Molecular cloning and characterization of brain and ovarian cytochrome P450 aromatase genes in the catfish Heteropneustes fossilis: Sex, tissue and seasonal variation in, and effects of gonadotropin on gene expression

Abstract

Cytochrome P450 aromatase (Cyp19arom) is the rate-limiting enzyme controlling estrogen biosynthesis, coded by Cyp19a1 in most gnathostomes. Most teleosts have two forms expressed differentially in ovary (cyp19a1a) and neural tissue (cyp19a1b). In this study, full length cDNAs of 2006bp and 1913bp with ORFs of 1575bp and 1488bp were isolated from the brain and ovary, respectively, of the catfish Heteropneustes fossilis, an air-breathing species with high aquaculture potential. The ORFs encode predicted proteins of 495 and 524 amino acid residues, respectively. The proteins show 62% identity with each other and cluster in two distinct clades (the brain type and ovary type) in the teleost taxon, separated from the tetrapod type. In the in situ localization study, both cyp19a1a and cyp19a1b transcripts were localized in the brain but the signal intensity was higher for the brain type paralog. The transcript signals were observed in the radial glial cells and in neuronal populations of the dorso-lateral region of the telencephalon, pre-tectum, hypothalamus and medulla oblongata. In the ovary, both paralogs were expressed in the follicular layer with a high signal intensity of the ovarian type (cyp19a1a). The differential expression of the gene paralogs was evident from qPCR analysis. Cyp19a1b has relatively a high abundance in the female brain, followed by other peripheral tissues (gonads, liver, gill, kidney and muscle). On the other hand, cyp19a1a has relatively a high transcript abundance in the ovary and female brain, followed by the testis and male brain, and female liver and muscle. The expression was low in male liver and muscle, and the lowest in the gill and kidney. The expression of the two paralogs exhibit brain regional differences; both types have relatively a high transcript abundance in telencephalon-preoptic area with the cyp19a1b expression higher in females than males. In hypothalamus, the expression of both types is higher in males than females. In medulla, the expression of the cyp19a1b is higher than cyp19a1a, and the transcript abundance of the ovarian type is higher in females than males. The expression of the gene paralogs elicits significant seasonal variations in the ovary and brain. In both tissues, the expression increases from the resting to preparatory phases, and decreases through the prespawning phase to low levels in spawning phase. In vivo and/or in vitro treatments with human chorionic gonadotropin (hCG) stimulated the expression of the gene paralogs in the brain and ovary, time-dependently. In conclusion, both paralogs have an overlapping distribution at different levels of the brain–pituitary–gonad axis and may function as a single functional unit as far as the estrogen synthesis is concerned.