Isolation and Characterization of Laminin-10/11 Secreted by Human Lung Carcinoma Cells
LAMININ-10/11 MEDIATES CELL ADHESION THROUGH INTEGRIN α3β1*
- From the Research Institute, Osaka Medical Center for Maternal and Child Health, 840 Murodo, Izumi, Osaka 594-1101, Japan
Abstract
A panel of human tumor cell lines was screened for selective expression of laminin α5 chain, a newly identified laminin subunit comprising laminin-10 (α5β1γ1) and -11 (α5β2γ1). The lung adenocarcinoma cell line A549 was found to express the α5 chain at relatively high levels but no detectable amounts of other α chains. The laminin variants containing α5 chain were purified from the conditioned medium of A549 cells by immunoaffinity chromatography using the anti-laminin monoclonal antibody 4C7 which was shown recently to recognize the laminin α5 chain (Tiger, C.-F., Champliaud, M.-F., Pedrosa-Domellof, F., Thornell, L.-E., Ekblom, P., and Gullberg, D. (1997) J. Biol. Chem. 272, 28590–28595). The purified laminin variants consisted of three chains with molecular masses of 350, 220, and 210 kDa. The 350-kDa chain was specifically recognized by another anti-α5 chain monoclonal antibody capable of recognizing denatured α5 chain on immunoblots, whereas the 210-kDa chain was recognized by an anti-γ1 chain antibody. The purified α5 chain-containing laminin variants (hereafter referred to as laminin-10/11) were highly active in mediating adhesion of A549 cells to the substratum with potency as high as that of laminin-5 and significantly higher than those of laminin-1, laminin-2/4, or fibronectin. Adhesion to substrata coated with laminin-10/11 was specifically inhibited by anti-integrin antibodies directed against the integrin α3 or β1 subunit but not by those against α2 or α6 subunit, indicating that laminin-10/11 is specifically recognized by integrin α3β1. Given the wide distribution of laminin-10/11 in the basement membrane of various tissue types and dominant expression of integrin α3β1 in most epithelial cells, specific interaction of laminin-10/11 with integrin α3β1 may play an important role in in vivo regulation of proliferation and differentiation of epithelial cells through the basement membrane.
Footnotes
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↵* This work was supported by special coordination funds from the Science and Technology Agencies of Japan.The costs of publication of this article were defrayed in part by the payment of page charges. The article must therefore be hereby marked “advertisement” in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.
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↵‡ To whom correspondence should be addressed. Tel.: 81-6-879-8617; Fax: 81-6-879-8619; E-mail: sekiguch{at}protein.osaka-u.ac.jp.
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↵1 The abbreviations used are: EHS, Engelbreth-Holm-Swarm; DMEM, Dulbecco’s modified Eagle’s medium; RT-PCR, reverse transcription-polymerase chain reaction; GST, glutathione S-transferase; PAGE, polyacrylamide gel electrophoresis.
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↵2 Y. Kikkawa, unpublished observation.
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- Received January 26, 1998.
- Revision received April 8, 1998.
- The American Society for Biochemistry and Molecular Biology, Inc.
