11 β-Hydroxysteroid dehydrogenase-2 is a high affinity corticosterone-binding protein

Abstract

In addition to mineralocorticoid and glucocorticoid receptors, a third category of corticosteroid binding sites has been described in the kidney, the Type III binding protein. This intracellular binder has high affinity for corticosterone, but binds neither aldosterone nor synthetic glucocorticoids. Based on similarities in steroid specificity and kinetic parameters, we hypothesized that these corticosterone binding sites belong to the type 2 isoform of 11 β-hydroxysteroid dehydrogenase (11 β-HSD2). The goal of this study was to express the recombinant rabbit 11 β-HSD2 in mammalian cells and test if such cells acquire both NAD-dependent 11 β-HSD2 activity as well as high affinity corticosterone binding sites. Stably transfected CHO cell lines expressed high, NAD-dependent, unidirectional 11 β-HSD2 activity. At the same time, the transfected cells also acquired a large number of corticosterone-specific binding sites (1.21±0.3×10 6), whereas non-transfected cells had no corticosterone binding above background. The K d for corticosterone was 25±8 nM. Neither the glucocorticoid receptor (GR) agonists dexamethasone and RU 28362 nor the mineralocorticoid receptor (MR) agonist aldosterone bound to these sites. The steroid specificity of the binding sites, as determined by competing [ 3H]corticosterone with unlabeled steroids, is identical to that of 11 β-HSD2: corticosterone≫11-hydroxyprogesterone>carbenoxolone>11 dehydrocorticosterone>cortisol>progesterone∼DOC⋙DEX>RU 28362∼aldosterone. These results strongly suggest that the previously described high affinity corticosterone binding sites are 11 β-HSD2. Thus, though Type III binding sites are not corticosteroid receptors as originally thought, they play an important role in regulating the activity of both mineralocorticoid- and glucocorticoid receptors.