Institute for Medical Microbiology, Virology and Hygiene, University Medical Center Eppendorf, Martinistr. 52, 20246 Hamburg, Germany.
J Cell Sci. 2013 Aug 15;126(Pt 16):3756-69. doi: 10.1242/jcs.129437. Epub 2013 Jun 7.
Regulation of cell-cell contacts is essential for integrity of the vascular endothelium. Here, a critical role of the F-actin-binding protein drebrin in maintaining endothelial integrity is revealed under conditions mimicking vascular flow. Drebrin knockdown leads to weakening of cell-cell contacts, characterized by loss of nectin from adherens junctions and its subsequent lysosomal degradation. Immunoprecipitation, FRAP and mitochondrial re-targeting experiments show that nectin stabilization occurs through a chain of interactions: drebrin binding to F-actin, interaction of drebrin and afadin through their polyproline and PR1-2 regions, and recruitment of nectin through the PDZ region of afadin. Key elements are modules in drebrin that confer binding to afadin and F-actin. Evidence for this was obtained using constructs containing the PDZ region of afadin coupled to the F-actin-binding region of drebrin or to lifeact, which restore junctional nectin under knockdown of drebrin or of both drebrin and afadin. Drebrin, containing binding sites for both afadin and F-actin, is thus uniquely equipped to stabilize nectin at endothelial junctions and to preserve endothelial integrity under vascular flow.
细胞间连接的调控对于血管内皮的完整性至关重要。在这里，揭示了在模拟血管流动条件下，F-肌动蛋白结合蛋白 drebrin 在维持内皮完整性方面的关键作用。drebrin 敲低导致细胞间连接减弱，特征是黏附连接处的 nectin 丢失及其随后的溶酶体降解。免疫沉淀、FRAP 和线粒体重靶向实验表明，nectin 的稳定是通过一系列相互作用实现的：drebrin 与 F-肌动蛋白结合，drebrin 和 afadin 通过其多脯氨酸和 PR1-2 区域相互作用，以及 afadin 的 PDZ 区域招募 nectin。关键元件是 drebrin 中与 afadin 和 F-肌动蛋白结合的模块。使用包含 afadin 的 PDZ 区域与 drebrin 或 lifeact 的 F-肌动蛋白结合区域偶联的构建体获得了这方面的证据，这些构建体能在 drebrin 或 drebrin 和 afadin 敲低的情况下恢复连接 nectin。含有 afadin 和 F-肌动蛋白结合位点的 drebrin 因此具有独特的能力，可以稳定内皮连接处的 nectin，并在血管流动下维持内皮完整性。
