title: Membrane type matrix metalloproteinase 1 activates pro-gelatinase A without furin cleavage of the N-terminal domain.
abstract: Membrane type matrix metalloproteinase 1 (MT-MMP1), a novel 63-kDa member of the matrix metalloproteinase family, is a membrane-anchored enzyme and an activator for gelatinase A. In addition to its C-terminal hydrophobic transmembrane domain, MT-MMP1 has an insertion of 11 amino acids between its propeptide and catalytic domain encrypted with a RRKR recognition motif for the paired basic amino acid cleaving enzyme, furin. In this report, we investigated whether the cleavage of the RRKR motif of MT-MMP1 by Golgi-associated furin is analogous to a similar enzyme activation mechanism observed with stromelysin-3. Mutant forms of MT-MMP1 were cotransfected into COS-1 cells with cDNAs for pro-gelatinase A and/or furin. Immunoprecipitation and immunoblotting using specific antibodies were employed to characterize cell proteins. Whereas furin readily cleaved soluble MT-MMP1 lacking the transmembrane domain (DeltaMT-MMP1), a soluble stromelysin-1/DeltaMT-MMP1 chimera without the RRKR basic motif was resistant to furin-induced cleavage. COS-1 cells cotransfected with wild type MT-MMP1 cDNA and furin cDNA demonstrated a 63-kDa protein (latent enzyme) on SDS-polyacrylamide gel electrophoresis rather than the anticipated lower molecular weight activated enzyme. Inhibition of furin activity with alpha1-protease inhibitorPittsburgh (a furin inhibitor) did not affect the pro-gelatinase A activation mechanism in COS-1 cells cotransfected with MT-MMP1 and pro-gelatinase A cDNAs. Furthermore, substitution of the RRKR motif of MT-MMP1 with alanine residues by site-directed mutagenesis resulted in the same 63-kDa protein without loss of pro-gelatinase A activation function. These data indicate that furin-induced activation of MT-MMP1 is not a prerequisite for pro-gelatinase A activation. The mechanism of activation of cell-bound MT-MMP1 remains to be elucidated.
authors.affiliation: Department of Medicine, the State University of New York, Stony Brook, New York 11794, USA.
authors.forename: J
authors.initials: J
authors.lastname: Cao
authors.forename: A
authors.initials: A
authors.lastname: Rehemtulla
authors.forename: W
authors.initials: W
authors.lastname: Bahou
authors.forename: S
authors.initials: S
authors.lastname: Zucker
mesh_terms: D000818:Animals; D019556:COS Cells; D002462:Cell Membrane / Q000201:enzymology; D017364:Collagenases / Q000235:genetics / Q000378:metabolism*; D018076:DNA, Complementary; D004789:Enzyme Activation; D004792:Enzyme Precursors / Q000235:genetics / Q000378:metabolism*; D045683:Furin; D018093:Gelatinases / Q000235:genetics / Q000378:metabolism*; D006868:Hydrolysis; D020781:Matrix Metalloproteinase 1; D008666:Metalloendopeptidases / Q000235:genetics / Q000378:metabolism*; D011499:Protein Processing, Post-Translational; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D013381:Subtilisins / Q000378:metabolism*; D014162:Transfection
chemical_list: D018076:DNA, Complementary; D004792:Enzyme Precursors; D011993:Recombinant Fusion Proteins; D013381:Subtilisins; D045683:Furin; D017364:Collagenases; D018093:Gelatinases; D008666:Metalloendopeptidases; C069860:progelatinase; D020781:Matrix Metalloproteinase 1