Nitric oxide (NO) may block apoptosis by inhibiting caspases via S-nitrosylation of cysteines. Here, we investigated whether effector caspases might cleave and thereby inhibit endothelial nitric oxide synthase (eNOS). Exposure of eNOS-transfected COS-7 cells and bovine aortic endothelial cells to staurosporine resulted in significant loss of 135-kDa eNOS protein and activity, and appearance of a 60-kDa eNOS fragment; effects were inhibited by the general caspase inhibitor, benzyloxycarbonyl-Val-Ala-Asp[OMe]-fluoromethyl ketone (zVAD-fmk). In eNOS-transfected COS-7 cells, staurosporine-induced activation of caspase-3 and poly(ADP-ribose) polymerase (PARP) cleavage coincided with increased eNOS degradation and decreased activity. Loss of eNOS activity was greater than the degree of proteolysis. Incubation of immunoprecipitated eNOS with caspase-3, caspase-6 or caspase-7 resulted in eNOS cleavage. Staurosporine, a general protein kinase inhibitor, also reduced phosphorylation and decreased calmodulin binding, an effect that may explain the reduction in activity. eNOS, therefore, is both an inhibitor of apoptosis and a target of apoptosis-associated proteolysis.

Tesauro, M., Thompson, W., Moss, J. (2006). Effect of staurosporine-induced apoptosis on endothelial nitric oxide synthase in transfected COS-7 cells and primary endothelial cells. CELL DEATH AND DIFFERENTIATION, 13(4), 597-606 [10.1038/sj.cdd.4401770].

Effect of staurosporine-induced apoptosis on endothelial nitric oxide synthase in transfected COS-7 cells and primary endothelial cells

TESAURO, MANFREDI;
2006-01-01

Abstract

Nitric oxide (NO) may block apoptosis by inhibiting caspases via S-nitrosylation of cysteines. Here, we investigated whether effector caspases might cleave and thereby inhibit endothelial nitric oxide synthase (eNOS). Exposure of eNOS-transfected COS-7 cells and bovine aortic endothelial cells to staurosporine resulted in significant loss of 135-kDa eNOS protein and activity, and appearance of a 60-kDa eNOS fragment; effects were inhibited by the general caspase inhibitor, benzyloxycarbonyl-Val-Ala-Asp[OMe]-fluoromethyl ketone (zVAD-fmk). In eNOS-transfected COS-7 cells, staurosporine-induced activation of caspase-3 and poly(ADP-ribose) polymerase (PARP) cleavage coincided with increased eNOS degradation and decreased activity. Loss of eNOS activity was greater than the degree of proteolysis. Incubation of immunoprecipitated eNOS with caspase-3, caspase-6 or caspase-7 resulted in eNOS cleavage. Staurosporine, a general protein kinase inhibitor, also reduced phosphorylation and decreased calmodulin binding, an effect that may explain the reduction in activity. eNOS, therefore, is both an inhibitor of apoptosis and a target of apoptosis-associated proteolysis.
Pubblicato
Rilevanza internazionale
Articolo
Esperti anonimi
Settore MED/09 - Medicina Interna
English
Amino Acid Chloromethyl Ketones; Animals; COS Cells; Calmodulin; Caspase 3; Caspase 6; Caspase 7; Caspase Inhibitors; Caspases; Cattle; Cercopithecus aethiops; Cysteine Proteinase Inhibitors; Dose-Response Relationship, Drug; Endothelial Cells; Nitric Oxide Synthase Type III; Poly(ADP-ribose) Polymerases; Protein Binding; Time Factors; Transfection; Apoptosis; Protein Kinase Inhibitors; Staurosporine
Tesauro, M., Thompson, W., Moss, J. (2006). Effect of staurosporine-induced apoptosis on endothelial nitric oxide synthase in transfected COS-7 cells and primary endothelial cells. CELL DEATH AND DIFFERENTIATION, 13(4), 597-606 [10.1038/sj.cdd.4401770].
Tesauro, M; Thompson, W; Moss, J
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Utilizza questo identificativo per citare o creare un link a questo documento: https://hdl.handle.net/2108/120756
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