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

An estrogen receptor beta (ER beta) cDNA fragment was amplified by RT-PCR o f total RNA extracted from liver and ovary of immature channel catfish. Thi s cDNA fragment was used to screen an ovarian cDNA library made from an imm ature female fish. A clone was obtained that contained an open reading fram e 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 2 5 full-length teleost and tetrapod ER sequences. The consensus tree obtaine d indicated the existence of two major vertebrate ER subtypes, alpha and be ta. Within each subtype, and in accordance with established phylogenetic re lationships, teleost and tetrapod ER were monophyletic confirming the resul ts 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 cDN A bound estrogen with high affinity (K-d = 0.21 nM) and specificity. The af finity of channel catfish ER beta for estrogen was higher than previously r eported 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 hea d kidney always corresponded to the pattern in whole blood. In conclusion, we isolated a channel catfish ER beta with ligand-binding affinity and tiss ue expression patterns different from ER alpha. Also, we confirmed the vali dity of our previously proposed general classification scheme for vertebrat e ER into alpha and beta subtypes and within each subtype, into teleost and tetrapod clades. (C) 2000 Academic Press.