title: Heterologous expression of selenium-dependent glutathione peroxidase affords cellular resistance to paraquat.
abstract: Transfection of murine NIH3T3 fibroblasts and human MCF7 breast carcinoma cells with a pSV2-derived eukaryotic expression vector for human cytosolic glutathione peroxidase resulted in clones with increased glutathione peroxidase activity. This heterologous expression indicates that murine cells recognize the human "selenocysteine insertion sequence" in the 3' untranslated region of the mRNA which facilitates insertion of selenocysteine directed by the opal codon. Though most clones from both cell lines eventually lost their enhanced glutathione peroxidase activity despite continuous selection on G418, some NIH3T3 clones retained enhanced enzyme activity without continuous G418 exposure. Transfection of MCF7 cells with an Epstein-Barr virus (EBV)-derived episomally replicating expression vector carrying the glutathione peroxidase gene also revealed increased glutathione peroxidase activity. These MCF7 cells, however, all required exposure to G418 to maintain enhanced glutathione peroxidase activity. Detailed biochemical analysis of a stably expressing NIH3T3 clone and MCF7 expressing cells revealed no alterations in activities of copper-zinc superoxide dismutase, manganese superoxide dismutase, catalase, phospholipid-glutathione peroxidase, glutathione reductase, glutathione transferase, or NADPH-P450 reductase. Both pSV2- and EBV-derived glutathione peroxidase-expressing clones exhibited enhanced resistance to paraquat as well as to peroxides.
mesh_terms: D016475:3T3 Cells; D000818:Animals; D002470:Cell Survival / Q000187:drug effects; D004351:Drug Resistance; D005822:Genetic Vectors; D005979:Glutathione Peroxidase / Q000096:biosynthesis* / Q000235:genetics; D004854:Herpesvirus 4, Human; D006801:Humans; D051379:Mice; D010269:Paraquat / Q000633:toxicity*; D012643:Selenium; D014162:Transfection; D014407:Tumor Cells, Cultured
chemical_list: D005979:Glutathione Peroxidase; D012643:Selenium; D010269:Paraquat