Endo Kazuhira, Takino Takahisa, Miyamori Hisashi, Kinsen Hidenori, Yoshizaki Tomokazu, Furukawa Mitsuru, Sato Hiroshi
Department of Molecular Virology and Oncology, Cancer Research Institute, Graduate School of Medical Science, Kanazawa University, 13-1 Takara-machi, Kanazawa, Ishikawa 920-0934, Japan.
J Biol Chem. 2003 Oct 17;278(42):40764-70. doi: 10.1074/jbc.M306736200. Epub 2003 Aug 6.
The transmembrane heparan sulfate proteoglycan syndecan-1 was identified from a human placenta cDNA library by the expression cloning method as a gene product that interacts with membrane type matrix metalloproteinase-1 (MT1-MMP). Co-expression of MT1-MMP with syndecan-1 in HEK293T cells promoted syndecan-1 shedding, and concentration of cell-associated syndecan-1 was reduced. Treatment of cells with MMP inhibitor BB-94 or tissue inhibitor of MMP (TIMP)-2 but not TIMP-1 interfered with the syndecan-1 shedding promoted by MT1-MMP expression. In contrast, syndecan-1 shedding induced by 12-O-tetradecanoylphorbol-13-acetate treatment was inhibited by BB-94 but not by either TIMP-1 or TIMP-2. Shedding of syndecan-1 was also induced by MT3-MMP but not by other MT-MMPs. Recombinant syndecan-1 core protein was shown to be cleaved by recombinant MT1-MMP or MT3-MMP preferentially at the Gly245-Leu246 peptide bond. HT1080 fibrosarcoma cells stably transfected with the syndecan-1 cDNA (HT1080/SDC), which express endogenous MT1-MMP, spontaneously shed syndecan-1. Migration of HT1080/SDC cells on collagen-coated dishes was significantly slower than that of control HT1080 cells. Treatment of HT1080/SDC cells with BB-94 or TIMP-2 induced accumulation of syndecan-1 on the cell surface, concomitant with further retardation of cell migration. Substitution of Gly245 of syndecan-1 with Leu significantly reduced shedding from HT1080/SDC cells and cell migration. These results suggest that the shedding of syndecan-1 promoted by MT1-MMP through the preferential cleavage of Gly245-Leu246 peptide bond stimulates cell migration.
通过表达克隆方法，从人胎盘cDNA文库中鉴定出跨膜硫酸乙酰肝素蛋白聚糖syndecan-1，它是一种与膜型基质金属蛋白酶-1（MT1-MMP）相互作用的基因产物。MT1-MMP与syndecan-1在HEK293T细胞中共表达促进了syndecan-1的脱落，细胞相关的syndecan-1浓度降低。用MMP抑制剂BB-94或MMP组织抑制剂（TIMP）-2处理细胞，但TIMP-1不影响MT1-MMP表达促进的syndecan-1脱落。相反，12-O-十四烷酰佛波醇-13-乙酸酯处理诱导的syndecan-1脱落被BB-94抑制，但不被TIMP-1或TIMP-2抑制。MT3-MMP也可诱导syndecan-1的脱落，但其他MT-MMP则不能。重组syndecan-1核心蛋白被证明可被重组MT1-MMP或MT3-MMP优先在Gly245-Leu246肽键处切割。稳定转染syndecan-1 cDNA的HT1080纤维肉瘤细胞（HT1080/SDC）表达内源性MT1-MMP，可自发脱落syndecan-1。HT1080/SDC细胞在胶原包被培养皿上的迁移明显慢于对照HT1080细胞。用BB-94或TIMP-2处理HT1080/SDC细胞可诱导syndecan-1在细胞表面积累，同时细胞迁移进一步减慢。将syndecan-1的Gly245替换为Leu可显著减少HT1080/SDC细胞的脱落和细胞迁移。这些结果表明，MT1-MMP通过优先切割Gly245-Leu246肽键促进syndecan-1的脱落，从而刺激细胞迁移。
