Xia Z, Gale W L, Chang X, Langenau D, Patiño R, Maule A G, Densmore L D
Texas Cooperative Fish & Wildlife Research Unit, Texas Tech University, Lubbock, Texas 79409-2120, USA.
Gen Comp Endocrinol. 2000 Apr;118(1):139-49. doi: 10.1006/gcen.1999.7447.
An estrogen receptor beta (ERbeta) 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 ERbeta 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-7 cells transfected with channel catfish ERbeta cDNA bound estrogen with high affinity (K(d) = 0.21 nM) and specificity. The affinity of channel catfish ERbeta for estrogen was higher than previously reported for channel catfish ERalpha. As determined by qualitative RT-PCR, the tissue distributions of ERalpha and ERbeta were similar but not identical. Both ER subtypes were present in ovary and testis. ERalpha was found in all other tissues examined from juvenile and mature fish of both sexes. ERbeta 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 ERbeta with ligand-binding affinity and tissue expression patterns different from ERalpha. 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.
通过逆转录聚合酶链反应（RT-PCR），从未成熟斑点叉尾鮰的肝脏和卵巢中提取的总RNA扩增出雌激素受体β（ERβ）cDNA片段。该cDNA片段用于筛选从未成熟雌鱼构建的卵巢cDNA文库。获得了一个克隆，其包含一个开放阅读框，编码一个575个氨基酸的蛋白质，推导分子量为63.9 kDa。基于25个硬骨鱼和四足动物的全长ER序列，使用最大简约法和邻接法分析来生成斑点叉尾鮰ERβ的系统发育分类。得到的共识树表明存在两种主要的脊椎动物ER亚型，α和β。在每个亚型内，根据已建立的系统发育关系，硬骨鱼和四足动物的ER是单系的，这证实了先前分析的结果（Z. Xia等人，1999年，《普通比较内分泌学》113卷，360 - 368页）。用斑点叉尾鮰ERβ cDNA转染的COS-7细胞提取物以高亲和力（K(d) = 0.21 nM）和特异性结合雌激素。斑点叉尾鮰ERβ对雌激素的亲和力高于先前报道的斑点叉尾鮰ERα。通过定性RT-PCR确定，ERα和ERβ的组织分布相似但不完全相同。两种ER亚型都存在于卵巢和睾丸中。在来自两性幼鱼和成熟鱼的所有其他检测组织中都发现了ERα。在大多数情况下，除了全血和头肾外，在大多数组织中也发现了ERβ。有趣的是，头肾中ER亚型的表达模式总是与全血中的模式相对应。总之，我们分离出了一种斑点叉尾鮰ERβ，其配体结合亲和力和组织表达模式与ERα不同。此外，我们证实了我们先前提出的脊椎动物ER分为α和β亚型以及在每个亚型内分为硬骨鱼和四足动物进化枝的一般分类方案的有效性。
