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

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)-Va(102) and Trp(105)-Asn(106) in loop 3 of the NH2-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.