Partial purification of estradiol receptor from Xenopus laevis liver and levels of receptor in relation to estradiol concentration.

Abstract

We have used ammonium sulphate precipitation followed by affinity chromatography to partially purify the estrogen receptor from Xenopus laevis liver which may control the genes for vitellogenin, the precursor of the egg yolk proteins. The rate at which receptor binds estradiol explains the kinetics of the induction of vitellogenin synthesis by estradiol, and the dissociation constant (0.5 x 10-9 M) explains the concentration dependence of the response, which has a threshold of 10-9 M estradiol, when 6707o of the receptor is bound to estradiol. The estradiol concentration in male liver, which does not make vitellogenin, is 0.18 x 10-9 M, sufficient to saturate 26%o of the receptor, while in female liver, which makes vitellogenin continuously, the estradiol concentration is 3.5 x 10-9 M, giving 88%o saturation of receptor, suggesting that the proportion of occupied receptor decides whether or not the vitellogenin genes are active. In the physiological concentration range, estradiol modulates the level of receptor, which varies between 100 binding sites per nucleus in males and 440 in females, but artificially high concentrations of estradiol raise the level to -1000 sites per nucleus.This suggests that the small increase in vitellogenin mRNA induced by physiological concentrations of estradiol is due to pre-existing receptor and that the much larger increases induced by very high concentrations depends on newlysynthesized receptor.

Activation of chromosomal vitellogenin genes in Xenopus oocytes by pure estrogen receptor and independent activation of albumin genes.

“…Albumin transcripts are found at about the same level in both female and male liver (Fig. 2), in which the estradiol concentrations are, respectively, 3.5 and 0.18 nM (36). Thus, our findings show that normal concentrations of estradiol do not destabilize albumin mRNA.…”

“…We estimate that for the oocyte vitellogenin B2 gene to be fully activated by pure human receptor, between 108 and 109 molecules have to enter the nucleus. By contrast, when vitellogenin genes are active in frog liver nuclei, only some 102 to 103 molecules of receptor are found in the nucleus (36).…”

Interactions of nitromusk parent compounds and their amino-metabolites with the estrogen receptors of rainbow trout (Oncorhynchus mykiss) and the South African clawed frog (Xenopus laevis)

“…These values are approximately 60-fold greater than the majority of the values reported in the literature (Westley and Knowland, 1978;Wright et al, 1983;Perlman et al, 1984). However, it should be noted that these previously reported values were all obtained in male xenopus of unknown age as well as in primary cell cultures of xenopus hepatocytes.…”

Transient administration of estradiol-17 beta establishes an autoregulatory loop permanently inducing estrogen receptor mRNA.

“…3). The persistent induction of xER mRNA is not due to residual estradiol from the injection, since estrogen levels in serum return to basal levels within 1 day after injection (29). Several years ago, we used hormone-binding studies to demonstrate that estradiol-17,( evoked a long-term increase in high-affinity estrogen binding sites (25).…”

“…A second class of target genes exemplified by RBP and xER exhibits long-term induction by estradiol. The seminal event that allows these genes to exhibit persistent estrogen induction is the establishment of an autoregulatory loop that maintains xER mRNA levels via a combination ofhighlevels ofinduced estrogen receptor protein and residual estrogen circulating even in the serum of male animals (25,29). This additional class of hormone-regulated genes presumably exhibits a lower threshold response to loaded hormone-receptor complex.…”