Evidence that steroid 5alpha-reductase isozyme genes are differentially methylated in human lymphocytes.
The synthesis of dihydrotestosterone (DHT) is catalyzed by steroid 5alpha-reductase isozymes 1 and 2, and this function determines the development of the male phenotype during embriogenesis and the growth of androgen sensitive tissues during puberty. The aim of this study was to determine the cytosine methylation status of 5alpha-reductase isozymes types 1 and 2 genes in normal and in 5alpha-reductase deficient men. Genomic DNA was obtained from lymphocytes of both normal subjects and patients with primary 5alpha-reductase deficiency due to point mutations in 5alpha-reductase 2 gene. Southern blot analysis of 5alpha-reductase types 1 and 2 genes from DNA samples digested with HpaII presented a different cytosine methylation pattern compared to that observed with its isoschizomer MspI, indicating that both genes are methylated in CCGG sequences. The analysis of 5alpha-reductase 1 gene from DNA samples digested with Sau3AI and its isoschizomer MboI which recognize methylation in GATC sequences showed an identical methylation pattern. In contrast, 5alpha-reductase 2 gene digested with Sau3AI presented a different methylation pattern to that of the samples digested with MboI, indicating that steroid 5alpha-reductase 2 gene possess methylated cytosines in GATC sequences. Analysis of exon 4 of 5alpha-reductase 2 gene after metabisulfite PCR showed that normal and deficient subjects present a different methylation pattern, being more methylated in patients with 5alpha-reductase 2 mutated gene. The overall results suggest that 5alpha-reductase genes 1 and 2 are differentially methylated in lymphocytes from normal and 5alpha-reductase deficient patients. Moreover, the extensive cytosine methylation pattern observed in exon 4 of 5alpha-reductase 2 gene in deficient patients, points out to an increased rate of mutations in this gene.
Rodríguez-Dorantes
M
M
Facultad de Química, UNAM, Ciudad Universitaria, Coyoacán 04510, Mexico.
Lizano-Soberón
M
M
Camacho-Arroyo
I
I
Calzada-León
R
R
Morimoto
S
S
Téllez-Ascencio
N
N
Cerbón
M A
MA
D013741:3-Oxo-5-alpha-Steroid 4-Dehydrogenase / Q000235:genetics*; D001483:Base Sequence; D016022:Case-Control Studies; D019175:DNA Methylation; D017931:DNA Primers; D006801:Humans; D007527:Isoenzymes / Q000235:genetics*; D008214:Lymphocytes / Q000201:enzymology*; D008297:Male; D016133:Polymerase Chain Reaction
D017931:DNA Primers; D007527:Isoenzymes; D013741:3-Oxo-5-alpha-Steroid 4-Dehydrogenase