Shaw A R, Domanska A, Mak A, Gilchrist A, Dobler K, Visser L, Poppema S, Fliegel L, Letarte M, Willett B J
Department of Oncology, University of Alberta, Edmonton, Canada.
J Biol Chem. 1995 Oct 13;270(41):24092-9. doi: 10.1074/jbc.270.41.24092.
Few molecules have been shown to confer cell motility. Although the motility-arresting properties of anti-CD9 monoclonal antibody (mAb) suggest the transmembrane 4 superfamily (TM4SF) member CD9 can induce a motorgenic signal, gene transfection studies have failed to confirm this hypothesis. We report here that ectopic expression of human CD9 (CD9h) and feline CD9 (CD9f) in the CD9-negative, poorly motile, human B cell line Raji dramatically enhances migration across fibronectin- and laminin-coated polycarbonate filters. Migration of Raji/CD9h and Raji/CD9f on either substrate was inhibited by the anti-CD9 mAb 50H.19 and by the anti-beta 1 integrin mAb AP-138. Migration of Raji/CD9h on laminin was potently inhibited by the anti-VLA-6 integrin mAb GoH3 and by the anti-VLA-4 integrin mAb 44H6, whereas migration of Raji/CD9h on fibronectin was inhibited only by mAb 44H6. Since CD9h-transfected Raji cells adhered to fibronectin as effectively as mock transfectants, expression of CD9 enhanced motility, but not adhesion. CD9-enhanced migration was inhibited by the protein tyrosine kinase inhibitor herbimycin A suggesting that tyrosine phosphorylation played a role in the generation of a motorgenic signal. Raji/CD9h transfectants adherent to fibronectin expressed 6-fold higher levels of phosphotyrosine than Raji. Raji/CD9f transfectants also phosphorylated proteins on tyrosine more effectively than Raji including a protein of 110 kDa which was phosphorylated on the motility-inducing substrates laminin and fibronectin, but not on bovine serum albumin. Our results support a role for CD9 in the amplification of a motorgenic signal in B cells involving beta 1 integrins and the activation of protein tyrosine kinases.
很少有分子被证明能赋予细胞运动性。尽管抗CD9单克隆抗体（mAb）的运动抑制特性表明跨膜4超家族（TM4SF）成员CD9可诱导运动信号，但基因转染研究未能证实这一假设。我们在此报告，在CD9阴性、运动能力差的人B细胞系Raji中异位表达人CD9（CD9h）和猫CD9（CD9f），可显著增强其跨纤连蛋白和层粘连蛋白包被的聚碳酸酯滤膜的迁移能力。抗CD9 mAb 50H.19和抗β1整合素mAb AP-138可抑制Raji/CD9h和Raji/CD9f在任一底物上的迁移。抗VLA-6整合素mAb GoH3和抗VLA-4整合素mAb 44H6可有效抑制Raji/CD9h在层粘连蛋白上的迁移，而Raji/CD9h在纤连蛋白上的迁移仅被mAb 44H6抑制。由于CD9h转染的Raji细胞与纤连蛋白的黏附效果与mock转染细胞相同，因此CD9的表达增强了运动能力，但未增强黏附能力。蛋白酪氨酸激酶抑制剂赫伯霉素A可抑制CD9增强的迁移，这表明酪氨酸磷酸化在运动信号的产生中起作用。黏附于纤连蛋白的Raji/CD9h转染细胞中磷酸酪氨酸的表达水平比Raji高6倍。Raji/CD9f转染细胞在酪氨酸上的蛋白磷酸化也比Raji更有效，包括一种110 kDa的蛋白，该蛋白在诱导运动的底物层粘连蛋白和纤连蛋白上发生磷酸化，但在牛血清白蛋白上未发生磷酸化。我们的结果支持CD9在B细胞中涉及β1整合素和蛋白酪氨酸激酶激活的运动信号放大中发挥作用。
