ISOLATION OF A GOLDFISH BRAIN CYTOCHROME-P450 AROMATASE CDNA - MESSENGER-RNA EXPRESSION DURING THE SEASONAL CYCLE AND AFTER STEROID TREATMENT

To investigate the molecular basis and physiological regulation of exc eptionally high levels of aromatase (P450arom) activity in the brain o f teleost fish, a 2927 bp P450arom cDNA encoding a 510 amino acid prot ein was isolated from a goldfish brain cDNA library. The brain-derived cDNA had 53% and 61-62% sequence identity when compared with human pl acental and fish ovarian P450arom forms, respectively, and higher homo logies in conserved functional domains. Goldfish brain poly(A) RNA was translatable in vitro to a 56 kDa P450arom immunoprecipitation produc t. Northern blot analysis using the brain cDNA revealed a major 3.0 kb transcript of high abundance in brain (FB, forebrain > M/HB, mid/hind brain), but no signal in ovary, testis or liver. P450arom mRNA varied seasonally in brain, with a peak at the onset of gonadal regrowth (Feb ruary) that preceded the annual rise in enzyme levels and was il 4-fol d (FB) or 50-fold (M/HB) higher than during reproductive inactivity (J uly-December). Known markers of neurogenesis and estrogen action in br ain (28S rRNA, beta-actin and beta-tubulin transcripts) each had uniqu e seasonal patterns which differed from P450arom mRNA. In vivo steroid treatment showed that estrogen and aromatizable androgen increase FB and M/HB levels of P450arom mRNA 8- and 4-fold, respectively. P450arom mRNA in pituitary and retina had a different regulation. Southern ana lysis provided no evidence for multiple genes encoding the brain deriv ed cDNA or for brain-specific gene amplification. Results imply that h igh accumulated levels of P450arom mRNA are the major determinant of h igh measured enzyme activity in goldfish brain, and that physiological regulation of mRNA expression in the natural environment is mediated by aromatization of androgen to estrogen. (C) 1998 Elsevier Science Ir eland Ltd. All rights reserved.