Analysis of a molecular mechanism by which JNK-binding proteins regulate turnover of focal adhesions and formAnalysis of a molecular mechanism by which JNK-binding proteins regulate turnover of focal adhesions and formation of cell polarity

膜型基质金属蛋白酶1（MT 1-MMP）与肿瘤的侵袭和转移有关。我们以前报道过MT 1-MMP降解细胞外基质通过调节持续的整合素介导的信号调节细胞迁移。在这项研究中，MT 1-MMP-表达细胞接种到纤连蛋白包被的平板上，并监测细胞基质粘附形成和纤连蛋白降解。纤连蛋白降解并与细胞迁移轨迹一致地去除。迁移的细胞呈极化形态，并通过细胞-基质粘附集中的前缘与纤连蛋白的边缘接触。通过与粘着斑激酶（FAK）的粘着斑靶向（FAT）结构域融合，靶向细胞-基质粘附的MT 1-MMP的表达促进了粘附后立即在细胞周围的初始纤连蛋白溶解。这些结果表明，纤连蛋白被位于细胞-基质粘附处的MT 1-MMP降解，所述细胞-基质粘附集中在迁移细胞的前缘。Ib抑制细胞-基质粘附的MT 1-MMP，MT 1-MMP的显性负性形式（MT 1-Pex）通过与FAT结构域（MT 1-Pex-FAT）融合而靶向细胞-基质粘附。MT 1-Pex-FAT在细胞-基质粘附处积累，并且比亲本MT 1-Pex更有效地抑制纤连蛋白降解以及FAK磷酸化。MT 1-Pex-FAT也显示出比MT 1-Pex更强地抑制肿瘤细胞侵入三维胶原凝胶。这些结果表明，MT 1-MMP-介导的细胞外基质裂解在细胞-基质粘附诱导细胞极性的建立，这有利于细胞-基质粘附周转和随后的细胞迁移。该模型强调了MT 1-MMP在迁移细胞前沿的作用。

Membrane-type 1 matrix metalloproteinase (MT1-MMP) has been implicated in tumor invasion and metastasis. We previously reported that extracellular matrix degradation by MT1-MMP regulates cell migration via modulating sustained integrin-mediated signals. In this study, MT1-MMP-expressing cells were plated onto fibronectin-coated plates and monitored for cell-matrix adhesion formation and fibronectin degradation. The fibronectin was degraded and removed in line with the cell migration track. The migrating cells showed a polarized morphology and were in contact with the edge of fibronectin through the leading edge in which cell-matrix adhesions are concentrated. Expression of MT1-MMP targeted to cell-matrix adhesions by fusing with the focal adhesion targeting (FAT) domain of focal adhesion kinase (FAK) promoted the initial fibronectin lysis at the cell periphery immediately after adhesion. These results suggest that fibronectin is degraded by MT1-MMP located at cell-matrix adhesions which are concentrated at the leading edge of the migrating cells. lb inhibit MT1-MMP at cell-matrix adhesion, the dominant negative form of MT1-MMP (MT1-Pex) was targeted to the cell-matrix adhesion by fusing with the FAT domain (MT1-Pex-FAT). MT1-Pex-FAT accumulated at cell-matrix adhesions and inhibited fibronectin degradation as well as FAK phosphorylation more effectively than parental MT1-Pex. MT1-Pex-FAT was also shown to suppress the invasion of tumor cells into 3-dimensional collagen gel more strongly than MT1-Pex. These results suggest that MT1-MMP-mediated extracellular matrix lysis at cell-matrix adhesions induces the establishment of cell polarity, which facilitates cell-matrix adhesion turnover and subsequent cell migration. This model highlights the role of MT1-MMP at the leading edge of migrating cells.