Mutations in aldosterone synthase gene of Milan hypertensive rats: phenotypic consequences.
Using in vitro and in vivo methods, we have demonstrated increased sensitivity of adrenocortical steroidogenesis to ACTH in Milan hypertensive (MHS) compared with normotensive (MNS) rats and have investigated whether this is caused by mutations of steroidogenic enzymes. Genes encoding aldosterone synthase (CYP11B2) and 11beta-hydroxylase (CYP11B1) in MHS and MNS have been cloned and sequenced. Nucleotide 752 (G) in exon 4 of MHS CYP11B2 differs from that of MNS (A); CYP11B1 sequences were identical. The nucleotide 752 mutation caused a Q251R substitution in the amino acid sequence of MHS CYP11B2. The phenotype of MHS CYP11B2 alleles, when expressed in COS-1 cells, differed from that of MNS alleles. The relative activities of the three reactions catalyzed by CYP11B2 (11beta-hydroxylation of deoxycorticosterone, 18-hydroxylation of corticosterone, and dehydrogenation of 18-hydroxycorticosterone) were estimated after incubation of transfected cells with [(14)C]deoxycorticosterone and analysis of radioactivity associated with deoxycorticosterone, corticosterone, 18 hydroxycorticosterone, and aldosterone. Both 11- and 18-hydroxylase activities were lower (19 and 12%, respectively; P < 0.01 and P < 0.05) in cells transfected with MHS compared with MNS alleles, whereas 18-oxidase activity was 42% higher (P < 0.01). To assess the significance of the CYP11B2 mutation in vivo, DNA from F2 hybrid MHS x MNS rats was genotyped. MHS alleles were associated with lower urine volumes in both sexes, lower ventricle weights in male rats, but no difference in systolic or diastolic blood pressures between the sexes. We conclude that a mutation in CYP11B2 may affect aldosterone secretion in MHS; however, under normal environmental circumstances, we were unable to demonstrate any influence of this mutation on blood pressure.
Lloyd-MacGilp
Susan A
SA
Molecular Medicine Centre, Western General Hospital, Edinburgh, EH4 2XU Scotland, UK.
Torielli
Lucia
L
Bechtel
Stephanie
S
Tripodi
Grazia
G
Gomez-Sanchez
Celso E
CE
Zagato
Laura
L
Bernhardt
Rita
R
Kenyon
Christopher J
CJ
D015069:18-Hydroxycorticosterone / Q000378:metabolism; D000302:Adrenal Cortex / Q000187:drug effects / Q000378:metabolism / Q000473:pathology; D000305:Adrenal Cortex Hormones / Q000096:biosynthesis; D000324:Adrenocorticotropic Hormone / Q000097:blood / Q000494:pharmacology; D000450:Aldosterone / Q000096:biosynthesis / Q000097:blood / Q000378:metabolism; D000483:Alleles; D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D001794:Blood Pressure / Q000235:genetics; D019556:COS Cells; D003345:Corticosterone / Q000097:blood / Q000378:metabolism; D019405:Cytochrome P-450 CYP11B2 / Q000235:genetics*; D004247:DNA / Q000032:analysis; D003900:Desoxycorticosterone / Q000378:metabolism; D005838:Genotype; D006900:Hydroxylation; D006973:Hypertension / Q000235:genetics* / Q000378:metabolism / Q000473:pathology; D008126:Lod Score; D008297:Male; D009154:Mutation; D009929:Organ Size; D010641:Phenotype; D051381:Rats; D011922:Rats, Mutant Strains; D012083:Renin / Q000097:blood; D020133:Reverse Transcriptase Polymerase Chain Reaction; D016415:Sequence Alignment; D013252:Steroid 11-beta-Hydroxylase / Q000235:genetics; D014162:Transfection
D000305:Adrenal Cortex Hormones; D003900:Desoxycorticosterone; D000450:Aldosterone; D015069:18-Hydroxycorticosterone; D000324:Adrenocorticotropic Hormone; D004247:DNA; D019405:Cytochrome P-450 CYP11B2; D013252:Steroid 11-beta-Hydroxylase; D012083:Renin; D003345:Corticosterone