A Study on the Expression of Estrogen-Responsive Genes in Rainbow Trout, Salmo gairdneri

Abstract

The expression of the estradiol-responsive genes in the liver of rainbow trout, Salmo gairdnerii, has been studied as a system which may permit quantitative measure of adverse sublethal effects of various xenobiotics. Upon administration of 17 β-estradiol to male or immature female rainbow trout, the expression of two genes are markedly enhanced. One of these genes codes for the egg yolk precursor protein, vitellogenin, whereas the other codes for another, as yet unidentified, protein. Complementary DNA (cDNA) genes of these two proteins have been employed as probes to characterize the dose-response relationship as well as the time course induction of these two classes of transcripts in liver both in vivo and in vitro. The maximal net transcription of pRTC 2 occurs at 20 ug β -estradiol per 100 gram fish body weight whereas those encoded for by pRTC 5 display maximal transcription at 5 μg β-estradiol per 100 gram fish body weight. With doses exceeding the dose required for maximal induction of transcripts, both classes of transcripts reach their maximal levels between 4 and 8 hours after primary and secondary induction. Transcripts homologous to pRTC 5 return to control levels by 16 hours and 2 days after primary and secondary induction respectively. Those homologous to pRTC 2 however, were not observed to recover to normal levels in the primary induction even 16 days after stimulation with estradiol. However, upon secondary induction pRTC 2 transcripts returned to control levels 8 days post-inoculation with estradiol. The effect of various xenobiotics on the induction of these two estradiol-responsive genes was investigated. Pretreatment with β-naphthoflavone resulted in neither the induction or repression of both classes of transcripts. Administration of Kepone prior to a subsequent stimulation with estradiol, resulted in a decrease in the accumulation of pRTC 2 transcripts only. In contrast, an isomer of DDT, p,p'-DDT, was found to enhance the expression of pRTC 5 transcripts in fish subsequently administered β-estradiol. Oncogenes have been found to be activated in transformed cells or cells treated with various xenobiotics. In this study, I have shown that the cellular oncogene, pRTC-myc 1-81, is expressed in the liver at an elevated level in response to estradiol treatment. The two classes of polyadenylated transcripts were detected (2.5 kb and 5.6 kb). The three estradiol-responsive genes mentioned above (pRTC 2, pRTC 5, and pRTC-myc 1-81) were also shown to be transcribed in an established rainbow trout hepatoma cell line. Transcriptional activities of all three genes, as well as the translational activity of vitellogenin, were diminished upon growth in serum stripped of endogenous hormones. Further characterization of this rainbow trout hepatoma cell line is warranted so that it may be employed in a sensitive and quantitative bioassay for assessing the pathobiochemical effects of environmental xenobiotics on fish reproduction. Such an index may be beneficial for establishing adequate guidelines for water contaminants that potentially could affect aquatic and human reproductive success.