Identification of cell binding sequences in mouse laminin gamma1 chain by systematic peptide screening.
Laminin-1, a major component of basement membranes, consists of three different chains designated alpha1, beta1, and gamma1 and has diverse biological functions. We have identified cell binding sites on the mouse laminin gamma1 chain, using systematic screening of 165 overlapping synthetic peptides covering the entire chain. We identified 12 cell binding sequences using HT-1080 human fibrosarcoma and B16-F10 mouse melanoma cells in two independent assays employing peptide-conjugated Sepharose beads and peptide-coated dishes. Four peptides (C-16, C-28, C-64, and C-68) located on the globular domains of the gamma1 chain were the most active and showed dose-dependent cell attachment. Cell attachment to C-68 was inhibited by EDTA and by anti-alpha2beta1 integrin antibodies. Cell attachment to C-16 and C-64 was partially inhibited by EDTA but was not inhibited by anti-integrin antibodies. EDTA and anti-integrin antibodies did not affect cell attachment to C-28. The four peptides were tested in adhesion and differentiation assays with endothelial, neuronal, and human salivary gland cells. C-16 was the most active for all of the cells, whereas the other three peptides showed cell type specificity in their activities. The active core sequences of C-16, C-28, C-64, and C-68 are YVRL, IRVTLN, TTVKYIFR, and SIKIRGTY, respectively. These sequences are highly conserved among the different species and in the laminin gamma2 chain. These results suggest that the specific sequences on the laminin gamma1 chain are biologically active and interact with distinct cell surface receptors.
Nomizu
M
M
Craniofacial Developmental Biology and Regeneration Branch, NIDR, National Institutes of Health, Bethesda, Maryland 20892, USA.
Kuratomi
Y
Y
Song
S Y
SY
Ponce
M L
ML
Hoffman
M P
MP
Powell
S K
SK
Miyoshi
K
K
Otaka
A
A
Kleinman
H K
HK
Yamada
Y
Y
D000595:Amino Acid Sequence; D000818:Animals; D001665:Binding Sites; D002448:Cell Adhesion / Q000187:drug effects / Q000276:immunology; D002460:Cell Line; D004492:Edetic Acid / Q000494:pharmacology; D006801:Humans; D016023:Integrins / Q000737:chemistry / Q000276:immunology / Q000378:metabolism; D007797:Laminin / Q000737:chemistry / Q000378:metabolism*; D051379:Mice; D008969:Molecular Sequence Data; D011956:Receptors, Cell Surface / Q000737:chemistry / Q000378:metabolism; D014407:Tumor Cells, Cultured
D016023:Integrins; D007797:Laminin; D011956:Receptors, Cell Surface; C089141:laminin gamma 1; D004492:Edetic Acid