Cleavage of the calpain inhibitor, calpastatin, during apoptosis.
Calpain activity is thought to be essential for the execution of apoptotic cell death in certain experimental models. In the present study, the physiological inhibitor of calpain, calpastatin, was found to be cleaved in three different apoptotic systems. The 110-120 kDa calpastatin protein of Jurkat T-lymphocytes and U937 monocytic leukemia cells was cleaved to a 65-70 kDa form after the induction of apoptosis with anti-CD95 monoclonal antibody, staurosporine or TNF. Cleavage of calpastatin in apoptotic cells occurred simultaneously with the cleavage of the DNA repair enzyme, poly(ADP-ribose) polymerase. The caspase inhibitors VAD-cmk and IETD-fmk prevented calpastatin cleavage in all three systems. Calpain inhibitor I, however, suppressed calpastatin cleavage only during TNF-induced apoptosis. Other protease inhibitors, such as lactacystin and pepstatin A, did not confer any significant protection against apoptotic calpastatin cleavage. The results from in vitro incubations with cell lysates and purified enzymes showed that calpain I, calpain II and recombinant caspase-3, all cleaved calpastatin, with varying efficiency. In conclusion, the results of the present study suggest that caspases may cleave calpastatin and thus, regulate calpain activity during apoptotic cell death.
Pörn-Ares
M I
MI
Institute of Environmental Medicine, Karolinska Institutet, Stockholm, Sweden. Isabella.Ares@imm.ki.se
Samali
A
A
Orrenius
S
S
D000111:Acetylcysteine / Q000031:analogs & derivatives / Q000494:pharmacology; D017209:Apoptosis / Q000502:physiology*; D002135:Calcium-Binding Proteins / Q000378:metabolism*; D002154:Calpain / Q000378:metabolism*; D053148:Caspase 3; D020169:Caspases / Q000378:metabolism; D015853:Cysteine Proteinase Inhibitors / Q000378:metabolism*; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005434:Flow Cytometry; D006801:Humans; D019169:Jurkat Cells / Q000166:cytology* / Q000187:drug effects / Q000201:enzymology; D010436:Pepstatins / Q000494:pharmacology; D011480:Protease Inhibitors / Q000494:pharmacology; D019311:Staurosporine / Q000494:pharmacology; D014409:Tumor Necrosis Factor-alpha / Q000494:pharmacology; D020298:U937 Cells; D019014:fas Receptor / Q000494:pharmacology
D002135:Calcium-Binding Proteins; D015853:Cysteine Proteinase Inhibitors; D004791:Enzyme Inhibitors; D010436:Pepstatins; D011480:Protease Inhibitors; D014409:Tumor Necrosis Factor-alpha; D019014:fas Receptor; C014195:Streptomyces pepsin inhibitor; C067713:lactacystin; C033668:calpastatin; C505407:CASP3 protein, human; D002154:Calpain; D053148:Caspase 3; D020169:Caspases; D019311:Staurosporine; C031375:pepstatin; D000111:Acetylcysteine