A Novel Nonstop Mutation in the Stop Codon and a Novel Missense Mutation in the Type II 3β-Hydroxysteroid Dehydrogenase (3β-HSD) Gene Causing, Respectively, Nonclassic and Classic 3β-HSD Deficiency Congenital Adrenal Hyperplasia

Abstract

We investigated two novel point mutations in the human type II 3beta-hydroxysteroid dehydrogenase (3beta-HSD) gene causing a mild and a severe form of 3beta-HSD deficiency congenital adrenal hyperplasia. The first is a nonstop mutation in the normal stop codon 373 of the gene in exon IV [TGA (Stop) --> TGC (Cys) = Stop373C) identified from one allele of a female child with premature pubarche whose second allele had an E142K mutation. The Stop373C mutation predictably results in an open reading frame and a mutant-type (MT) II 3beta-HSD protein containing 467 amino acid residues, compared with the 372 amino acid residues of wild-type (WT) protein. The second is a homozygous missense mutation in codon 222 [CCA (Pro) --> ACT (Thr) = P222T] in the gene identified from a female neonate with salt-wasting disorder. The pcDNA vectors containing the constructs of WT II 3beta-HSD cDNA, WT cDNA with the open reading frame (WT cDNA(+)), MT Stop373C with the open reading frame (Stop373C(+)) and MT P222T cDNA were transfected in COS-I and 293T cells and expressed a similar amount of 3beta-HSD mRNA. The enzyme activity in intact cells using pregnenolone and dehydroepiandrosterone as substrate in the medium (1 micromol/liter) was identical between the WT cDNA and the WT cDNA(+), but was decreased to 27% of the WT enzymes at 6 h by MT Stop373C(+) enzyme, and was undetectable by P222T enzyme. In the homogenates of the cells, both MT Stop373C(+) and P222T enzyme activities and enzymes were undetectable despite clear detection of WT enzyme activities and WT enzymes. LH response to an LHRH analog stimulation in the pubertal female with the Stop373C/E142K genotypes and in a pubertal female with compound 273/318 frameshift genotypes were comparable to and higher than control females, respectively. In conclusion, a structurally lengthy MT II 3beta-HSD enzyme due to a nonstop mutation was relatively detrimental in intact cells causing the nonclassic phenotype of 3beta-HSD deficiency. A missense P222T mutation was seriously detrimental, causing the classic phenotype of 3beta-HSD deficiency. The undetectable Stop373C and P222T enzymes on Western blottings, together with the respective in vivo and in vitro data, suggest that a relative instability of Stop373C enzyme and a profound instability of the P222T enzyme are likely the detrimental molecular mechanisms. The increased LH in the female with the frameshift genotype and the appropriate LH response in the female with the nonstop genotype correlated with predictably severe and mild ovarian type II 3beta-HSD deficiency, respectively.

Human 3β-hydroxysteroid dehydrogenase deficiency seems to affect fertility but may not harbor a tumor risk: lesson from an experiment of nature

“…Treatment resulted in testicular growth and pubertal advancement, but effect on gynecomastia was not further described. In HSD3B2 deficiency, development of secondary sexual characteristics and marked gynecomastia at puberty has been described in 46,XY DSD individuals and seems not to correlate with the severity of enzyme deficiency (Table 3) (2,5,6,7,8,9,10,11,12,13,14,19,20,22). Gynecomastia may rather correspond to peripheral precursor conversion to estrogens which depends on accuracy of replacement therapy and peripheral HSD3B1 activity.…”

“…Case Report M-A Burckhardt and others Histology of HSD3B2 deficiency 173:5 K6 described pubertal development (Table 3) (2,5,6,7,8,9,10,11,12,13,14,19,20). Remarkably, the adrenal phenotype was mostly more severe than the DSD phenotype, and all subjects spontaneously virilized at puberty (when not gonadectomized).…”

3?-hydroxysteroid dehydrogenase type II deficiency on newborn screening test

“…A different mutation was also described in the same position at the protein, the p.P222T, and was associated with premature pubarche, but not salt-wasting form. Notably, the mutated protein 222T was not detected on in vitro assays with transiently transfected 293 cells, bringing up an additional pathogenic mechanism of a severe impairment in protein stability (32).…”

A New Homozygous Frameshift Mutation in the <b><i>HSD3B2</i></b> Gene in an Apparently Nonconsanguineous Italian Family

“…At birth, she was misdiagnosed as having 21-hydroxylase deficiency CAH, based on elevated 17-OHP and testosterone levels, and the correct diagnosis was carried out at 7 years of age. A nonstop mutation in the stop codon 373 (Stop373C), was identified in a female child with late-onset CAH [25]: the Stop373C leads to an open reading frame of 95 additional amino acid residues, resulting in a mutant enzyme with 467 amino acid residues compared to the 372 amino acid residues of the normal protein. The authors suggest that this extra tail of the enzyme produces an unstable protein, so that the protein is made but quickly falls apart.…”