Identification of CD63 as a tissue inhibitor of metalloproteinase-1 interacting cell surface protein
Identification of CD63 as a tissue inhibitor of metalloproteinase-1 interacting cell surface protein
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DOI：
10.1038/sj.emboj.7601281
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发表时间：
2006-09-06
期刊：
影响因子：
11.4
通讯作者：
Kim, Hyeong-Reh Choi
中科院分区：
文献类型：
--
作者：
Jung, Ki-Kyung;Liu, Xu-Wen;Kim, Hyeong-Reh Choi
本研究通过酵母双杂交筛选，确定了四白蛋白家族成员CD63是TIMP-1相互作用蛋白。免疫沉淀和共聚焦显微镜分析证实了CD63与TIMP-1、整合素β 1的相互作用，以及它们在人乳腺上皮MCF10A细胞表面的共定位。TIMP-1的表达与细胞表面活性整合素β 1的水平相关，与细胞粘附无关。当MCF10A细胞在三维（3D）基质中形成极化的腺泡样结构时，TIMP-1过表达破坏了乳腺上皮细胞的极化，抑制了位于中心位置的caspase介导的细胞凋亡，这是中空腺泡样结构形成和维持所必需的。小发夹RNA （Small hairpin RNA, shRNA）介导的CD63下调有效降低了TIMP-1与细胞表面的结合、TIMP-1与整合素β 1的共定位，从而逆转了TIMP-1介导的整合素β 1活化、细胞存活信号传导和细胞凋亡抑制。CD63下调也恢复了3d基质中TIMP-1过表达MCF10A细胞的极化和凋亡。综上所述，本研究确定CD63是TIMP-1的细胞表面结合伙伴，通过TIMP-1调节四跨蛋白/整合素信号复合物来调节细胞存活和极化。
This study identified CD63, a member of the tetraspanin family, as a TIMP-1 interacting protein by yeast two-hybrid screening. Immunoprecipitation and confocal microscopic analysis confirmed CD63 interactions with TIMP-1, integrin beta 1, and their co-localizations on the cell surface of human breast epithelial MCF10A cells. TIMP-1 expression correlated with the level of active integrin beta 1 on the cell surface independent of cell adhesion. While MCF10A cells within a three-dimensional (3D) matrigel matrix form polarized acinar-like structures, TIMP-1 over-expression disrupted breast epithelial cell polarization and inhibited caspase-mediated apoptosis in centrally located cells, necessary for the formation and maintenance of the hollow acinar-like structures. Small hairpin RNA (shRNA)-mediated CD63 downregulation effectively reduced TIMP-1 binding to the cell surface, TIMP-1 co-localization with integrin beta 1, and consequently reversed TIMP-1-mediated integrin beta 1 activation, cell survival signaling and apoptosis inhibition. CD63 downregulation also restored polarization and apoptosis of TIMP-1 over-expressing MCF10A cells within a 3D-matrigel matrix. Taken together, the present study identified CD63 as a cell surface binding partner for TIMP-1, regulating cell survival and polarization via TIMP-1 modulation of tetraspanin/integrin signaling complex.