An IKLLI-containing peptide derived from the laminin alpha1 chain mediating heparin-binding, cell adhesion, neurite outgrowth and proliferation, represents a binding site for integrin alpha3beta1 and heparan sulphate proteoglycan.
We synthesized and characterized several peptides containing the IKLLI sequence in the alpha1 chain of laminin-1. The IKLLI-containing peptides, such as LA4 (CSRNLSEIKLLISRARK), LA5 (EIKLLIS) and LA5L (SEIKLLIS), were found to mediate heparin binding and cell adhesion, while also promoting neurite outgrowth in PC12 cells. Furthermore, peptides LA4 and LA5 also mediated proliferation. However, a scrambled peptide, LA5S (ILEKSLI), did not show any of these activities. Anti-LA4 antibodies inhibited laminin- and LA5-mediated cell adhesion and neurite outgrowth, and anti-(integrin alpha3) and anti-(integrin beta1) antibodies inhibited LA5-mediated cell adhesion and neurite outgrowth. Heparin and heparan sulphate inhibited LA5-mediated heparin binding and PC12 cell adhesion in a dose- dependent manner. The IC50 for inhibition of heparin binding and cell adhesion was observed with 9 microM and 8 microM heparin/heparan sulphate respectively. Furthermore, heparan sulphate proteoglycan also inhibited LA5-mediated PC12 cell adhesion with an IC50 of 100 micrograms/ml. However, chondroitin sulphate (dermatan sulphate) did not inhibit cell adhesion. These data suggest that an IKLLI-containing peptide derived from the laminin alpha1 chain may be an active site of laminin and that its cell adhesion may thus interact with both integrin alpha3beta1 and cell- surface heparan sulphate proteoglycan.
Tashiro
K
K
Department of Psychiatry, Miyazaki Medical College, 5200, Kihara, Kiyotake-cho, Miyazaki-gun, Miyazaki, 889-1692, Japan. ktashiro@hf.rim.or.jp
Monji
A
A
Yoshida
I
I
Hayashi
Y
Y
Matsuda
K
K
Tashiro
N
N
Mitsuyama
Y
Y
D000595:Amino Acid Sequence; D000818:Animals; D000906:Antibodies / Q000494:pharmacology; D001665:Binding Sites; D002448:Cell Adhesion / Q000187:drug effects; D002455:Cell Division / Q000187:drug effects; D002460:Cell Line; D048429:Cell Size / Q000187:drug effects; D004247:DNA / Q000096:biosynthesis; D006025:Glycosaminoglycans / Q000378:metabolism / Q000494:pharmacology; D006493:Heparin / Q000031:analogs & derivatives* / Q000378:metabolism* / Q000494:pharmacology; D006801:Humans; D020128:Inhibitory Concentration 50; D039201:Integrin alpha3beta1; D016023:Integrins / Q000276:immunology / Q000378:metabolism*; D007797:Laminin / Q000737:chemistry / Q000276:immunology / Q000378:metabolism* / Q000494:pharmacology; D008969:Molecular Sequence Data; D016501:Neurites / Q000502:physiology*; D016716:PC12 Cells; D010446:Peptide Fragments / Q000737:chemistry / Q000276:immunology / Q000378:metabolism* / Q000494:pharmacology; D011485:Protein Binding / Q000187:drug effects; D011509:Proteoglycans / Q000378:metabolism* / Q000494:pharmacology; D051381:Rats; D012965:Sodium Chloride / Q000494:pharmacology
D000906:Antibodies; D006025:Glycosaminoglycans; D039201:Integrin alpha3beta1; D016023:Integrins; D007797:Laminin; D010446:Peptide Fragments; D011509:Proteoglycans; C036090:heparin proteoglycan; C084098:laminin A; D012965:Sodium Chloride; D006493:Heparin; D004247:DNA