Estrogen and antiestrogen binding to rat uterine and pituitary estrogen receptor: evidence for at least two physicochemical forms of the estrogen receptor.

Our laboratory has previously reported that calf uterine cytosol prepared in buffer containing 10 mM molybdate and chromatographed on DEAE-Sephadex contains two forms of the unactivated estrogen receptor, Peak I and Peak II; however, cytosol receptor bound to the high-affinity antiestrogen, H1285 (4-(N,N-diethylaminoethoxy)-4'-methoxy-alpha- (p-hydroxyphenyl)-alpha'-ethylstilbene), eluted only as Peak I. We have extended these studies to the rat uterus and pituitary in order to determine the organ and species specificity of this phenomenon. Cytosol prepared in Tris-molybdate buffer from immature and adult rat uteri or pituitaries was labelled with 10 nM [3H]estradiol or [3H]H1285 and chromatographed on QAE-Sephadex. Uterine estrogen receptors bound to either [3H]estradiol or [3H]H1285 eluted from QAE-Sephadex as a large Peak I (approximately 0.21 M KCl) and a smaller Peak II (approximately 0.25 M KCl). Analyses of these partially purified estrogen receptor fractions using high-salt sucrose density gradients showed that Peak I [3H]estradiol-receptor complexes sedimented predominantly as a lighter form (4.0S). In contrast, Peak I [3H]H1285-receptor complexes sedimented primarily as a heavier form (5.5S) often accompanied by a smaller lighter form (4.0S). Peak II [3H]estradiol- and [3H]H1285-receptor complexes sedimented as the heavier form (5.3-5.5S). These data suggest a monomer-dimer relationship between estrogen receptor forms with antiestrogen binding favoring the formation of the dimeric form. Further analysis of these Peak I receptor complexes by gel filtration chromatography yielded molecular forms of approx 70 KDaltons for [3H]estradiol-receptor complexes and 73 KDaltons and 165 KDaltons for [3H]H1285-receptor complexes, supporting the monomer-dimer concept. Data from experiments with the pituitary also suggest that H1285 causes the formation of the dimeric receptor form whereas estradiol interaction with the receptor results only in the monomeric form. These differences in estrogen receptor forms when bound by estrogen versus antiestrogen may be related to the different biological responses induced by these ligands.