Regulation of focal complex composition and disassembly by the calcium-dependent protease calpain.
Regulation of focal complex composition and disassembly by the calcium-dependent protease calpain.
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发表时间：
2002-09
影响因子：
4
通讯作者：
A. Bhatt;I. Kaverina;C. Otey;A. Huttenlocher
中科院分区：
文献类型：
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作者：
A. Bhatt;I. Kaverina;C. Otey;A. Huttenlocher
细胞迁移需要粘合剂复合物的调节和动态周转。我们之前已经证明，钙依赖性蛋白酶钙蛋白酶在细胞迁移过程中调节粘附复合物的组织和细胞分离。现在有证据表明，通过钙蛋白酶内源性抑制剂、钙蛋白酶抑制素和药物抑制剂的过度表达来抑制钙蛋白酶，可抑制粘附复合物的分解，并稳定细胞外周的 GFP-vinculin 和 GFP/RFP-zyxin。诺考达唑冲洗后，微管介导的粘附复合物位点的周转也需要钙蛋白酶，这表明钙蛋白酶可能介导微管下游的局灶性复合物解体。使用 RFP-zyxin 和 GFP-α-辅肌动蛋白的双重成像，我们观察到 α-辅肌动蛋白定位到焦点接触以及随后在细胞回缩区域中含有焦点复合物的 RFP-zyxin 分解或易位之间的时间和空间关系。钙蛋白酶抑制破坏了α-辅肌动蛋白定位到含有zyxin的焦点接触以及焦点复合物解体或易位到细胞中心。此外，通过表达α-辅肌动蛋白杆结构域（而不是头结构域）破坏α-辅肌动蛋白对焦点复合物的定位，导致与钙蛋白酶抑制类似的粘着斑分解的抑制。我们的研究提出了一种新的作用机制，钙蛋白酶可以调节α-辅肌动蛋白定位到焦点复合物及其随后的分解或易位。
Cell migration requires the regulated and dynamic turnover of adhesive complexes. We have previously demonstrated that the calcium-dependent protease, calpain, regulates the organization of adhesive complexes and cell detachment during cell migration. Evidence is now provided that inhibiting calpain through over-expression of the endogenous inhibitor of calpain, calpastatin, and pharmacological inhibitors results in an inhibition of adhesive complex disassembly with stabilization of GFP-vinculin and GFP/RFP-zyxin at the cell periphery. Calpain was also required for the microtubule-mediated turnover of adhesive complex sites after nocodazole wash-out, suggesting that calpain may mediate focal complex disassembly downstream of microtubules. Using dual imaging of RFP-zyxin and GFP-alpha-actinin, we observed a temporal and spatial relationship between alpha-actinin localization to focal contacts and the subsequent disassembly or translocation of RFP-zyxin containing focal complexes in areas of cell retraction. Calpain inhibition disrupted alpha-actinin localization to zyxin-containing focal contacts and focal complex disassembly or translocation to the cell center. In addition, disrupting alpha-actinin localization to focal complexes through expression of the alpha-actinin rod domain, but not the head domain, resulted in inhibition of focal adhesion disassembly similar to calpain inhibition. Our studies suggest a novel mechanism of action whereby calpain may modulate alpha-actinin localization into focal complexes and their subsequent disassembly or translocation.