Kulkarni S, Saido T C, Suzuki K, Fox J E
Joseph J. Jacobs Center for Thrombosis and Vascular Biology, Department of Molecular Cardiology, The Lerner Research Institute, Cleveland Clinic Foundation, Cleveland, Ohio 44195, USA.
J Biol Chem. 1999 Jul 23;274(30):21265-75. doi: 10.1074/jbc.274.30.21265.
Integrin-induced adhesion leads to cytoskeletal reorganizations, cell migration, spreading, proliferation, and differentiation. The details of the signaling events that induce these changes in cell behavior are not well understood but they appear to involve activation of Rho family members which activate signaling molecules such as tyrosine kinases, serine/threonine kinases, and lipid kinases. The result is the formation of focal complexes, focal adhesions, and bundles and networks of actin filaments that allow the cell to spread. The present study shows that mu-calpain is active in adherent cells, that it cleaves proteins known to be present in focal complexes and focal adhesions, and that overexpression of mu-calpain increased the cleavage of these proteins, induced an overspread morphology and induced an increased number of stress fibers and focal adhesions. Inhibition of calpain with membrane permeable inhibitors or by expression of a dominant negative form of mu-calpain resulted in an inability of cells to spread or to form focal adhesions, actin filament networks, or stress fibers. Cells expressing constitutively active Rac1 could still form focal complexes and actin filament networks (but not focal adhesions or stress fibers) in the presence of calpain inhibitors; cells expressing constitutively active RhoA could form focal adhesions and stress fibers. Taken together, these data indicate that calpain plays an important role in regulating the formation of focal adhesions and Rac- and Rho-induced cytoskeletal reorganizations and that it does so by acting at sites upstream of both Rac1 and RhoA.
整合素诱导的黏附导致细胞骨架重组、细胞迁移、铺展、增殖和分化。引发细胞行为这些变化的信号事件细节尚未完全清楚，但似乎涉及Rho家族成员的激活，这些成员进而激活诸如酪氨酸激酶、丝氨酸/苏氨酸激酶和脂质激酶等信号分子。结果是形成粘着斑复合体、粘着斑以及肌动蛋白丝束和网络，使细胞得以铺展。本研究表明，μ-钙蛋白酶在贴壁细胞中具有活性，它能切割已知存在于粘着斑复合体和粘着斑中的蛋白质，且μ-钙蛋白酶的过表达增加了这些蛋白质的切割，诱导了过度铺展的形态，并增加了应力纤维和粘着斑的数量。用膜通透性抑制剂抑制钙蛋白酶或通过表达μ-钙蛋白酶的显性负性形式，导致细胞无法铺展或形成粘着斑、肌动蛋白丝网络或应力纤维。在存在钙蛋白酶抑制剂的情况下，组成型激活的Rac1表达细胞仍能形成粘着斑复合体和肌动蛋白丝网络（但不能形成粘着斑或应力纤维）；组成型激活的RhoA表达细胞能形成粘着斑和应力纤维。综上所述，这些数据表明钙蛋白酶在调节粘着斑的形成以及Rac和Rho诱导的细胞骨架重组中起重要作用，并且它是通过作用于Rac1和RhoA上游位点来实现的。
