Activation of type IV procollagenases by human tumor-associated trypsin-2

Increased production of proteinases, such as matrix metalloproteinases (MMPs), is a characteristic feature of malignant tumors. Some human cancers :Ind cell lines derived from them also express trypsinogen, but the function of the extrapancreatic trypsin has remained unclear, In this study we cloned and sequenced trypsinogen-a cDNA from human COLO 205 colon carcinoma cells and characterized the ability of the enzyme to activate latent human type TV procollagenases (proMMP-2 and proMMP-9). As shown by cloning and N-terminal amino acid sequencing, the amino acid sequence of tumor associated trypsin-a is identical to that of pancreatic trypsin-a. We found that both pancreatic trypsin-a and tumor cell-derived trypsin-a are efficient activators of proMMP-9 and are capable of activating proMMP-9 at a molar ratio of 1:1000, the lowest reported so far. Human trypsin-a was a more efficient activator than widely used bovine trypsin and converted the 92-kDa proMMP-9 to a single 77-kDa product that was not fragmented further. The single peptide bond cleaved by trypsin-a in proMMP-9 was Arg(87)-Phe(88), The generation of the 77-kDa species coincided with the increase in specific activity of MMP-9. In contrast, trypsin-2 only partially activated proMMP-2. Trypsin-a cleaved the Arg(99)-Lys(100) peptide bond of proMMP-2 generating 62-65-kDa MMP-2 species. Trypsin-a-induced proMMP-2 and -9 conversions were inhibited by tumor-associated trypsin inhibitor added either prior to or during activation indicating that proMMPs were not activated autocatalytically. Trypsin-2 also activated proMMPs associated with tissue inhibitor of matrix metalloproteinases, the complexes of which are thought to be the major MMP forms in vivo. The ability of human tumor cell-derived trypsin-a to activate latent MMPs suggests a role fbr trypsin-a in initiating the proteinase cascade that mediates tumor invasion and metastasis formation.