Molecular cloning of an estrogen receptor beta subtype from the goldfish, Carassius auratus

The brain of many teleost fish species, including the goldfish Carassius au ratus, expresses exceptionally high levels of cytochrome P450 aromatase (es trogen synthetase). To begin investigating the molecular and cellular targe ts of estrogen action in goldfish brain, a polymerase chain reaction (PCR) cloning strategy was used to isolate an estrogen receptor (ER) complementar y DNA (cDNA). The 2283-bp cDNA isolated from goldfish liver encoded a prote in of 568 amino acids (aa) with an estimated molecular weight of 63,539. Th e goldfish ER had high overall sequence identity when compared to other ver tebrate ER sequences: eel (64%), human beta subtype (59%), human alpha subt ype (46%), medaka (46%), and rainbow trout (47%). The highest degree of con servation was seen in the DNA-binding (94-100%) and ligand-binding (67-79%) domains. Phylogenetic analysis of the ER gene family indicated that the go ldfish and eel ER are most closely related to mammalian ER beta subtypes, w hereas previously identified fish, amphibian, and avian ER forms cluster se parately with mammalian ER alpha subtypes. Using the goldfish ER cDNA (here designated gfER beta), multiple mRNA species (3.1-8.6 kb) were detected by Northern blot analysis in goldfish liver and ovary but expression was belo w detection in brain. Using reverse transcription-PCR analysis, gfERP mRNA was detected in forebrain, mid/hindbrain, pituitary, retina, liver, ovary, and testis. Further studies are required to determine whether an additional ER alpha subtype is present in the goldfish and whether ER alpha or ER bet a forms have evolutionary precedence invertebrates. (C) 1999 Academic Press .