Molecular cloning of two isoforms of Xenopus (Silurana) tropicalis estrogen receptor mRNA and their expression during development

Abstract

Native estrogens and estrogenic chemicals exert their actions primarily through the nuclear estrogen receptors (ERs). The present study is directed towards providing a molecular basis for detection of estrogenic and antiestrogenic effects of compounds. In the present study, we isolated successfully two isoforms of a full-length ER mRNA from the liver of Xenopus (Silurana) tropicalis, and analyzed their expression in various tissues during the development by RT-PCR method. The isolated full-length ERα and ERβ cDNAs contained a 1755-bp and a 1644-bp coding regions, respectively. The deduced amino acid sequence of X. tropicalis ERα was highly homologous to previously published X. tropicalis ERα, Xenopus laevis ERs (xer3, ERα1, and ERα2), and X. laevis ER-related proteins. The N-terminal region of the X. tropicalis ERα amino acid sequence cloned in the present study was more highly homologous to that of the xer3 amino acid sequence than the previously published X. tropicalis ERα sequence. Some amino acid residues in the D domain were absent in this sequence. In contrast, the deduced amino acid sequence of the C and E/F domains in X. tropicalis ERβ was highly homologous to those in teleosts, quail, and human ERβs. The RT-PCR analysis showed that expression of the two ER transcripts was relatively abundant in the brain, liver, and gonad/kidney complex of 2-month-old froglet, but weakly expressed in the heart, stomach, leg muscle, and back skin. Gonadal sex differentiation occurred histologically in the X. tropicalis tadpole between Nieuwkoop and Faber stages 54 and 59. The ERα transcripts in the brain, liver and gonad/kidney complex, and ERβ in the brain were expressed at low levels in the tadpole at stage 51, but became extremely abundant in the tadpole at stage 60, and remained at relatively high levels in the froglet after metamorphosis. In contrast, ERβ transcript was expressed abundantly in a consistent manner in the liver and gonad/kidney complex from stage 51 through metamorphosis. Expression of the two ER transcripts was recognized at almost the same levels in both sexes during the development, except for relatively lower expression of ERβ in the female gonad after metamorphosis. These results taken together suggest that the brain, liver, and gonad/kidney complex of the X. tropicalis tadpole may be more susceptible to toxic influences by exogenous estrogenic substances after sex differentiation.