Saoncella S, Echtermeyer F, Denhez F, Nowlen J K, Mosher D F, Robinson S D, Hynes R O, Goetinck P F
Cutaneous Biology Research Center, Massachusetts General Hospital, Harvard Medical School, Charlestown, MA 02129, USA.
Proc Natl Acad Sci U S A. 1999 Mar 16;96(6):2805-10. doi: 10.1073/pnas.96.6.2805.
The assembly of focal adhesions and actin stress fibers by cells plated on fibronectin depends on adhesion-mediated signals involving both integrins and cell-surface heparan sulfate proteoglycans. These two cell-surface receptors interact with different domains of fibronectin. To attempt to identify the heparan sulfate proteoglycans involved, we used fibronectin-null (FN-/-) mouse fibroblasts to eliminate the contribution of endogenous fibronectin during the analysis. FN-/- fibroblasts plated on the cell-binding domain of fibronectin or on antibodies directed against mouse beta1 integrin chains attach but fail to spread and do not form focal adhesions or actin stress fibers. When such cells are treated with antibodies directed against the ectodomain of mouse syndecan-4, they spread fully and assemble focal adhesions and actin stress fibers indistinguishable from those seen in cells plated on intact fibronectin. These results identify syndecan-4 as a heparan sulfate proteoglycan involved in the assembly process. The antibody-stimulated assembly of focal adhesions and actin stress fibers in cells plated on the cell-binding domain of fibronectin can be blocked with C3 exotransferase, an inhibitor of the small GTP-binding protein Rho. Treatment of cells with lysophosphatidic acid, which activates Rho, results in full spreading and assembly of focal adhesions and actin stress fibers in fibroblasts plated on the cell-binding domain of fibronectin. We conclude that syndecan-4 and integrins can act cooperatively in generating signals for cell spreading and for the assembly of focal adhesions and actin stress fibers. We conclude further that these joint signals are regulated in a Rho-dependent manner.
细胞接种在纤连蛋白上时，粘着斑和肌动蛋白应力纤维的组装依赖于涉及整合素和细胞表面硫酸乙酰肝素蛋白聚糖的粘附介导信号。这两种细胞表面受体与纤连蛋白的不同结构域相互作用。为了试图鉴定所涉及的硫酸乙酰肝素蛋白聚糖，我们使用纤连蛋白缺失（FN-/-）的小鼠成纤维细胞，以在分析过程中消除内源性纤连蛋白的影响。接种在纤连蛋白的细胞结合结构域或针对小鼠β1整合素链的抗体上的FN-/-成纤维细胞能够附着，但无法铺展，也不会形成粘着斑或肌动蛋白应力纤维。当用针对小鼠syndecan-4胞外结构域的抗体处理这些细胞时，它们会完全铺展，并组装出与接种在完整纤连蛋白上的细胞中所见无异的粘着斑和肌动蛋白应力纤维。这些结果确定syndecan-4是参与组装过程的一种硫酸乙酰肝素蛋白聚糖。接种在纤连蛋白细胞结合结构域上的细胞中，抗体刺激的粘着斑和肌动蛋白应力纤维组装可被小GTP结合蛋白Rho的抑制剂C3外切转移酶阻断。用激活Rho的溶血磷脂酸处理细胞，会导致接种在纤连蛋白细胞结合结构域上的成纤维细胞完全铺展，并组装粘着斑和肌动蛋白应力纤维。我们得出结论，syndecan-4和整合素可以协同作用，产生细胞铺展以及粘着斑和肌动蛋白应力纤维组装所需的信号。我们进一步得出结论，这些联合信号以Rho依赖的方式受到调节。
