Accessible Unlicensed Requires Authentication Published by De Gruyter June 1, 2005

Endosomal-Lysosomal Proteolysis Mediates Death Signalling by TNFα, Not by Etoposide, in L929 Fibrosarcoma Cells: Evidence for an Active Role of Cathepsin D

M. Démoz, R. Castino, P. Cesaro, F. M. Baccino, G. Bonelli and C. Isidoro
From the journal

Abstract

In several in vitro models of apoptosis, lysosomal proteolysis has been shown to play an active role in mediating the death signal by cytokines or antiblastic drugs. Depending on the experimental cell model and the cytotoxic stimulus applied, an increased expression and the cytosolic translocation of either cathepsin D or B have been reported in apoptotic cells. We have analysed the involvement of these lysosomal proteases in a canonical apoptotic cell model, namely L929 fibroblasts, in which apoptosis was induced by cytotoxic agents acting through different mechanisms: (i) the cytokine TNFα, which triggers the cell suicide via interaction with its membrane receptor, and (ii) the topoisomerase IIinhibitor etoposide (VP16), which directly causes DNA damage. In both cases the activity of cathepsins B and D increased in apoptosing cultures. CA074-Me, a specific inhibitor of cathepsin B, and Leupeptin, a broad inhibitor of serine and cysteine proteases (among which is cathepsin B), did not exert any protection from TNFα. In contrast, preloading the cells with pepstatin A, a specific inhibitor of cathepsin D, protected L929 cells from TNFα cytotoxicity by more than 50%. However, no protection was observed if pepstatin A was added concomitantly with the cytokine. Inhibition of either cathepsin B or D did not impede apoptosis induced by etoposide. Lysosomal integrity was preserved and cathepsin D remained still confined in vesicular structures in apoptotic cells treated with either TNFα or etoposide. It follows that proteolysis by cathepsin D is likely to represent an early event in the death pathway triggered by TNFα and occurs within the endosomal lysosomal compartment.

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Published Online: 2005-06-01
Published in Print: 2002-08-27

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