title: Laminin-1 and the RKRLQVQLSIRT laminin-1 alpha1 globular domain peptide stimulate matrix metalloproteinase secretion by PC12 cells.
abstract: Here we have investigated the ability of laminin-1 and specific laminin-1-derived synthetic peptides to stimulate neuronal cell matrix metalloproteinase secretion. Zymographic analysis of conditioned media from laminin-1-treated PC12 and NG108-15 cells revealed a 72-kDa matrix metalloproteinase which was not secreted by untreated cells. Laminin-1 alpha1 chain-derived synthetic peptides, AASIKVAVSADR (LAM-L) and RKRLQVQLSIRT (AG-73), also stimulated PC12 cell secretion of a 72-kDa matrix metalloproteinase. We further investigated the structural requirements of AG-73 for cell attachment, neurite outgrowth, and matrix metalloproteinase secretion using a series of AG-73 analogs that had single amino acids substituted with alanine. At the substrate levels tested, the AG-73 peptide promoted the adhesion of 67% of the PC12 cells and neurite outgrowth in 71% of the PC12 cells. Substitutions in any one of the amino acids within the central LQVQ sequence resulted in a large reduction in cell attachment whereas substitution in the carboxyl terminal proximal amino acids L, S, and R had little effect on attachment. Alanine substitution of any of the amino terminal proximal LQV amino acids and the carboxyl terminal L, I, and R residues resulted in a 65-91% reduction in neurite outgrowth. These data demonstrate that the sequence requirements for cell attachment and neurite outgrowth were not necessarily coupled but that the sequence requirements for neurite outgrowth and matrix metalloproteinase secretion were identical. We conclude that laminin-1 is able to stimulate neuronal cells to secrete a matrix metalloproteinase. Further, this study identifies the LQVXLXIR laminin-1 alpha1 globular domain peptide to be capable of stimulating both neurite outgrowth and matrix metalloproteinase secretion.
authors.affiliation: Department of Biology, Adelphi University, Garden City, New York 11530, USA. weeks@adlibv.adelphi.edu
authors.forename: B S
authors.initials: BS
authors.lastname: Weeks
authors.forename: M
authors.initials: M
authors.lastname: Nomizu
authors.forename: R S
authors.initials: RS
authors.lastname: Ramchandran
authors.forename: Y
authors.initials: Y
authors.lastname: Yamada
authors.forename: H K
authors.initials: HK
authors.lastname: Kleinman
mesh_terms: D000818:Animals; D005109:Extracellular Matrix / Q000201:enzymology; D018093:Gelatinases / Q000378:metabolism; D007797:Laminin / Q000378:metabolism* / Q000494:pharmacology; D008666:Metalloendopeptidases / Q000378:metabolism*; D016501:Neurites / Q000187:drug effects; D016716:PC12 Cells; D010446:Peptide Fragments / Q000378:metabolism / Q000494:pharmacology; D051381:Rats
chemical_list: C105786:AG 73; D007797:Laminin; D010446:Peptide Fragments; C084098:laminin A; D018093:Gelatinases; D008666:Metalloendopeptidases