The primary cilium functions as a mechanical and calcium signaling nexus.
The primary cilium functions as a mechanical and calcium signaling nexus.
复制标题
作者：
Lee KL;Guevarra MD;Nguyen AM;Chua MC;Wang Y;Jacobs CR
初级纤毛是从大多数哺乳动物细胞类型的表面延伸的触角样非运动结构，并且对于包括软骨、骨和肾在内的各种组织中的化学传感和机械传感至关重要。血流诱导的肾上皮细胞内钙离子（Ca 2+）增加依赖于初级纤毛和初级纤毛定位的Ca 2+渗透通道多囊蛋白2（PC 2）和瞬时受体电位香草酸4（TRPV 4）。虽然原纤毛已经牵连在骨细胞mechanotransduction，介导这一过程的分子机制还没有完全理解。我们通过将生物传感器序列与主要纤毛特异性蛋白Arl 13 b的序列融合，将基于荧光共振能量转移（FRET）的Ca 2+生物传感器定向到纤毛。使用这个工具，我们研究了几个Ca 2+渗透性通道的作用，可能介导流量诱导的Ca 2+进入：PC 2，TRPV 4和PIEZO 1。在这里，我们报告的第一个测量的钙离子信号在骨细胞初级纤毛使用FRET为基础的生物传感器融合ARL 13 B。我们发现，流体流动诱导钙增加骨细胞初级纤毛依赖于细胞内钙释放和细胞外钙进入。使用siRNA介导的敲除，我们证明TRPV 4，而不是PC 2或PIEZO 1，介导流动诱导的纤毛Ca 2+增加和负载诱导的考克斯-2 mRNA增加，成骨反应。在这项研究中，我们表明，初级纤毛形成一个Ca 2+微区依赖于Ca 2+进入通过TRPV 4。这些结果表明，由初级纤毛介导的机械转导的机制在不同的组织环境中是不同的。此外，我们预计这项工作是更多研究TRPV 4在机械转导中作用的起点。本文的在线版本（doi：10.1186/s13630-015-0016-y）包含补充材料，可供授权用户使用。
The primary cilium is an antenna-like, nonmotile structure that extends from the surface of most mammalian cell types and is critical for chemosensing and mechanosensing in a variety of tissues including cartilage, bone, and kidney. Flow-induced intracellular calcium ion (Ca2+) increases in kidney epithelia depend on primary cilia and primary cilium-localized Ca2+-permeable channels polycystin-2 (PC2) and transient receptor potential vanilloid 4 (TRPV4). While primary cilia have been implicated in osteocyte mechanotransduction, the molecular mechanism that mediates this process is not fully understood. We directed a fluorescence resonance energy transfer (FRET)-based Ca2+ biosensor to the cilium by fusing the biosensor sequence to the sequence of the primary cilium-specific protein Arl13b. Using this tool, we investigated the role of several Ca2+-permeable channels that may mediate flow-induced Ca2+ entry: PC2, TRPV4, and PIEZO1. Here, we report the first measurements of Ca2+ signaling within osteocyte primary cilia using a FRET-based biosensor fused to ARL13B. We show that fluid flow induces Ca2+ increases in osteocyte primary cilia which depend on both intracellular Ca2+ release and extracellular Ca2+ entry. Using siRNA-mediated knockdowns, we demonstrate that TRPV4, but not PC2 or PIEZO1, mediates flow-induced ciliary Ca2+ increases and loading-induced Cox-2 mRNA increases, an osteogenic response. In this study, we show that the primary cilium forms a Ca2+ microdomain dependent on Ca2+ entry through TRPV4. These results demonstrate that the mechanism of mechanotransduction mediated by primary cilia varies in different tissue contexts. Additionally, we anticipate that this work is a starting point for more studies investigating the role of TRPV4 in mechanotransduction. The online version of this article (doi:10.1186/s13630-015-0016-y) contains supplementary material, which is available to authorized users.