Glutamate-cysteine ligase attenuates TNF-induced mitochondrial injury and apoptosis.
Glutathione (GSH) is important in free radical scavenging, maintaining cellular redox status, and regulating cell survival in response to a wide variety of toxicants. The rate-limiting enzyme in GSH synthesis is glutamate-cysteine ligase (GCL), which is composed of catalytic (GCLC) and modifier (GCLM) subunits. To determine whether increased GSH biosynthetic capacity enhances cellular resistance to tumor necrosis factor-alpha- (TNF-alpha-) induced apoptotic cell death, we have established several mouse liver hepatoma (Hepa-1) cell lines overexpressing GCLC and/or GCLM. Cells overexpressing GCLC alone exhibit modest increases in GCL activity, while cells overexpressing both subunits have large increases in GCL activity. Importantly, cells overexpressing both GCL subunits exhibit increased resistance to TNF-induced apoptosis as judged by a loss of redox potential; mitochondrial membrane potential; translocation of cytochrome c to the cytoplasm; and activation of caspase-3, caspase-8, and caspase-9. Analysis of the effects of TNF on these parameters indicates that maintaining mitochondrial integrity mediates this protective effect in GCL-overexpressing cells.
Botta
Dianne
D
Department of Environmental and Occupational Health Sciences, University of Washington, Seattle, WA 98195, USA.
Franklin
Christopher C
CC
White
Collin C
CC
Krejsa
Cecile M
CM
Dabrowski
Michael J
MJ
Pierce
Robert H
RH
Fausto
Nelson
N
Kavanagh
Terrance J
TJ
D000818:Animals; D017209:Apoptosis / Q000187:drug effects*; D006528:Carcinoma, Hepatocellular; D045744:Cell Line, Tumor; D005721:Glutamate-Cysteine Ligase / Q000378:metabolism*; D005978:Glutathione / Q000378:metabolism; D006801:Humans; D008113:Liver Neoplasms; D051379:Mice; D008928:Mitochondria / Q000187:drug effects / Q000473:pathology*; D011994:Recombinant Proteins / Q000378:metabolism; D014162:Transfection; D014409:Tumor Necrosis Factor-alpha / Q000633:toxicity*
D011994:Recombinant Proteins; D014409:Tumor Necrosis Factor-alpha; D005721:Glutamate-Cysteine Ligase; D005978:Glutathione