THE PRESENCE OF 2 CYTOCHROME-P450 ALDOSTERONE SYNTHASE MESSENGER-RNAS IN THE HAMSTER ADRENAL

We isolated a cDNA from a hamster adrenal cDNA library which was similar in sequence to those of the mouse and rat P450(c18) cDNAs. The hamster P450(c18) cDNA, however, was shorter than the rat and mouse P450(c18) cDNAs at its 5'-end and the peptide leader sequence was absent. From a hamster genomic library we isolated and sequenced the first seven exons and a 5'-flanking region of the first P450(c18) gene exon. With this information we were able to generate a P450(c18) cDNA containing the peptide leader sequence using the polymerase chain reaction. Northern analyses were performed on adrenals from hamsters maintained on a low sodium diet for 0, 4, 7 and 10 days using a P-32-labeled sequence specific to P450(c18); two mRNA bands were found at 2 and 3.4 kb. The intensity of both bands was increased about 3- to 5-fold under sodium restriction compared to controls. A distinct mRNA band of 2.3 kb hybridized with an oligonucleotide specific to P450(11 beta) and its intensity did not change following low sodium intake. Immunoblotting analyses were performed using an antibovine adrenal P450(11 beta) antibody that does not discriminate between P450(11 beta) and P450(c18) proteins. Three bands were detected at 52, 48 and 45 kDa in homogenate preparations of entire glands. Furthermore, the 45 kDa protein band was present in homogenates of the zona glomerulosa and absent in homogenates of the zone fasciculata-reticularis. In conclusion, these results show that the hamster adrenals express P450(c18) as do mouse, rat and human adrenal glands. Furthermore, two P450(c18) mRNAs, which are inducible by a low sodium intake, are present in the hamster adrenal vs one for the rat. The physiological role of these two hamster adrenal mRNA species remains to be elucidated.