Abstract
The laminin alpha 1 chain carboxyl-terminal globular domain has been identified as a site of multiple biological activities. Using a systematic screening for cell binding sites with 113 overlapping synthetic peptide beads that covered this domain, we found 19 potential active sequences. Corresponding synthetic peptides were evaluated for direct cell attachment, spreading, and inhibition of cell spreading to a laminin-1 substrate using several cell lines. Five peptides (AG-10, AG-22, AG-32, AG-56, and AG-73) showed cell attachment activities with cell-type specificities. Cell spreading on AG-10 was inhibited by beta 1 and alpha 6 integrin antibodies and on AG-32 was inhibited by beta 1, alpha 2, and alpha 6 integrin antibodies. In contrast, cell adhesion and spreading on peptide AG-73 were not inhibited by these antibodies. The minimum active sequences of AG-10, AG-32, and AG-73 were determined to be SIYITRF, IAFQRN, and LQVQLSIR, respectively. These sequences are highly conserved among the different species and different laminin alpha chains, suggesting that they play a critical role for biological function and for interaction with cell surface receptors.
Highlights
Laminin-I is a major component and cell adhesion protein of the basement membrane matrix [1,2,3]
An IKVAV sequence located on the COOH-terminal end of the long arm of the a1 chain was found to promote cell adhesion, neurite outgrowth, experimental metastasis, collagenase IV secretion, angiogenesis, cell growth, and tumor growth [15,16,17,18]
E8, a proteolytic fragment containing the COOH-terminallong arm and the NH2-terminal 60% of the G domain, possesses a major cell binding activity that is mediated through a6/31 integrin [19,20,21]
Summary
Laminin-I is a major component and cell adhesion protein of the basement membrane matrix [1,2,3]. An IKVAV sequence located on the COOH-terminal end of the long arm of the a1 chain was found to promote cell adhesion, neurite outgrowth, experimental metastasis, collagenase IV secretion, angiogenesis, cell growth, and tumor growth [15,16,17,18]. We describe the systematic screening of'biologically active sequences in the mouse laminin al chain G domain (positions 2111-3060) using a large set of overlapping peptides covalently bound to resin beads. Using this assay system as the first stage of screening, we examined the cell attachment activities of 113 different peptide beads. Specific antibodies to integrins blocked adhesion to two of these active peptides
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