Is the renal type III corticosteroid-binding site the collecting duct-specific isoform of 11 beta-hydroxysteroid dehydrogenase?

Abstract

In addition to mineralocorticoid and glucocorticoid receptors, the kidney contains a third high affinity binding site for endogenous glucocorticoids, the type III receptor. These binding sites have been localized to the collecting duct, but their biological function has not yet been identified. We have used immunodissected rabbit cortical collecting duct (CCD) cells to further characterize these binding sites. Experiments using intact cells revealed very high levels of [3H]corticosterone (CS) binding (2.99 +/- 0.38 x 10(6) sites/cell), about 100-fold higher than the number of mineralocorticoid or glucocorticoid receptors in CCD cells. Among the two cell types of the collecting duct, principal cells, the putative targets of aldosterone, contain approximately 10 times more CS-binding sites than intercalated cells. The Kd of the binding sites for CS averaged 54.3 +/- 3.48 nM at 0 C. The relative affinity of unlabeled steroids for the binding sites is CS > carbenoxolone congruent to glycyrrhetinic acid > or = 11-dehydrocorticosterone (11-DHCS) > cortisol congruent to cortisone > deoxycorticosterone > progesterone. Synthetic glucocorticoids (dexamethasone, RU 28362, and RU 486) and aldosterone did not compete for [3H]CS binding. Based on their preferential localization to CCD cells, which are the main targets of aldosterone, we hypothesized that these CS-binding sites are involved in conferring aldosterone specificity on mineralocorticoid receptors in these cells. As 11 beta-hydroxysteroid dehydrogenase (11-OHSD) is thought to play a key role in this process, we studied the relationship between CS-binding sites and the collecting duct isoform of this enzyme (11-OHSD/CD) in purified CCD cells. Freshly isolated cells rapidly converted [3H]CS to 11-DHCS with an apparent Km of about 50 nM, a value close to the Kd of the CS-binding sites for CS in these cells. The rank order of potency of unlabeled steroids to decrease the conversion of [3H]CS to [3H]11-DHCS was identical to their relative affinity for the CS-binding sites. Also, there is a close correlation (r = 0.783; P < 0.0001) between the activity of 11-OHSD and the number of CS-binding sites in different CCD preparations. Based on the similarities between the abundant CS-binding sites and avid CS metabolism in CCD cells, we suggest that these binding sites belong to the collecting duct isoform of 11-OHSD, which, by decreasing the intracellular levels of active glucocorticoids, plays an important role in conferring aldosterone selectivity on mineralocorticoid receptors in CCD cells.