Signaling Through Membrane Type 1 Matrix Metalloproteinase

描述（由申请人提供）：膜1型基质金属蛋白酶（MT 1-MMP）是一种跨膜蛋白酶，具有细胞外蛋白水解结构域和短胞质尾区。MT 1-MMP的蛋白水解活性在生理上被基质金属蛋白酶组织抑制剂-2（TIMP-2）抑制。TIMP-2与MT 1-MMP结合，复合物通过蛋白酶胞质尾区介导的内吞作用内化。我们广泛的前期工作表明，TIMP-2与MT 1-MMP的结合可迅速诱导ERK 1/2 MAP激酶活化，从而上调细胞增殖和迁移。这些作用需要胞质尾区，但不需要MT 1-MMP的蛋白水解活性。 因此，我们建议： 1.确定TIMP-2与MT 1-MMP结合产生的ERK 1/2活化的生理意义。为此，将使用野生型和MT 1-MMP敲除小鼠胚胎成纤维细胞来确定该信号传导机制及其产生的生物学效应是否在表达生理量的MT 1-MMP的基质细胞中具有活性，并且依赖于细胞质尾区而不是蛋白酶的蛋白水解活性。 2.探讨MT 1-MMP介导细胞内信号转导的分子机制。我们建议，以确定蛋白质的相互作用与细胞质尾的MT 1-MMP TIMP-2添加后，并表征的作用，内吞作用的TIMP-2-MT 1-MMP复合物的控制MT 1-MMP介导的细胞内信号。 拟议研究项目的结果将阐明细胞表面蛋白之间的相互作用产生细胞内信号的新机制，这些信号在许多细胞功能的调节中具有重要作用。因此，对这些机制的详细了解可以对我们理解各种生理和病理过程以及开发旨在控制它们的药理学工具产生相关影响。

DESCRIPTION (provided by applicant): Membrane-type 1 matrix metalloproteinase (MT1-MMP) is a transmembrane proteinase with an extracellular proteolytic domain and a short cytoplasmic tail. The proteolytic activity of MT1-MMP is inhibited physiologically by tissue inhibitor of matrix metalloproteinase-2 (TIMP-2). TIMP-2 binds to MT1-MMP and the complex is internalized by endocytosis mediated by the cytoplasmic tail of the proteinase. Our extensive preliminary work has shown that binding of TIMP-2 to MT1-MMP rapidly induces ERK1/2 MAP kinase activation, which upregulates cell proliferation and migration. These effects require the cytoplasmic tail but not the proteolytic activity of MT1 -MMP. We therefore propose to: 1. Determine the physiologic significance of ERK1/2 activation generated by TIMP-2 binding to MT1-MMP. For this purpose, wild-type and MT1-MMP knockout mouse embryonic fibroblasts will be used to determine if this signaling mechanism and its resulting biological effects are active in stromal cells expressing physiologic amounts of MT1 -MMP and dependent on the cytoplasmic tail but not the proteolytic activity of the proteinase. 2. Investigate the molecular mechanism of intracellular signaling by MT1-MMP. We propose to identify proteins that interact with the cytoplasmic tail of MT1-MMP upon TIMP-2 addition, and to characterize the role of endocytosis of the TIMP-2-MT1-MMP complex in the control of MT1-MMP-mediated intracellular signaling. The results of the proposed research project will elucidate novel mechanisms through which interactions among cell surface proteins generate intracellular signaling that has important roles in the regulation of numerous cell functions. A detailed understanding of these mechanisms can therefore have relevant implications for our comprehension of a variety of physiological and pathological processes, and for the development of pharmacological tools aimed to control them.