The
laminin alpha3 chain is mainly expressed at the skin, and its C-terminal G domain has a critical role in multiple
biological functions. We screened for biologically active sites on the mouse
laminin alpha3 chain G domain using 107 synthetic
peptides on coated plates and conjugated to
Sepharose beads with HT1080 human
fibrosarcoma cells, HaCaT human skin keratinocyte cells, and human dermal fibroblasts (HDFs). Eleven
peptides exhibited cell attachment activity with respect to the
peptide-coated plates and/or
peptide-
Sepharose beads. MA3G28 (WTIQTTVDRGLL) strongly binds to HaCaT cells. Four
peptides promoted PC12 cell neurite outgrowth.
Heparin inhibited attachment of HDFs to eight
peptides on the coated plates. In contrast,
EDTA significantly inhibited attachment of HDFs to MA3G27 (NAPFPKLSWTIQ) and MA3G28 but had no effect on the attachment of the other
peptides. HDF cells formed well-organized actin stress fibers and focal contacts with
vinculin accumulation on MA3G27. Additionally, attachment of HDFs to MA3G27 was inhibited by anti-alpha6 and anti-beta1
integrin antibodies, suggesting that MA3G27 promotes
alpha6beta1 integrin-mediated cell adhesion. MA3G57 (NQRLASFSNAQQS) exhibited cell attachment activity only in the
peptide bead assay. MA3G57 conjugated to a
chitosan membrane promoted HDF attachment and spreading with well-organized actin stress fibers. The anti-beta1
integrin antibody partially inhibited attachment of HDFs to the MA3G57-chitosan membrane, suggesting that the MA3G57 site is involved in
beta1 integrin-mediated cell attachment. These active sites are likely important in the
biological activities of the
laminin alpha3 chain G domain and would be useful for the study of molecular mechanisms of
laminin-receptor interactions.