently present near the basal bodies at an early step of their duplication. VFL3-A localizes at the junction between the striated rootlet and the basal body.
Our results demonstrate the conserved role of OFD1 in the anchoring mechanisms of motile cilia and establish its relations with FOR20 and Centrin 2. They support the hypothesis of its association with microtubule doublets. They suggest that the primary defect of VFL3 depletion is a loss of the rotational asymmetry of the basal body which specifies the sites of assembly of the appendages which guide the movement of basal bodies toward the cell surface. The localization of VFL3 outside of the basal body suggests that extrinsic factors could control this asymmetry.
纤毛轴丝的发育需要基体在细胞质囊泡或质膜上正确对接。在多纤毛细胞中，三种保守蛋白FOR20、中心蛋白2和中心蛋白3参与此过程，FOR20和中心蛋白2参与过渡区的组装。我们研究了另外两种可能参与此过程的进化保守蛋白OFD1和VFL3的功能。
在[具体生物]中，单个基因编码OFD1，而四个基因编码VFL3的四种异构体，分为两个家族，VFL3 - A和VFL3 - B。OFD1和唯一的VFL3 - A家族的缺失会损害基体对接。OFD1的缺失导致基体远端部分组装缺陷。与FOR20一样，OFD1在基体组装早期被招募，并定位在轴丝和膜之间微管双联体水平的过渡区。虽然OFD1和中心蛋白2的招募是独立进行的，但OFD1和FOR20在基体上的定位是相互依赖的。相比之下，在VFL3 - A缺失的细胞中，未锚定的基体具有完全组织化的远端部分，但显示其相关小根的分布异常，这些小根标志着它们的旋转不对称性。VFL3 - A是中心蛋白3招募所必需的，在基体复制的早期阶段短暂出现在基体附近。VFLA - 3定位在横纹小根和基体之间的连接处。
我们的结果证明了OFD1在运动纤毛锚定机制中的保守作用，并确定了它与FOR20和中心蛋白2的关系。这些结果支持了它与微管双联体相关的假设。结果表明，VFL3缺失的主要缺陷是基体旋转不对称性的丧失，而这种不对称性决定了附属物的组装位点，这些附属物引导基体向细胞表面移动。VFL3在基体外部的定位表明外在因素可能控制这种不对称性。
