Kundranda Madappa N, Ray Sanhita, Saria Margaret, Friedman David, Matrisian Lynn M, Lukyanov Pavel, Ochieng Josiah
Department of Biochemistry, Meharry Medical College, 1005 Dr. D.B. Todd Jr. Blvd. Nashville, TN 37208-3599, USA.
Biochim Biophys Acta. 2004 Aug 23;1693(2):111-23. doi: 10.1016/j.bbamcr.2004.06.005.
Fetuin-A is a major constituent of the fetal bovine serum used extensively in cell culture media. We hereby present data demonstrating that breast carcinoma cells can adhere to immobilized fetuin-A in a calcium-dependent fashion. Interestingly, the cells can also divide and attain confluency under these conditions. Using a proteomic approach, we have identified annexin-II and -VI as the putative cell surface receptors for fetuin-A in the presence of Ca2+ ions. Biotinylation of cell surface proteins followed by immunoprecipitation revealed that annexin-VI was expressed on the extracytoplasmic surface of the cell membranes. Finally, to demonstrate that annexin-II and -VI were the adhesive receptors for fetuin-A, siRNA knockdown of expression of the annexins significantly reduced the calcium-mediated adhesion. Interestingly, we demonstrated that the tumor cells could also adhere to immobilized fetuin-A in the presence of magnesium ions, and that this adhesion was most likely mediated by integrins because neutralizing antibodies against beta1 integrins substantially reduced the adhesion. Our studies suggest that the expression of annexin-II and -VI and possibly other members of the family mediate novel adhesion and signaling mechanisms in tumor cells.
胎球蛋白-A是细胞培养基中广泛使用的胎牛血清的主要成分。我们在此展示的数据表明，乳腺癌细胞能够以钙依赖的方式黏附于固定化的胎球蛋白-A。有趣的是，在这些条件下细胞还能分裂并达到汇合状态。通过蛋白质组学方法，我们已鉴定出膜联蛋白-II和-VI是在Ca2+离子存在时胎球蛋白-A的假定细胞表面受体。对细胞表面蛋白进行生物素化标记后进行免疫沉淀显示，膜联蛋白-VI在细胞膜的胞外表面表达。最后，为证明膜联蛋白-II和-VI是胎球蛋白-A的黏附受体，对膜联蛋白表达进行小干扰RNA敲低显著降低了钙介导的黏附。有趣的是，我们证明肿瘤细胞在镁离子存在时也能黏附于固定化的胎球蛋白-A，并且这种黏附很可能由整合素介导，因为针对β1整合素的中和抗体显著降低了黏附。我们的研究表明，膜联蛋白-II和-VI以及该家族的其他成员可能介导肿瘤细胞中的新型黏附和信号传导机制。
