Institute for Biomedical Research, Birmingham University Medical School, Vincent Drive, Birmingham B15 2TT, UK.
Biochem Soc Trans. 2009 Dec;37(Pt 6):1214-7. doi: 10.1042/BST0371214.
We have applied search algorithms to expression databases to identify genes whose expression is restricted to the endothelial cell. Such genes frequently play a critical role in endothelial biology and angiogenesis. Two such genes are the roundabout receptor Robo4 and the ECSCR (endothelial-cell-specific chemotaxis regulator). Endothelial cells express both Robo1 and Robo4, which we have knocked down using siRNA (small interfering RNA) and then studied the effect in a variety of in vitro assays. Both Robo4 and Robo1 knockdown inhibited in vitro tube formation on Matrigel. Transfection of Robo4 into endothelial cells increased the number of filopodial extensions from the cell, but failed to do so in Robo1-knockdown cells. Separate immunoprecipitation studies showed that Robo1 and Robo4 heterodimerize. We conclude from this and other work that a heteroduplex of Robo1 and Robo4 signals through WASP (Wiskott-Aldrich syndrome protein) and other actin nucleation-promoting factors to increase the number of filopodia and cell migration. Knockdown of the transmembrane ECSCR protein in endothelial cells also reduced chemotaxis and impaired tube formation on Matrigel. Yeast two-hybrid analysis and immunoprecipitation studies showed that, in contrast with the roundabouts, ECSCR binds to the actin-modulatory filamin A. We conclude that all three of these genes are critical for effective endothelial cell migration and, in turn, angiogenesis.
我们已经将搜索算法应用于表达数据库，以鉴定其表达仅限于内皮细胞的基因。此类基因通常在血管生成内皮生物学中起着至关重要的作用。两个这样的基因是圆体受体 Robo4 和 ECSCR（内皮细胞特异性趋化调节因子）。内皮细胞表达 Robo1 和 Robo4，我们使用 siRNA（小干扰 RNA）敲低了这两种基因，然后在各种体外测定中研究了其作用。Robo4 和 Robo1 敲低均抑制了 Matrigel 上的体外管状形成。Robo4 转染到内皮细胞中增加了细胞丝状伪足的数量，但在 Robo1 敲低的细胞中却没有这样做。单独的免疫沉淀研究表明，Robo1 和 Robo4 异二聚化。根据这一结果和其他工作，我们得出结论，Robo1 和 Robo4 的异二聚体通过 WASP（Wiskott-Aldrich 综合征蛋白）和其他肌动蛋白成核促进因子发出信号，以增加丝状伪足的数量并促进细胞迁移。内皮细胞中跨膜 ECSCR 蛋白的敲低也减少了趋化性，并损害了 Matrigel 上的管状形成。酵母双杂交分析和免疫沉淀研究表明，与圆体相反，ECSCR 与肌动蛋白调节蛋白细丝蛋白 A 结合。我们得出结论，这三个基因对于有效的内皮细胞迁移，进而对于血管生成至关重要。
