11-hydroxysteroid dehydrogenase type 1 (11-HSD1) increases intracellular glucocorticoid action by converting inactive to active glucocorticoids (cortisol, corticosterone) within cells. It is highly expressed in glucocorticoid target tissues including liver and lung, and at modest levels in adipose tissue and brain. A selective increase in adipose 11-HSD1 expression occurs in obese humans and rodents and is likely to be of pathogenic importance in the Metabolic Syndrome. Here we have used 5' RACE to identify a novel promoter, P1, of the gene encoding 11-HSD1. P1 is located 23kb 5' to the previously described promoter, P2. Both promoters are active in liver, lung, adipose tissue and brain. However, P1 (encoding exon 1A) predominates in lung and P2 (encoding exon 1B) predominates in liver, adipose tissue and brain. Adipose tissue of obese leptin-deficient C57BL/6J-Lep^ob mice showed higher expression only of the P2 associated exon 1B-containing 11-HSD1 mRNA variant. In contrast to P2 which is CAAAT/Enhancer Binding Protein (C/EBP)- inducible in transiently transfected cells, the P1 promoter was unaffected by C/EBP in transfected cells. Consistent with these findings mice lacking C/EBP had normal 11-HSD1 mRNA levels in lung, but showed a dramatic reduction in levels of 11-HSD1 mRNA in liver and brown adipose tissue. These results therefore demonstrate tissue-specific differential regulation of 11-HSD1 mRNA through alternate promoter usage and suggest that increased adipose 11-HSD1 expression in obesity is due to a selective increase in activity of the C/EBP-regulated P2 promoter.