Overexpression of γ-glutamylcysteine synthetase suppresses tumor necrosis factor-induced apoptosis and activation of nuclear transcription factor-kappa B and activator protein-1

Abstract

Tumor necrosis factor (TNF) is a highly pleiotropic cytokine whose activity is at least partially regulated by the redox status of the cell. The cellular redox status is controlled primarily by glutathione, a major cellular antioxidant, whose synthesis is regulated by the rate-limiting enzyme γ-glutamylcysteine synthetase (γ-GCS). In the present report we investigated the effect of γ-GCS overexpression on the TNF-induced activation of nuclear transcription factors NF-κB and AP-1, stress-activated protein kinase/c-Jun amino-terminal kinase (JNK) and apoptosis. Transfection of cells with γ-GCS cDNA blocked TNF-induced NF-κB activation, cytoplasmic IκBα degradation, nuclear translocation of p65, and NF-κB-dependent gene transcription. γ-GCS overexpression also completely suppressed NF-κB activation induced by phorbol ester and okadaic acid, whereas that induced by H₂O₂, ceramide, and lipopolysaccharide was minimally affected. γ-GCS also abolished the activation of AP-1 induced by TNF and inhibited TNF-induced activation of JNK and mitogen-activated protein kinase kinase. TNF-mediated cytotoxicity and activation of caspase-3 were both abrogated in γ-GCS-overexpressing cells. Overall, our results indicate that most of the pleiotropic actions of TNF are regulated by the glutathione-controlled redox status of the cell.