Cathepsin B mediates the pH-dependent proinvasive activity of tumor-shed microvesicles.
Vesicles shed by cancer cells are known to mediate several tumor-host interactions. Tumor microenvironment may, in turn, influence the release and the activity of tumor-shed microvesicles. In this study, we investigated the molecular mediators of the pH-dependent proinvasive activity of tumor-shed vesicles. Gelatinase zymography showed increased microvesicle activity of matrix metalloproteinases 9 and 2 as a result of acid exposure (pH 5.6) compared to pH 7.4. Thus, we reasoned that the cysteine protease cathepsin B might play a role in mediating the pH-dependent activation of gelatinases. Cathepsin B expression in tumor-shed microvesicles was confirmed by Western blot analysis and zymography. The activity of vesicle-associated cathepsin B measured using Z-Arg-Arg-pNA as substrate was significantly increased at acidic pH values. Inhibition of protease activity by the cysteine protease inhibitor, E-64, and treatment of ovarian cancer cells with small interfering RNA against cathepsin B suppressed the ability of tumor-shed microvesicles to stimulate both gelatinase activation and the invasiveness of endothelial cells observed at low pH values. We conclude that microvesicle shedding is a major secretory pathway for cathepsin B release from tumor cells. Hence, the acidic microenvironment found in most solid tumors may contribute to cathepsin B-mediated proinvasive capabilities of tumor-shed vesicles.
Giusti
Ilaria
I
Department of Experimental Medicine, University of L'Aquila, L'Aquila, Italy.
D'Ascenzo
Sandra
S
Millimaggi
Danilo
D
Taraboletti
Giulia
G
Carta
Gaspare
G
Franceschini
Nicola
N
Pavan
Antonio
A
Dolo
Vincenza
V
D002401:Cathepsin B / Q000235:genetics / Q000378:metabolism*; D045744:Cell Line, Tumor; D002465:Cell Movement; D017077:Culture Media, Conditioned / Q000494:pharmacology; D004789:Enzyme Activation; D005260:Female; D006801:Humans; D006863:Hydrogen-Ion Concentration; D020778:Matrix Metalloproteinase 2 / Q000378:metabolism; D020780:Matrix Metalloproteinase 9 / Q000378:metabolism; D021962:Membrane Microdomains / Q000378:metabolism* / Q000473:pathology*; D009361:Neoplasm Invasiveness; D010051:Ovarian Neoplasms / Q000201:enzymology* / Q000235:genetics / Q000473:pathology; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D034741:RNA, Small Interfering / Q000494:pharmacology; D020133:Reverse Transcriptase Polymerase Chain Reaction
D017077:Culture Media, Conditioned; D012333:RNA, Messenger; D034741:RNA, Small Interfering; D002401:Cathepsin B; D020778:Matrix Metalloproteinase 2; D020780:Matrix Metalloproteinase 9