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First published online May 4, 2004
doi: 10.1242/10.1242/jcs.01068

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Processing and activation of latent heparanase occurs in lysosomes

Anna Zetser^1,*, Flonia Levy-Adam^1,*, Victoria Kaplan¹, Svetlana Gingis-Velitski¹, Yulia Bashenko¹, Shay Schubert¹, Moshe Y. Flugelman², Israel Vlodavsky^1, and Neta Ilan¹

¹ Cancer and Vascular Biology Research Center, The Bruce Rappaport Faculty of Medicine, Technion, Haifa 31096, Israel
² Department of Cardiology, Lady Davis Carmel Medical Center, Haifa 34632, Israel

Author for correspondence (e-mail: vlodavsk{at}cc.huji.ac.il)

Accepted 22 December 2003

Heparanase is a heparan sulfate degrading endoglycosidase participating in extracellular matrix degradation and remodeling. Heparanase is synthesized as a 65 kDa non-active precursor that subsequently undergoes proteolytic cleavage, yielding 8 kDa and 50 kDa protein subunits that heterodimerize to form an active enzyme. The protease responsible for heparanase processing is currently unknown, as is the sub-cellular processing site. In this study, we characterize an antibody (733) that preferentially recognizes the active 50 kDa heparanase form as compared to the non-active 65 kDa heparanase precursor. We have utilized this and other anti-heparanase antibodies to study the cellular localization of the latent 65 kDa and active 50 kDa heparanase forms during uptake and processing of exogenously added heparanase. Interestingly, not only the processed 50 kDa, but also the 65 kDa heparanase precursor was localized to perinuclear vesicles, suggesting that heparanase processing occurs in lysosomes. Indeed, heparanase processing was completely inhibited by chloroquine and bafilomycin A1, inhibitors of lysosome proteases. Similarly, processing of membrane-targeted heparanase was also chloroquine-sensitive, further ruling out the plasma membrane as the heparanase processing site. Finally, we provide evidence that antibody 733 partially neutralizes the enzymatic activity of heparanase, suggesting that the N-terminal region of the molecule is involved in assuming an active conformation. Monoclonal antibodies directed to this region are likely to provide specific heparanase inhibitors and hence assist in resolving heparanase functions under normal and pathological conditions.

Key words: Heparanase, Localization, Processing, Uptake, Lysosome, Antibody

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