Calpain-mediated Proteolysis of Paxillin Negatively Regulates Focal Adhesion Dynamics and Cell Migration
Calpain-mediated Proteolysis of Paxillin Negatively Regulates Focal Adhesion Dynamics and Cell Migration
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DOI：
10.1074/jbc.m110.187294
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发表时间：
2011-03-25
影响因子：
4.8
通讯作者：
Huttenlocher, Anna
中科院分区：
文献类型：
--
作者：
Cortesio, Christa L.;Boateng, Lindsy R.;Huttenlocher, Anna
整合素介导的黏附的动态转换对细胞迁移很重要。桩蛋白是一种定位于黏着斑的衔接蛋白，与细胞运动有关。我们之前报道过，钙蛋白酶介导的踝蛋白1和黏着斑激酶的蛋白水解作用介导了运动细胞中黏附的解体。为了确定钙蛋白酶介导的桩蛋白水解是否调节黏着斑动力学和细胞运动，我们绘制了桩蛋白中钙蛋白酶偏好的蛋白水解位点。切割位点位于桩蛋白的LD1和LD2基序之间，并产生一个大小与替代产物桩蛋白δ相似的C末端片段。钙蛋白酶产生的蛋白水解片段与桩蛋白δ一样，作为桩蛋白的拮抗剂发挥作用，并损害黏着斑解体和迁移。我们构建了带有一个点突变（S95G）的突变型桩蛋白，使其对钙蛋白酶水解具有部分抗性。通过延时显微镜观察，表达桩蛋白S95G的桩蛋白缺陷细胞显示含斑联蛋白的黏附的转换增加，并且迁移也增加。此外，桩蛋白中与癌症相关的体细胞突变在LD1和LD2基序之间的N末端区域很常见，并赋予部分钙蛋白酶抗性。综上所述，这些发现表明钙蛋白酶介导的桩蛋白水解作为黏着斑动力学和迁移的负调节因子具有一种新的作用，可能起到限制癌细胞侵袭的作用。
The dynamic turnover of integrin-mediated adhesions is important for cell migration. Paxillin is an adaptor protein that localizes to focal adhesions and has been implicated in cell motility. We previously reported that calpain-mediated proteolysis of talin1 and focal adhesion kinase mediates adhesion disassembly in motile cells. To determine whether calpain-mediated paxillin proteolysis regulates focal adhesion dynamics and cell motility, we mapped the preferred calpain proteolytic site in paxillin. The cleavage site is between the paxillin LD1 and LD2 motifs and generates a C-terminal fragment that is similar in size to the alternative product paxillin delta. The calpain-generated proteolytic fragment, like paxillin delta, functions as a paxillin antagonist and impairs focal adhesion disassembly and migration. We generated mutant paxillin with a point mutation (S95G) that renders it partially resistant to calpain proteolysis. Paxillin-deficient cells that express paxillin S95G display increased turnover of zyxin-containing adhesions using time-lapse microscopy and also show increased migration. Moreover, cancer-associated somatic mutations in paxillin are common in the N-terminal region between the LD1 and LD2 motifs and confer partial calpain resistance. Taken together, these findings suggest a novel role for calpain-mediated proteolysis of paxillin as a negative regulator of focal adhesion dynamics and migration that may function to limit cancer cell invasion.