title: Identification of a novel 82 kDa proMMP-9 species associated with the surface of leukaemic cells: (auto-)catalytic activation and resistance to inhibition by TIMP-1.
abstract: MMP-9 (matrix metalloproteinase 9) plays a critical role in tumour progression. Although the biochemical properties of the secreted form of proMMP-9 are well characterized, little is known about the function and activity of cell surface-associated proMMP-9. We purified a novel 82 kDa species of proMMP-9 from the plasma membrane of THP-1 leukaemic cells, which has substantial differences from the secreted 94 kDa proMMP-9. The 82 kDa form was not detected in the medium even upon stimulation with a phorbol ester. It is truncated by nine amino acid residues at its N-terminus, lacks O-linked oligosaccharides present in the 94 kDa proMMP-9, but retains N-linked carbohydrates. Incubation of 94 kDa proMMP-9 with MMP-3 generated the well-known 82 kDa active form, but the 82 kDa proMMP-9 was converted into an active species of 35 kDa, which was also produced by autocatalytic processing in the absence of activating enzymes. The activated 35 kDa MMP-9 efficiently degraded gelatins, native collagen type IV and fibronectin. The enzyme was less sensitive to TIMP-1 (tissue inhibitor of metalloproteinase 1) inhibition with IC50 values of 82 nM compared with 1 nM for the 82 kDa active MMP-9. The synthetic MMP inhibitor GM6001 blocked the activity of both enzymes, with similar IC50 values below 1 nM. The 82 kDa proMMP-9 is also produced in HL-60 and NB4 leukaemic cell lines as well as ex vivo leukaemic blast cells. It is, however, absent from neutrophils and mononuclear cells isolated from peripheral blood of healthy individuals. Thus, the 82 kDa proMMP-9 expressed on the surface of malignant cells may escape inhibition by natural TIMP-1, thereby facilitating cellular invasion in vivo.
authors.affiliation: Division of Clinical Chemistry and Clinical Biochemistry in the Surgical Department, Ludwig-Maximilians-University of Munich, 80336 Munich, Germany. christian.ries@med.uni-muenchen.de
authors.forename: Christian
authors.initials: C
authors.lastname: Ries
authors.forename: Thomas
authors.initials: T
authors.lastname: Pitsch
authors.forename: Reinhard
authors.initials: R
authors.lastname: Mentele
authors.forename: Stefan
authors.initials: S
authors.lastname: Zahler
authors.forename: Virginia
authors.initials: V
authors.lastname: Egea
authors.forename: Hideaki
authors.initials: H
authors.lastname: Nagase
authors.forename: Marianne
authors.initials: M
authors.lastname: Jochum
mesh_terms: D000595:Amino Acid Sequence; D002384:Catalysis; D045744:Cell Line, Tumor; D004789:Enzyme Activation; D004792:Enzyme Precursors / Q000037:antagonists & inhibitors / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D005354:Fibrosarcoma / Q000378:metabolism; D015971:Gene Expression Regulation, Enzymologic; D006801:Humans; D007938:Leukemia / Q000378:metabolism* / Q000473:pathology*; D020780:Matrix Metalloproteinase 9 / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D061965:Matrix Metalloproteinase Inhibitors; D008565:Membrane Proteins; D021381:Protein Transport; D013379:Substrate Specificity; D019715:Tissue Inhibitor of Metalloproteinase-1 / Q000235:genetics / Q000378:metabolism*
chemical_list: D004792:Enzyme Precursors; D061965:Matrix Metalloproteinase Inhibitors; D008565:Membrane Proteins; D019715:Tissue Inhibitor of Metalloproteinase-1; C120487:pro-matrix metalloproteinase 9; D020780:Matrix Metalloproteinase 9