Calpain activation is upstream of caspases in radiation-induced apoptosis.
The molecular events involved in apoptosis induced by ionizing radiation remain unresolved. In this paper we show that the cleavage of fodrin to a 150 kDa fragment is an early proteolytic event in radiation-induced apoptosis in the Burkitts' Lymphoma cell line BL30A and requires 100 microM zVAD-fmk for inhibition. Caspases-1, -3, -6 and -7 were shown to cleave fodrin to the 150 kDa fragment in vitro and all were inhibited by 10 microM zVAD-fmk. We also show that the in vitro cleavage of fodrin by calpain is inhibited by 100 microM zVAD-fmk as was the calpain-mediated hydrolysis of casein. We demonstrate that calpain is activated within 15 min after radiation exposure, concomitant with the cleavage of fodrin to the 150 kDa fragment whereas caspase-3 is activated at 2 h correlating with the cleavage of fodrin to the 120 kDa fragment. These results support a role for calpain in the early phases of the radiation-induced apoptosis pathway, upstream of the caspases.
Waterhouse
N J
NJ
Queensland Cancer Fund Research Unit, Queensland Institute of Medical Research, Royal Brisbane Hospital, Herston, Australia.
Finucane
D M
DM
Green
D R
DR
Elce
J S
JS
Kumar
S
S
Alnemri
E S
ES
Litwack
G
G
Khanna
K
K
Lavin
M F
MF
Watters
D J
DJ
D000590:Amino Acid Chloromethyl Ketones / Q000494:pharmacology; D000595:Amino Acid Sequence; D017209:Apoptosis / Q000187:drug effects / Q000502:physiology* / Q000528:radiation effects; D002051:Burkitt Lymphoma; D002154:Calpain / Q000037:antagonists & inhibitors / Q000378:metabolism*; D002352:Carrier Proteins / Q000235:genetics / Q000378:metabolism; D020170:Caspase 1 / Q000378:metabolism; D053148:Caspase 3; D053178:Caspase 6; D053179:Caspase 7; D061945:Caspase Inhibitors; D020169:Caspases / Q000378:metabolism*; D015853:Cysteine Proteinase Inhibitors / Q000494:pharmacology; D004305:Dose-Response Relationship, Drug; D004789:Enzyme Activation; D006801:Humans; D008565:Membrane Proteins / Q000235:genetics / Q000378:metabolism; D008840:Microfilament Proteins / Q000235:genetics / Q000378:metabolism; D008969:Molecular Sequence Data; D010446:Peptide Fragments / Q000378:metabolism; D014407:Tumor Cells, Cultured / Q000166:cytology / Q000201:enzymology / Q000528:radiation effects; D019014:fas Receptor; D013049:Spectrin
D000590:Amino Acid Chloromethyl Ketones; D002154:Calpain; D002352:Carrier Proteins; D020170:Caspase 1; D053148:Caspase 3; D053178:Caspase 6; D053179:Caspase 7; D061945:Caspase Inhibitors; D020169:Caspases; D015853:Cysteine Proteinase Inhibitors; D008565:Membrane Proteins; D008840:Microfilament Proteins; D010446:Peptide Fragments; D019014:fas Receptor; D013049:Spectrin; C096713:benzyloxycarbonylvalyl-alanyl-aspartyl fluoromethyl ketone; C505407:CASP3 protein, human; C505514:CASP6 protein, human; C505520:CASP7 protein, human