Regulation of Cu/Zn-superoxide dismutase expression via the phosphatidylinositol 3 kinase/Akt pathway and nuclear factor-kappaB.
Aerobic cells adjust the expression of antioxidant enzymes to maintain reactive oxygen species within tolerable levels. In addition, phosphatidylinositol 3 kinase (PI3K) and its downstream protein kinase effector Akt adapt cells to survive in the presence of oxidative stress. Here we provide evidence for an association between these two defense systems via transcriptional regulation of Cu/Zn-superoxide dismutase (Cu/Zn-SOD). PC12 pheochromocytoma cells expressing active Akt1 exhibit lower ROS levels in response to hydrogen peroxide, as determined with the superoxide-sensitive probe hydroethidine. Transfection of constitutive or 4-hydroxytamoxifen-inducible versions of Akt1 results in higher messenger RNA and protein levels of Cu/Zn-SOD. Luciferase reporter constructs, carrying different length fragments of the human sod1 gene promoter, have identified a region between -552 and -355 that is targeted by PI3K and Akt and that contains a putative site of regulation by nuclear factor-kappaB (NF-kappaB). Nerve growth factor (NGF) and Akt augment the transactivating activity and produce higher nuclear levels of p65-NF-kappaB. Electrophoretic mobility shift assays indicate that the putative NF-kappaB regulatory sequence binds p65-NF-kappaB more efficiently in nuclear extracts from these cells. A dominant-negative mutant of IkappaBalpha further demonstrates that the PI3K/Akt axis targets the sod1 promoter at the level of the newly characterized NF-kappaB site. These results illustrate a new mechanism by which the PI3K/Akt pathway protects cells against oxidative stress, involving the upregulation of Cu/Zn-SOD gene expression, and the results identify NF-kappaB as a key mediator in the regulation of this gene.
Institute of Biomedical Investigation and Department of Biochemistry, Faculty of Medicine, Autónoma University of Madrid, 28029 Madrid, Spain.
Ana I
AI
Rojo
Marta
M
Salinas
Daniel
D
Martín
Rosario
R
Perona
Antonio
A
Cuadrado
D000818:Animals; D015971:Gene Expression Regulation, Enzymologic; D006801:Humans; D006861:Hydrogen Peroxide / Q000494:pharmacology; D016328:NF-kappa B / Q000378:metabolism*; D009474:Neurons / Q000187:drug effects / Q000201:enzymology / Q000378:metabolism*; D018384:Oxidative Stress; D016716:PC12 Cells; D019869:Phosphatidylinositol 3-Kinases / Q000378:metabolism*; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism*; D011518:Proto-Oncogene Proteins / Q000378:metabolism*; D051057:Proto-Oncogene Proteins c-akt; D051381:Rats; D015398:Signal Transduction; D013482:Superoxide Dismutase / Q000096:biosynthesis / Q000235:genetics*; D000072105:Superoxide Dismutase-1; D015533:Transcriptional Activation
D006861:Hydrogen Peroxide; D016328:NF-kappa B; D019869:Phosphatidylinositol 3-Kinases; D017346:Protein Serine-Threonine Kinases; D011518:Proto-Oncogene Proteins; D051057:Proto-Oncogene Proteins c-akt; D013482:Superoxide Dismutase; D000072105:Superoxide Dismutase-1; C000606290:SOD1 protein, human; C000606292:Sod1 protein, rat; C494918:AKT1 protein, human; C494920:Akt1 protein, rat