Binding of activated α2-macroglobulin to its cell surface receptor GRP78 in 1-LN prostate cancer cells regulates PAK-2-dependent activation of LIMK
Binding of activated α2-macroglobulin to its cell surface receptor GRP78 in 1-LN prostate cancer cells regulates PAK-2-dependent activation of LIMK
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DOI：
10.1074/jbc.m414467200
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发表时间：
2005-07-15
影响因子：
4.8
通讯作者：
Pizzo, SV
中科院分区：
文献类型：
--
作者：
Misra, UK;Deedwania, R;Pizzo, SV
高度恶性细胞的两个特征是它们增加的运动性和蛋白酶分泌，允许这些细胞穿透周围的基底膜并转移。21-kDa活化激酶（PAK）的活化是增加细胞运动性的重要机制。最近，我们报道了受体识别形式的蛋白酶抑制剂α 2-巨球蛋白（α M-2*）与1-LN人前列腺癌细胞表面的GRP 78的结合诱导促有丝分裂信号传导和细胞增殖。在本研究中，我们检测了α 2 M * 激活PAK-1和PAK-2的能力。1-LN细胞暴露于α M-2* 导致磷酸化PAK-2增加2至3倍，其对髓鞘碱性蛋白的激酶活性也有类似的增加。相比之下，PAK-1的磷酸化仅受到可忽略的影响。沉默GRP 78基因的表达，使用两种不同的mRNA序列中的任何一种，大大减弱了α M-2* 刺激细胞中磷酸化PAK-2的出现。用α M-2* 处理1-LN细胞导致PAK-2与NCK结合易位到细胞表面，这通过来自质膜的GRP 78免疫沉淀物中PAK-2和NCK的共免疫沉淀来证明。α M-2* 诱导的PAK-2活化可通过预先将细胞与酪氨酸激酶和磷脂酰肌醇3-激酶的特异性抑制剂孵育来抑制。PAK-2激活伴随着磷酸化LIMK和磷酸化cofilin水平的显著增加。沉默PAK-2基因的表达大大减弱了LIMK的磷酸化。总之，我们首次发现蛋白酶抑制剂α（2）-巨球蛋白可激活1-LN前列腺癌细胞中的PAK-2。这些研究表明了α M-2* 增强这些细胞转移潜力的机制。
Two characteristics of highly malignant cells are their increased motility and secretion of proteinases allowing these cells to penetrate surrounding basement membranes and metastasize. Activation of 21-kDa activated kinases (PAKs) is an important mechanism for increasing cell motility. Recently, we reported that binding of receptor-recognized forms of the proteinase inhibitor alpha(2)-macroglobulin (alpha M-2*) to GRP78 on the cell surface of 1-LN human prostate cancer cells induces mitogenic signaling and cellular proliferation. In the current study, we have examined the ability of alpha 2M* to activate PAK-1 and PAK-2. Exposure of 1-LN cells to alpha M-2* caused a 2- to 3-fold increase in phosphorylated PAK-2 and a similar increase in its kinase activity toward myelin basic protein. By contrast, the phosphorylation of PAK-1 was only negligibly affected. Silencing the expression of the GRP78 gene, using either of two different mRNA sequences, greatly attenuated the appearance of phosphorylated PAK-2 in alpha M-2*-stimulated cells. Treatment of 1-LN cells with alpha M-2* caused translocation of PAK-2 in association with NCK to the cell surface as evidenced by the coimmunoprecipitation of PAK-2 and NCK in the GRP78 immunoprecipitate from plasma membranes. alpha M-2*-induced activation of PAK-2 was inhibited by prior incubation of the cells with specific inhibitors of tyrosine kinases and phosphatidylinositol 3-kinase. PAK-2 activation was accompanied by significant increases in the levels of phosphorylated LIMK and phosphorylated cofilin. Silencing the expression of the PAK-2 gene greatly attenuated the phosphorylation of LIMK. In conclusion, we show for the first time the activation of PAK-2 in 1-LN prostate cancer cells by a proteinase inhibitor, alpha(2)-macroglobulin. These studies suggest a mechanism by which alpha M-2* enhances the metastatic potential of these cells.