LIVER-TYPE 11-BETA-HYDROXYSTEROID DEHYDROGENASE CDNA ENCODES REDUCTASE BUT NOT DEHYDROGENASE-ACTIVITY IN INTACT MAMMALIAN COS-7 CELLS

11 beta-Hydroxysteroid dehydrogenase (11 beta-HSD) catalyses the metab olism of corticosterone to inert 11-dehydrocorticosterone, thus preven ting glucocorticoid access to otherwise non-selective renal mineraloco rticoid receptors (MRs), producing aldosterone selectivity in vivo. At least two isoforms of 11 beta-HSD exist. One isoform (11 beta-HSD1) h as been purified from rat liver and an encoding cDNA cloned from a rat liver library. Transfection of rat 11 beta-HSD, cDNA into amphibian c ells with a mineralocorticoid phenotype encodes 11 beta-reductase acti vity (activation of inert 11-dehydrocorticosterone) suggesting that 11 beta-HSD1 does not have the necessary properties to protect renal MRs from exposure to glucocorticoids. This function is likely to reside i n a second 11 beta-HSD isoform. 11 beta-HSD1 is co-localized with gluc ocorticoid receptors (GRs) and may modulate glucocorticoid access to t his receptor type. To examine the predominant direction of 11 beta-HSD 1 activity in intact mammalian cells, and the possible role of 11 beta -HSD in regulating glucocorticoid access to GRs, we transfected rat 11 beta-HSD1 cDNA into a mammalian kidney-derived cell system (COS-7) wh ich has little endogenous 11 beta-HSD activity or mRNA expression. Hom ogenates of COS-7 cells transfected with increasing amounts of 11 beta -HSD cDNA exhibited a dose-related increase in 11 beta-dehydrogenase a ctivity. In contrast, intact cells did not convert corticosterone to 1 1-dehydrocorticosterone over 24 h, but showed a clear dose-related 11 beta-reductase activity, apparent within 4 h of addition of 11-dehydro corticosterone to the medium. To demonstrate that this reflected a cha nge in functional intracellular glucocorticoids, COS-7 cells were co-t ransfected with an expression vector encoding GR and a glucocorticoid- inducible MMTV-LTR luciferase reporter construct, with or without 11 b eta-HSD. Corticosterone induced MMTV-LTR luciferase expression in the presence or absence of 11 beta-HSD. 11-Dehydrocorticosterone was witho ut activity in the absence of 11 beta-HSD, but induced MMTV-LTR lucife rase activity in the presence of 11 beta-HSD. These results indicate t hat rat 11 beta-HSD1 can behave exclusively as a reductase in intact m ammalian cells. Thus in some tissues in vivo, 11 beta-HSD1 may regulat e ligand access to GRs by reactivating inert glucocorticoids.