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Extracellular superoxide dismutase (EC-SOD or SOD3) is an important protective enzyme against the toxicity of superoxide radicals that are produced under both physiological and pathophysiological conditions. We have isolated and characterized over 11 kb of the mouse EC-SOD gene and its 5'- and 3'-flanking regions. The gene consists of two exons, with the entire coding region located within exon 2. In order to study the mechanism of cell-specific gene regulation for mouse EC-SOD, we characterized 2500 bp of its 5'-flanking region using cultured cells derived from mouse lung fibroblasts (MLg), kidney medulla (mIMCD3) and hepatocytes (Hepa 1-6). Real-time PCR showed that basal expression of EC-SOD was considerably higher in MLg cells compared with the other cell types. Reporter-gene assays revealed that the proximal promoter region was sufficient to support this high expression in MLg cells. Although no obvious TATA box was identified, our results show that a highly purine-rich region from -208 to +104 contains active binding sites for both the Kruppel-like and Ets families of transcription factors. Using electrophoretic mobility shift, DNase footprinting and reporter gene assays, we identified myeloid zinc finger 1 and gut-enriched Kruppel-like-factor-like nuclear transcription factors as repressors of EC-SOD expression, whereas nuclear transcription factors from the Ets family, such as Elf-1 and GA-binding protein alpha and beta, were potent activators of EC-SOD transcription. We propose a model that highlights competition between Ets activators and Kruppel-like repressors within the proximal promoter region that determines the level of EC-SOD expression in a particular cell type.

biochemical journal

Myeloid zinc finger (MZF)-like, Kruppel-like and Ets families of transcription factors determine the cell-specific expression of mouse extracellular superoxide dismutase