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We have analyzed the nucleotide sequence of complementary and genomic DNAs of the human androgen receptor (AR) gene in two siblings (patients 9006 and 9030) with receptor-positive complete androgen insensitivity (Rec(+)-CAI). Northern analysis indicated that mRNA of the AR was normal in size. However, its expression was relatively reduced in both patients. Consistent with the normal androgen-binding capacity (496 and 552 fmol/mg DNA for patients 9006 and 9030, respectively) but decreased DNA-binding ability (168 fmol/mg DNA) measured in genital skin fibroblasts, no mutation was found in both N-terminal and ligand-binding domains of the AR. However, a single base substitution (G-->A) was found in the second zinc finger of the DNA-binding domain at nucleotide 2372 of the AR cDNA in both cases. This resulted in the replacement of a highly conserved arginine residue (amino acid 614) by a histidine. When the mutated receptor plasmid was cotransfected into PC-3 cells together with the reporter chloramphenicol acetyltransferase gene, chloramphenicol acetyltransferase activity was not induced by 5 alpha-dihydrotestosterone treatment, confirming that the mutation renders the AR nonfunctional and can, therefore, be held responsible for the clinical features in these patients. These results highlight the importance of Arginine-614 in the second zinc finger of the DNA-binding domain of the AR in the protein-DNA interaction.

Dihydrotestosterone (DHT), the 5 alpha-reduced metabolite of testosterone, is the active molecule triggering androgen action, and 5 alpha-reductase (5 alpha-R), the enzyme converting testosterone to DHT, is a key step in this mechanism. Skin, like prostate, is a DHT- dependent tissue. Our laboratory demonstrated, many years ago, that 5 alpha-R in external genitalia was not regulated by androgens, whereas it was androgen dependent in public skin. As two genes, 5 alpha-R types 1 and 2, encoding for 5 alpha-R enzymes have been recently cloned, we undertook the present study to determine whether the two enzymes we had postulated on the basis of regulation studies were coincident with the cloned isoforms. The expression of the two isoforms was studied in genital and pubic skin fibroblasts from normal men, normal women, and hirsute patients. Messenger ribonucleic acid analysis, using Northern blot and RT-PCR techniques, indicated that both 5 alpha-R1 and -2 messenger ribonucleic acids are expressed in genital skin as well as in public skin fibroblasts. In contrast, studies using specific inhibitors of 5 alpha-R1 (LY306089) and 5 alpha-R2 (finasteride) showed that 5 alpha-R2 is predominant in pubic skin of normal men, normal women, and hirsute patients. These data raise the question of the possible use of specific 5 alpha-R1 inhibitors in the treatment of idiopathic hirsutism.

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