MOLECULAR-BASIS OF HUMAN 3-BETA-HYDROXYSTEROID DEHYDROGENASE-DEFICIENCY

The enzyme 3 beta-hydroxysteroid dehydrogenase (3 beta-HSD) catalyses an essential step in the biosynthesis of all classes of steroid hormon es. Classical 3 beta-HSD deficiency is responsible for CAHII, a severe form of congenital adrenal hyperplasia (CAH) that impairs steroidogen esis in both the adrenals and gonads. Newborns affected by 3 beta-HSD deficiency exhibit signs and symptoms of adrenal insufficiency of vary ing degrees associated with pseudohermaphroditism in males, whereas fe males exhibit normal sexual differentiation or mild virilization. Elev ated ratios of 5-ene-to 4-ene-steroids appear as the best biological p arameter for the diagnosis of 3 beta-HSD deficiency. The nonclassical form has been suggested to be related to an allelic variant of the cla ssical form of 3 beta-HSD as described for steroid 21-hydroxylase defi ciency. To elucidate the molecular basis of the classical form of 3 be ta-HSD deficiency, we have analysed the structure of the highly homolo gous type I and II 3 beta-HSD genes in 12 male pseudohermaphrodite 3 b eta-HSD deficient patients as well as in four female patients. The 14 different point mutations characterized were all detected in the type II 3 beta-HSD gene, which is the gene predominantly expressed in the a drenals and gonads, while no mutation was detected in the type I 3 bet a-HSD gene predominantly expressed in the placenta and peripheral tiss ues. The finding of a normal type I 3 beta-HSD gene provides the expla nation for the intact peripheral intracrine steroidogenesis in these p atients and increased androgen manifestations at puberty. The influenc e of the detected mutations on enzymatic activity was assessed by in v itro expression analysis of mutant enzymes generated by site-directed mutagenesis in COS-1 cells. The mutant type II 3 beta-HSD enzymes carr ying mutations detected in patients affected by the salt-losing form e xhibit no detectable activity in intact tranfected cells, whereas thos e with mutations found in nonsalt-loser index cases have some residual activity ranging from similar to 1-10% compared to the wild-type enzy me. Although in general, our findings provide a molecular explanation for the enzymatic heterogeneity ranging from the severe salt-losing fo rm to the clinically inapparent salt-wasting form of the disease, we h ave observed that the mutant L108W or P186L enzymes found in a compoun d heterozygote male presenting the salt-wasting form of the disease, h as some residual activity (similar to 1%) similar to that observed for the mutant N100S enzyme detected in an homozygous male patient suffer ing from a nonsalt-losing form of this disorder. Unlike the classical 3 beta-HSD deficiency, our study in women presenting nonclassical 3 be ta-HSD deficiency strongly suggests that this disorder is not due to a mutant type II 3 beta-HSD.