Mast cell tissue inhibitor of metalloproteinase-1 is cleaved and inactivated extracellularly by alpha-chymase

• Published in: The Journal of immunology (1950) Vol. 166; no. 4; pp. 2783 - 2792
• Main Authors: Frank, B T, Rossall, J C, Caughey, G H, Fang, K C
• Format: Journal Article
• Language: English
• Published: United States 15.02.2001
• Subjects: a-chymase; Acute-Phase Proteins; Amino Acid Sequence; Animals; Binding Sites; Carrier Proteins - metabolism; Cell Degranulation; Cells, Cultured; Chymases; Dogs; Enzyme Activation; Enzyme Precursors - metabolism; Extracellular Space - enzymology; Extracellular Space - metabolism; Gelatinases - metabolism; Humans; Hydrolysis; Leukocyte Elastase - metabolism; Lipocalin-2; Lipocalins; Macromolecular Substances; Mast Cells - enzymology; Mast Cells - metabolism; Mast-Cell Sarcoma - enzymology; Mast-Cell Sarcoma - metabolism; Metalloendopeptidases - metabolism; Mice; Mice, Inbred C57BL; Molecular Sequence Data; Neutrophils - enzymology; Neutrophils - metabolism; Oncogene Proteins; Protein Processing, Post-Translational; Proto-Oncogene Proteins; Serine Endopeptidases - metabolism; TIMP-1 protein; Tissue Inhibitor of Metalloproteinase-1 - antagonists & inhibitors; Tissue Inhibitor of Metalloproteinase-1 - biosynthesis; Tissue Inhibitor of Metalloproteinase-1 - metabolism; Tumor Cells, Cultured
• ISSN: 0022-1767; 1550-6606
• DOI: 10.4049/jimmunol.166.4.2783

We previously reported that mast cell alpha-chymase cleaves and activates progelatinase B (progel B). Outside of cells, progel B is complexed with tissue inhibitor of metalloproteinase (TIMP)-1, which hinders zymogen activation and inhibits activity of mature forms. The current work demonstrates that dog BR mastocytoma cells, HMC-1 cells, and murine bone marrow-derived mast cells secrete TIMP-1 whose electrophoretic profile in supernatants suggests degranulation-dependent proteolysis. Alpha-chymase cleaves uncomplexed TIMP-1, reducing its ability to inhibit gel B, whereas tryptase has no effect. Sequencing of TIMP-1's alpha-chymase-mediated cleavage products reveals hydrolysis at Phe(12)-Cys(13) and Phe(23)-Val(24) in loop 1 and Phe(101)-Val(102) and Trp(105)-Asn(106) in loop 3 of the NH(2)-terminal domain. TIMP-1 in a ternary complex with progel B and neutrophil gelatinase-associated lipocalin is also susceptible to alpha-chymase cleavage, yielding products like those resulting from processing of free TIMP-1. Thus, alpha-chymase cleaves free and gel B-bound TIMP-1. Incubation of the progel B-TIMP-1-neutrophil gelatinase-associated lipocalin complex with alpha-chymase increases gel B activity 2- to 5-fold, suggesting that alpha-chymase activates progel B whether it exists as free monomer or as a complex with TIMP-1. Furthermore, inhibition of alpha-chymase blocks degranulation-induced TIMP-1 processing (absent in alpha-chymase-deficient HMC-1 cells). Purified alpha-chymase processes TIMP-1 in BR supernatants, generating products like those induced by degranulation. In summary, these results suggest that controlled exocytosis of mast cell alpha-chymase activates progel B even in the presence of TIMP-1. This is the first identification of a protease that overcomes inhibition by bound TIMP-1 to activate progel B without involvement of other proteases.