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Caspase‐dependent cleavage of 170‐kDa P‐glycoprotein during apoptosis of human T‐lymphoblastoid CEM cells
Author(s) -
Mantovani Irina,
Cappellini Alessandra,
Tazzari Pier Luigi,
Papa Veronica,
Cocco Lucio,
Martelli Alberto M.
Publication year - 2006
Publication title -
journal of cellular physiology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.529
H-Index - 174
eISSN - 1097-4652
pISSN - 0021-9541
DOI - 10.1002/jcp.20628
Subject(s) - apoptosis , programmed cell death , caspase , microbiology and biotechnology , biology , ly294002 , lymphoblast , cell culture , inhibitor of apoptosis , protein kinase b , caspase 3 , caspase 8 , biochemistry , genetics
Multidrug resistance (MDR) mediated by the drug efflux protein, 170‐kDa P‐glycoprotein (P‐gp), is one mechanism that tumor cells use to escape cell death induced by chemotherapeutic drugs. Moreover, evidence suggests that cell lines expressing high levels of 170‐kDa P‐gp are less sensitive to caspase‐mediated apoptosis induced by a wide range of death stimuli, including Fas ligand, tumor necrosis factor, and ultraviolet irradiation. However, the fate of 170‐kDa P‐gp during apoptosis is unknown. In this study, we demonstrate for the first time that 170‐kDa P‐gp is cleaved during apoptosis of VBL100 human T‐lymphoblastoid CEM cells. Apoptotic cell death was induced by LY294002 (a pharmacological inhibitor of the phosphoinositide 3‐kinase/Akt survival pathway), H 2 O 2 , and Z‐LEHD‐FMK (a caspase‐9 inhibitor which has been recently reported to induce apoptosis in CEM cells). Using an antibody to a common epitope present in both the third and the sixth extracellular loop of P‐gp, two cleavage products were detected, with an apparent molecular weight of 80 and 85 kDa. DEVD‐FMK (a caspase‐3 inhibitor), but not VEID‐CHO (a caspase‐6 inhibitor), blocked 170‐kDa P‐gp cleavage. Recombinant caspase‐3 was able to cleave in vitro 170‐kDa P‐gp yielding two fragments of equal size to those generated in vivo. Considering the size of the cleaved fragments and their reactivity with antibodies, which recognize either the N‐half or the C‐half region of the protein, it is conceivable that the cleavage occurs intracytoplasmically. Since 170‐kDa P‐gp has been reported to counteract apoptosis, its cleavage may be a mechanism aimed at blocking an important cell survival component. J. Cell. Physiol. © 2006 Wiley‐Liss, Inc.