Pro-urokinase-type plasminogen activator is a substrate for hepsin.
Hepsin, a type II transmembrane serine protease, is strongly up-regulated in prostate cancer. Hepsin overexpression in a mouse prostate cancer model resulted in tumor progression and metastasis, associated with basement membrane disorganization. We investigated whether hepsin enzymatic activity was linked to the basement membrane defects by examining its ability to initiate the plasminogen/plasmin proteolytic pathway. Because plasminogen is not processed by hepsin, we investigated the upstream activators, urokinase-type plasminogen activator (uPA) and tissue-type plasminogen activator. Enzymatic assays with a recombinant soluble form of hepsin demonstrated that hepsin did not cleave pro-tissue-type plasminogen activator but efficiently converted pro-uPA into high molecular weight uPA by cleavage at the Lys158-Ile159 (P1-P1') peptide bond. uPA generated by hepsin displayed enzymatic activity toward small synthetic and macromolecular substrates indistinguishable from uPA produced by plasmin. The catalytic efficiency of pro-uPA activation by hepsin (kcat/Km 4.8 x 10(5) m(-1) s(-1)) was similar to that of plasmin, which is considered the most potent pro-uPA activator and was about 6-fold higher than that of matriptase. Conversion of pro-uPA was also demonstrated with cell surface-expressed full-length hepsin. A stable hepsinoverexpressing LnCaP cell line converted pro-uPA into high molecular weight uPA at a rate of 6.6 +/- 1.9 nm uPA h(-1), which was about 3-fold higher than LnCaP cells expressing lower hepsin levels on their surface. In conclusion, the ability of hepsin to efficiently activate pro-uPA suggests that it may initiate plasmin-mediated proteolytic pathways at the tumor/stroma interface that lead to basement membrane disruption and tumor progression.
Moran
Paul
P
Department of Protein Engineering, Genentech, Inc., South San Francisco, California 94080, USA.
Li
Wei
W
Fan
Bin
B
Vij
Rajesh
R
Eigenbrot
Charles
C
Kirchhofer
Daniel
D
D000818:Animals; D004195:Disease Models, Animal; D005341:Fibrinolysin / Q000378:metabolism; D006801:Humans; D008297:Male; D051379:Mice; D008807:Mice, Inbred BALB C; D009362:Neoplasm Metastasis; D011471:Prostatic Neoplasms / Q000378:metabolism; D011485:Protein Binding; D012697:Serine Endopeptidases / Q000737:chemistry*; D013379:Substrate Specificity; D010959:Tissue Plasminogen Activator / Q000378:metabolism; D014568:Urokinase-Type Plasminogen Activator / Q000737:chemistry* / Q000378:metabolism
D012697:Serine Endopeptidases; C055645:hepsin; C485595:matriptase; D010959:Tissue Plasminogen Activator; D005341:Fibrinolysin; D014568:Urokinase-Type Plasminogen Activator