THE CLEAVAGE OF PROUROKINASE TYPE PLASMINOGEN-ACTIVATOR BY STROMELYSIN-1

Membrane binding of urokinase type plasminogen activator (u-PA) is tho ught to play a pivotal role in connective tissue remodeling and invasi ve processes. We compare the ability of different matrix-metalloprotei nases involved in connective tissue turnover to cleave pro-urokinase t ype plasminogen activator between the catalytic domain and the recepto r binding part to investigate a potential role for matrix-metalloprote inases in the regulation of membrane-associated proteolytic activity. We employed several forms of human stromelysin-l (full length, C-trunc ated, and recombinant catalytic domain), rabbit C-truncated stromelysi n-1, the human gelatinases A and B and the human catalytic domain of n eutrophil collagenase. The gelatinases and the collagenase did not sep arate the receptor binding domain of pro-urokinase type plasminogen ac tivator from the catalytic domain, whereas all stromelysin-l forms cle aved the glutamic acid 143-leucine 144 bond of pro-urokinase type plas minogen activator. This reaction could be inhibited by specific inhibi tors of matrix metalloproteinases and was not affected by inhibitors o f serine proteinases. The M-r 31000 cleavage product with leucine 144 as N-terminus displayed no proteolytic activity towards the prourokina se type plasminogen activator substrate pyro-Glu-Gly-Arg-pNA-HCl (S244 4), but it could be activated by an additional treatment with plasmin. Comparison between full length stromelysin-l and its C-truncated form s, showed that both exhibited the same cleavage properties towards pro -urokinase type plasminogen activator. Thus, the cleavage of pro-uroki nase type plasminogen activator by stromelysin-l is not influenced by the presence or absence of the C-terminal domain. The recombinant cata lytic domain of MMP-3 generated pro-urokinase type plasminogen activat or, whereas incubation of pro-urokinase type plasminogen activator wit h the native forms of human or rabbit stromelysin-l led to a moderate activation of pro-uPA due to an additional cleavage that is catalyzed by a serine proteinase.