细胞间接触形成调控血管内皮细胞中 TRPC4 的 Ca2+信号:TRPC4-β-连环蛋白相互作用的调控证据。

Cell-cell contact formation governs Ca2+ signaling by TRPC4 in the vascular endothelium: evidence for a regulatory TRPC4-beta-catenin interaction.

机构信息

From the Institute of Pharmaceutical Sciences-Pharmacology and Toxicology, University of Graz, Universitätsplatz 2, 8010 Graz, Austria.

the Institute of Anatomy and Cell Biology, University of Würzburg, Koellikerstrasse 6, 97070 Würzburg, Germany.

出版信息

J Biol Chem. 2010 Feb 5;285(6):4213-4223. doi: 10.1074/jbc.M109.060301. Epub 2009 Dec 8.

Abstract

TRPC4 is well recognized as a prominent cation channel in the vascular endothelium, but its contribution to agonist-induced endothelial Ca(2+) entry is still a matter of controversy. Here we report that the cellular targeting and Ca(2+) signaling function of TRPC4 is determined by the state of cell-cell adhesions during endothelial phenotype transitions. TRPC4 surface expression in human microvascular endothelial cells (HMEC-1) increased with the formation of cell-cell contacts. Epidermal growth factor recruited TRPC4 into the plasma membrane of proliferating cells but initiated retrieval of TRPC4 from the plasma membrane in quiescent, barrier-forming cells. Epidermal growth factor-induced Ca(2+) entry was strongly promoted by the formation of cell-cell contacts, and both siRNA and dominant negative knockdown experiments revealed that TRPC4 mediates stimulated Ca(2+) entry exclusively in proliferating clusters that form immature cell-cell contacts. TRPC4 co-precipitated with the junctional proteins beta-catenin and VE-cadherin. Analysis of cellular localization of fluorescent fusion proteins provided further evidence for recruitment of TRPC4 into junctional complexes. Analysis of TRPC4 function in the HEK293 expression system identified beta-catenin as a signaling molecule that enables cell-cell contact-dependent promotion of TRPC4 function. Our results place TRPC4 as a Ca(2+) entry channel that is regulated by cell-cell contact formation and interaction with beta-catenin. TRPC4 is suggested to serve stimulated Ca(2+) entry in a specific endothelial state during the transition from a proliferating to a quiescent phenotype. Thus, TRPC4 may adopt divergent, as yet unappreciated functions in endothelial Ca(2+) homeostasis and emerges as a potential key player in endothelial phenotype switching and tuning of cellular growth factor signaling.

摘要

TRPC4 是血管内皮中一种重要的阳离子通道,但其在激动剂诱导的内皮细胞 Ca(2+)内流中的作用仍存在争议。本文报道了在血管内皮细胞表型转化过程中,细胞间黏附状态决定了 TRPC4 的细胞靶向和 Ca(2+)信号转导功能。在人微血管内皮细胞(HMEC-1)中,随着细胞间接触的形成,TRPC4 的表面表达增加。表皮生长因子将 TRPC4 募集到增殖细胞的质膜,但在静止的、形成屏障的细胞中,TRPC4 从质膜中被内吞。表皮生长因子诱导的 Ca(2+)内流强烈促进细胞间接触的形成,siRNA 和显性负性敲低实验表明,TRPC4 仅在形成不成熟细胞间接触的增殖簇中介导刺激的 Ca(2+)内流。TRPC4 与连接蛋白β-连环蛋白和 VE-钙黏蛋白共沉淀。荧光融合蛋白的细胞定位分析进一步证明了 TRPC4 被募集到连接复合物中。在 HEK293 表达系统中分析 TRPC4 的功能,鉴定出β-连环蛋白是一种信号分子,它使细胞间接触依赖性促进 TRPC4 功能。我们的研究结果将 TRPC4 定位为一种 Ca(2+)内流通道,其受到细胞间接触形成和与β-连环蛋白相互作用的调节。TRPC4 可能在从增殖到静止表型的转变过程中,在特定的内皮状态下发挥刺激的 Ca(2+)内流作用。因此,TRPC4 在血管内皮细胞 Ca(2+)稳态中可能具有不同的、尚未被认识的功能,并成为内皮细胞表型转换和细胞生长因子信号转导调节的潜在关键因子。

https://cdn.ncbi.nlm.nih.gov/pmc/blobs/bee2/2823560/a517395569da/zbc0091005150001.jpg

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