Cell binding sequences in mouse laminin alpha1 chain.
Laminin-1, a multifunctional glycoprotein of the basement membrane, consists of three different subunits, alpha1, beta1, and gamma1 chains. Previously, we used synthetic peptides to screen for biologically active sequences in the laminin alpha1 chain C-terminal globular domain (G domain) and identified several cell binding sequences (Nomizu, M., Kim, W. H., Yamamura, K., Utani, A., Song, S. Y., Otaka, A., Roller, P. P., Kleinman, H. K., and Yamada, Y. (1995) J. Biol. Chem. 270, 20583-20590). Here, we identify new cell binding sequences on the remainder of the laminin alpha1 chain by systematic peptide screening, using 208 overlapping synthetic peptides encompassing the central and N-terminal portions of the alpha1 chain. HT-1080 cell attachment activity to the peptides was evaluated using peptide-coated plastic substrates and peptide-conjugated Sepharose beads. Twenty five peptides showed cell attachment activities on either the peptide-coated plastic substrates and/or the peptide-conjugated Sepharose beads. A-13 (RQVFQVAYIIIKA) showed strongest cell attachment activity in both the assays. Cell attachment to 14 of the peptides was inhibited by heparin. EDTA and integrin antibodies inhibited cell adhesion to two of the peptides, A-13 and A-25, suggesting that these sites likely bind to integrins. These peptides inhibited cell attachment to laminin-1 but not to collagen I, suggesting these active sites are available on the intact molecule. Most of active sequences were localized on globular domains suggesting that these structures play a critical role in binding to cell-surface receptors.
Nomizu
M
M
Craniofacial Developmental Biology and Regeneration Branch, NIDR, National Institutes of Health, Bethesda, Maryland 20892, USA.
Kuratomi
Y
Y
Malinda
K M
KM
Song
S Y
SY
Miyoshi
K
K
Otaka
A
A
Powell
S K
SK
Hoffman
M P
MP
Kleinman
H K
HK
Yamada
Y
Y
D000595:Amino Acid Sequence; D000818:Animals; D000906:Antibodies / Q000276:immunology; D002448:Cell Adhesion; D004492:Edetic Acid / Q000276:immunology; D006801:Humans; D016023:Integrins / Q000276:immunology; D007797:Laminin / Q000737:chemistry / Q000378:metabolism*; D051379:Mice; D008969:Molecular Sequence Data; D010446:Peptide Fragments / Q000378:metabolism; D011485:Protein Binding; D014407:Tumor Cells, Cultured
D000906:Antibodies; D016023:Integrins; D007797:Laminin; D010446:Peptide Fragments; D004492:Edetic Acid