Ectopic Expression of Human and Feline CD9 in a Human B Cell Line Confers 
1 Integrin-dependent Motility on Fibronectin and Laminin Substrates and Enhanced Tyrosine Phosphorylation (*)
- Andrew R. E. Shaw(§),
- Agatha Domanska,
- Allan Mak,
- Anita Gilchrist,
- Kelly Dobler,
- Lydia Visser,
- Sibrand Poppema(1),
- Larry Fliegel(2),
- Michelle Letarte(3) and
- Brian J. Willett(4)(¶)
- From the (1)Department of Oncology, University of Alberta, and Cross Cancer Institute, Edmonton, Alberta T6G 1Z2, Canada, the Department of Pathology and the
- (2) Department of Pediatrics, University of Alberta, Edmonton, Alberta T6G 2E1, the
- (3) Division of Immunology and Cancer Research, the Hospital for Sick Children, Toronto, Ontario M5G 1X8, Canada, and the
- (4) Department of Veterinary Pathology, University of Glasgow, Glasgow G12 8Q9, United Kingdom
- § To whom correspondence should be addressed: Dept. of Oncology, Cross Cancer Institute, 11560, University Avenue, Edmonton, Alberta T6G 1Z2, Canada.
Abstract
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-β1 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 β1 integrins and the activation of protein tyrosine kinases.
Footnotes
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↵¶ Supported by the Wellcome Trust.
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↵* This work was supported in part by an operating award from the Medical Research Council of Canada (to A. R. E. S.). The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore by hereby marked “advertisement” in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.
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↵1 The abbreviations used are:
- TM4SF
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transmembrane 4 superfamily
- VLA
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very late antigen or β1 integrins
- RHAMM
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receptor for hyaluronic acid-mediated motility
- mAb
-
monoclonal antibody
- PAGE
-
polyacrylamide gel electrophoresis
- BSA
-
bovine serum albumin
- FACS
-
fluorescence-activated cell sorter.
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- Received April 21, 1995.
- Revision received June 14, 1995.
- © 1995 by The American Society for Biochemistry and Molecular Biology, Inc.
