Calpain-dependent cleavage of cain/cabin1 activates calcineurin to mediate calcium-triggered cell death.
Cain/cabin1 is an endogenous inhibitor of calcineurin (Cn), a calcium-dependent serine/threonine phosphatase involved in various cellular functions including apoptosis. We show here that during apoptosis cain/cabin1 is cleaved by calpain at the carboxyl terminus to generate a cleavage product with a molecular mass of 32 kDa as a necessary step leading to Cn-mediated cell death. Mouse cain/cabin1 was identified from a thymus cDNA library by an in vitro substrate-screening assay with calpain. Exposure of Jurkat cells to the calcium ionophore, induced cain/cabin1 cleavage and cell death, accompanied by activation of calpain and Cn. The calpain inhibitors, calpeptin and zLLY, suppressed both -induced cain/cabin1 cleavage and Cn activation, indicating that Cn activation and cain/cabin1 cleavage are calpain-dependent. Expression of cain/cabin1 or a catalytically inactive Cn mutant [CnA beta(2)(1-401/H160N)] and treatment with FK506 reduced -induced cell death. In vitro calpain cleavage and immunoprecipitation assays with deletion mutants of cain/cabin1 showed that cleavage occurred in the Cn-binding domain of cain/cabin1, indicating that the cleavage at its C terminus by calpain prevented cain/cabin1 from binding to Cn. In addition, in vitro binding assays showed that cain/cabin1 bound to the Cn B-binding domain of Cn A. Taken together, these results indicate that calpain cleaves the calcineurin-binding domain of cain/cabin1 to activate Cn and elicit calcium-triggered cell death.
Department of Life Science, Kwangju Institute of Science and Technology, 1 Oryong-dong, Puk-gu, Kwangju 500-712, Korea.
Min-Jung
MJ
Kim
Dong-Gyu
DG
Jo
Gil-Sun
GS
Hong
Byung Ju
BJ
Kim
Michael
M
Lai
Dong-Hyung
DH
Cho
Ki-Woo
KW
Kim
Arun
A
Bandyopadhyay
Yeon-Mi
YM
Hong
Do Han
DH
Kim
Chunghee
C
Cho
Jun O
JO
Liu
Solomon H
SH
Snyder
Yong-Keun
YK
Jung
D048868:Adaptor Proteins, Signal Transducing; D000818:Animals; D017209:Apoptosis / Q000187:drug effects / Q000502:physiology*; D051017:Apoptosis Regulatory Proteins; D001665:Binding Sites; D015153:Blotting, Western; D000001:Calcimycin / Q000494:pharmacology; D019703:Calcineurin / Q000378:metabolism*; D002118:Calcium / Q000502:physiology*; D002122:Calcium Chloride / Q000494:pharmacology; D002154:Calpain / Q000378:metabolism*; D002352:Carrier Proteins / Q000378:metabolism*; D053148:Caspase 3; D020169:Caspases / Q000378:metabolism; D016923:Cell Death / Q000502:physiology*; D003001:Cloning, Molecular; D015853:Cysteine Proteinase Inhibitors / Q000494:pharmacology; D004789:Enzyme Activation; D015723:Gene Library; D006801:Humans; D047908:Intracellular Signaling Peptides and Proteins; D019169:Jurkat Cells; D007700:Kinetics; D051379:Mice; D010750:Phosphoproteins / Q000378:metabolism*; D016133:Polymerase Chain Reaction; D051381:Rats; D011994:Recombinant Proteins / Q000378:metabolism; D013601:T-Lymphocytes / Q000502:physiology; D016559:Tacrolimus / Q000494:pharmacology
D048868:Adaptor Proteins, Signal Transducing; D051017:Apoptosis Regulatory Proteins; C491861:CABIN1 protein, human; C113151:Cabin1 protein, mouse; C113573:Cabin1 protein, rat; D002352:Carrier Proteins; D015853:Cysteine Proteinase Inhibitors; D047908:Intracellular Signaling Peptides and Proteins; D010750:Phosphoproteins; D011994:Recombinant Proteins; D000001:Calcimycin; D019703:Calcineurin; C505407:CASP3 protein, human; D002154:Calpain; C505409:Casp3 protein, mouse; C505410:Casp3 protein, rat; D053148:Caspase 3; D020169:Caspases; D002122:Calcium Chloride; D002118:Calcium; D016559:Tacrolimus