The Binding of 5α-Pregnane-3,20-dione by Cytosol and Nuclear Preparations of Guinea Pig Uterus*

Abstract

We compared the binding characteristics of [3H]progesterone and 5 alpha-[3H]pregnanedione in cytosol and nuclear preparations of ovariectomized, estrogen-treated guinea pig uterus. There were similarities as well as differences in binding behavior. [3H]Progesterone and 5 alpha-[3H]pregnanedione were bound in cytosol with approximately the same apparent association constants and concentrations of binding sites. When centrifuged on sucrose gradients in 5 mM phosphate buffer, binding peaks with sedimentation coefficients of 7S were found with both. 5 alpha-[3H]Pregnanedione was bound by uterine nuclei and, like [3H]progesterone, required temperature-activated cytosol of estrogen-stimulated uterus. A series of unlabeled steroids had similar relative abilities to displace both [3H]progesterone and 5 alpha-[3H]pregnanedione from receptor complexes in cytosol or nuclei. When cytosol was incubated with increasing concentrations of [3H]progesterone or 5 alpha-[3H]pregnanedione the amount of 5 alpha-[3H]pregnanedione bound exceeded the amount of [3H]progesterone bound at steroid concentrations above 5 x 10(-9) M. This suggested that 5 alpha-pregnanedione was bound by additional components of the cytosol and the suggestion was strengthened by sucrose gradient analysis. At greater than saturating-steroid concentrations, the partition between 7S and 4 S binding proteins was different. [3H]Progesterone bound to 7S binding proteins in preference to 4S binders, whereas 5 alpha-pregnanedione showed relatively more 4S binding.

The binding of rat uterine cytosol oestrogen receptors to oligodeoxythymidylate-cellulose. Its relationship to a stable form of receptor complex with separate ligand- and oligonucleotide-binding sites

“…Using oligo(dT)-cellulose binding of receptor complexes as an index, we find that dilution, dialysis or (NH4)2SO4 precipitation increased 'activation' of receptor, in agreement with other workers studying the oestrogen receptor (Sato et al, 1978(Sato et al, , 1979) and other steroid receptors (Bailly et al, 1977;Goidl et al, 1977;Saffran et al, 1976). The increased binding to oligo(dT)-cellulose observed after these treatments may be the result of removal of lowmolecular-weight components which inhibit either the interaction of the two subunits necessary to form 5S receptor or the interaction of 5 S receptor with oligo(dT)-cellulose.…”

Activation, Transformation, and Subunit Structure of Steroid Hormone Receptors

“…Fleischmann and Beato (70) reported that both the transformed and untransformed forms of this receptor sediment at about 4S, whereas in the study of Saffran et al (71), there was a shift from 7S to 5.5S during transformation. This held true as well for the receptor of guinea pig uterus (71).…”

Inhibitory Effect of 5-Alpha-Dihydroprogesterone on Nuclear Estrogen Receptor Binding of the Anterior Pituitary and Uterus in the Rat

“…5 a -D H P probably acts by a similar receptor-mediated mechanism, but it is unclear at this moment if this action is mediated by its own receptor or by the progester one receptor system. It has been shown that 5 a -D H P binds al most as well as progesterone to the progesterone receptor in the pituitary [29] and hypothalamus o f the rat [30], oviduct o f estro gen-primed chicks [31 ] and guinea pig uterus [32]. The involve ment o f the progesterone receptor system in 5 a -D H P action was addressed in experiments with the antiprogestin RU486 that showed antagonism o f the 5a-DHP-induced decrease in prolactin release in estrogen-primed immature rats [Brann et al, unpubl.…”