Expression of human catalase in acatalasemic murine SV-B2 cells confers protection from oxidative damage.
Reactive oxygen species have been implicated in aerobic organisms as causative agents in damage to DNA, proteins, and lipids. Catalase is a major enzyme in the defense against such oxidant damage. To determine whether increased catalase expression confers greater resistance to oxidant stress, a eukaryotic expression vector harboring a human catalase cDNA clone was constructed. Acatalasemic murine fibroblasts were then co-transfected with that catalase expression vector and pSV2-neo, and successfully transfected cells were identified by their ability to grow in the presence of geneticin. Clones that contained integrated copies of the catalase expression vector were identified by Polymerase Chain Reaction (PCR) analysis. Stably transfected geneticin-resistant cell lines that overexpressed catalase in potentially positive cell lines were confirmed by catalase enzyme assays. To examine the physiological relevance of catalase overexpression, cells were exposed to oxidant stresses (hydrogen peroxide and hyperoxia), and survival rates were determined. Results demonstrated a significant resistance to oxidative stress in cells overexpressing catalase when compared to controls. These transfected cell lines will provide important models for further evaluation of the role of catalase in protecting cells against the toxic effects of oxygen-derived free radicals and their derivatives.
Lindau-Shepard
B A
BA
Wadsworth Center for Laboratories and Research, New York State Department of Health, Albany 12201-0509.
Shaffer
J B
JB
D020642:Acatalasia; D000818:Animals; D002374:Catalase / Q000235:genetics* / Q000378:metabolism*; D002455:Cell Division / Q000187:drug effects; D002460:Cell Line; D002470:Cell Survival / Q000187:drug effects; D015870:Gene Expression; D005822:Genetic Vectors; D006801:Humans; D006861:Hydrogen Peroxide / Q000378:metabolism / Q000633:toxicity; D051379:Mice; D017382:Reactive Oxygen Species / Q000378:metabolism* / Q000633:toxicity; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D014162:Transfection
D017382:Reactive Oxygen Species; D011994:Recombinant Proteins; D006861:Hydrogen Peroxide; D002374:Catalase