Phylogenetic sequence analysis, recombinant expression, and tissue distribution of a channel catfish estrogen receptor β

An estrogen receptor beta (ER beta) cDNA fragment was amplified by RT-PCR of total RNA extracted from liver and ovary of immature channel catfish. This cDNA fragment was used to screen an ovarian cDNA library made from an immature female fish. A clone was obtained that contained an open reading frame encoding a 575-amino-acid protein with a deduced molecular weight of 63.9 kDa. Maximum parsimony and Neighbor Joining analyses were used to generate a phylogenetic classification of channel catfish ER beta on the basis of 25 full-length teleost and tetrapod ER sequences. The consensus tree obtained indicated the existence of two major vertebrate ER subtypes, alpha and beta. Within each subtype, and in accordance with established phylogenetic relationships, teleost and tetrapod ER were monophyletic confirming the results of a previous analysis (Z. Xia et al., 1999, Gen. Comp. Endocrinol. 113, 360-368). Extracts of COS-T cells transfected with channel catfish ERP cDNA bound estrogen with high affinity (K-d = 0.21 nM) and specificity. The affinity of channel catfish ER beta for estrogen was higher than previously reported for channel catfish ER alpha. As determined by qualitative RT-PCR, the tissue distributions of ER alpha and ERP were similar but not identical. Both ER subtypes were present in ovary and testis. ER alpha was found in all other tissues examined from juvenile and mature fish of both sexes. ER beta was also found in most tissues except, in most cases, whole blood and head kidney. Interestingly, the pattern of expression of ER subtypes in head kidney always corresponded to the pattern in whole blood. In conclusion, we isolated a channel catfish ER beta with ligand-binding affinity and tissue expression patterns different from ER alpha. Also, we confirmed the validity of our previously proposed general classification scheme for vertebrate ER into alpha and beta subtypes and within each subtype, into teleost and tetrapod clades. (C) 2000 Academic Press.