Multiple forms of oviduct progesterone receptors analyzed by ion exchange filtration and gel electrophoresis

Sherman, Merry R.; Tuazon, Fe B.; Diaz, Soledad C.; Miller, Leslie

doi:10.1021/bi00650a006

Cited by 99 publications

(30 citation statements)

References 29 publications

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“…The molecular weight of the non-aggregated form determined from the stokes radius and the sedimentation coefficient is much lower (80000) indicating a high asymmetry of the receptor molecule V/fo = 1.40). A similar asymmetry has been found for the estrogen receptor from calf uterus [28] and rat uterus [29] as well as for the progesterone receptor from chicken oviduct [15]. The question now arises, why does the binding protein from crude nuclei show neither aggregation nor the higher molecular weight form.…”

Section: Discussionsupporting

confidence: 66%

“…The asymmetry of the receptor molecule, however, is lost, indicating the cleavage of an asymmetric portion of the molecule from a globular estrogen-binding entity. These properties of the receptor fragment obtained after trypsination are very similar to those of the hero-receptor', a small form of the progesterone receptor in chicken oviduct obtained by treatment of the receptor with certain divalent cations [15,27]. Treatment of chromatin with trypsin also yields receptor fragments.…”

Section: Discussionmentioning

confidence: 62%

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Estrogen-Binding Sites of Chicken Liver. Preliminary Characterization of Nuclear Components

Gschwendt

Schneider

1978

Eur J Biochem

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1. Liver nuclei from estrogenized chickens contain high-affinity low-capacity estrogen-binding sites, which are in part salt-extractable (60 -70 %) and in part tightly sticking to the nuclear residue During the preparation of chromatin with low salt buffers part of the salt-extractable nuclearbinding sites remains together with the non-extractable sites on the chromatin. Extractable and non-extractable sites can be separated by agarose as well as hydroxyapatite chromatography.2. The estrogen-binding protein extracted from crude nuclei was characterized as follows : sedimentation coefficient of 3.9 S, Stokes' radius (a) of 3.3 nm, molecular weight ( M , ) of 56000 and frictional ratio (f/b) of 1.20. Trypsination results in a globular receptor fragment ( M , = 41 000), which has lost a small asymmetric portion of the receptor molecule but still binds estradiol.3. In contrast to the binding protein from crude nuclei the estrogen-binding protein extracted from purified nuclei at pH 7.4 is found mainly in an aggregated form. Dissociation of the aggregates is achieved in high saltlurea resulting in a receptor molecule with an apparent molecular weight of 130000. Aggregation of the binding sites can be prevented to some extent by raising the pH of the extraction medium. At pH 8.7 the non-aggregated part of the binding protein from pure nuclei could be characterized as follows: 4.4 S, a = 4.3 nm, M , = 80000 (apparent molecular weight of 4. Mixing experiments indicate that an extranuclear protease present in a crude nuclear preparation converts the large receptor (the binding protein from pure nuclei) to the smaller one (the binding protein from crude nuclei) by digesting off an asymmetric portion of the molecule. This portion seems to be responsible for the strong tendency of the binding protein from pure nuclei to associate with other nuclear components.(30-40%). 1500OO),fl'~ = 1.40.The chicken liver as site of the estrogen-induced egg yolk protein synthesis contains high-affinity lowcapacity estrogen-binding proteins in the cytosol [l] and in the nucleus [2-101. The number of nuclear binding sites is increased several-fold after estrogentreatment of the animals [2,6,8 -lo]. Most likely this is due to a translocation of the cytoplasmic estrogen-receptor complex to the nucleus [1,9]. These characteristics of the estrogen-binding in the chicken liver are in agreement with those of steroid hormonebinding in other target organs (for review see [ l l , 121). Part of the nuclear binding sites can be easily extracted from nuclei with salt-containing buffers [2 -4, 91, whereas part remains sticking to the chromatin Enzyme. Trypsin (EC 3.4.21.4).Triviaf Numes. Estradiol or estradiol-17a, 1,3,5(10)-estratriene-3,17b-diol; estradiol-I7a, 1,3,5(1O)-estratriene-3,17~-diol; estrone, 3-hydroxy-l,3,5(10)-estratriene-l7-one; diethylstilbestrol, 4,4-dihydroxy-trans-7,7'-diethylstilbene; testosterone, 17P-hydroxy-4-androsten-3-one; progesterone, 4-pregnene-3,20-dione; cortisol, 11b. 17a,21-trihydroxy-4-pregene-3,20-dione.

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Section: Discussionsupporting

confidence: 66%

Section: Discussionmentioning

confidence: 62%

Estrogen-Binding Sites of Chicken Liver. Preliminary Characterization of Nuclear Components

Gschwendt

Schneider

1978

Eur J Biochem

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“…These receptors render hormone specificity to the target tissues and interaction with these proteins is a necessary first step for steroids to exert their effects at the nuclear level. Several laboratories have studied the characteristics of cytoplasmic progesterone receptor of the chick oviduct (Sherman et al, 1970(Sherman et al, , 1976Smith et al, 1974;Spelsberg & Cox, 1976;Toft et al, 1977) and our laboratory reported their purification Coty et al, 1979). Experiments in vitro showed that purified progesterone-receptor complexes can influence directly transcription of chick oviduct chromatin.…”

mentioning

confidence: 75%

“…Oviducts were excised from the oestrogenized animals and treated as before (Sherman et al, 1970(Sherman et al, , 1976Smith et al, 1974;Spelsberg & Cox, 1976;Toft et al, 1977). Cytosol was prepared by using a standard buffer containing 20mM-triethanolamine, pH 8.0, 1 mM-EGTA, and 12mM-1-thioglycerol.…”

Section: Preparation Of Crude Receptorfractionmentioning

confidence: 99%

Chemical cross-linking of chick oviduct progesterone-receptor subunits by using a reversible bifunctional cross-linking agent

Birnbaumer¹,

Schrader²,

O’Malley³

1979

Biochemical Journal

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Chick oviduct progesterone-receptor proteins were treated in cytosol with the reversible cross-linking reagent methyl 4-mercaptobutyrimidate. The product of the reaction was a 7S complex that could be detected and recovered after sucrose-density-gradient centrifugation in 0.3 M-KCl. The extent of the reaction was dependent on the concentration of methyl 4-mercaptobutyrimidate and independent of the presence of bound hormone, since unlabelled receptors could also be cross-linked. The cross-linking reaction required conditions in which the cytosol 6S complex was preserved. A Stokes radius of 7.3 nm was determined by gel filtration in Agarose A-1.5m in 0.3M-KCI. The sedimentation coefficient, which was also determined in 0.3M-KCI, allowed us to calculate a mol.wt. of 228 000. We were also able to cross-link partially purified receptor forms isolated by using an Agarose A-15m column. On reduction with 16-mercaptoethanol the complex broke down to 4S monomers that were identified by DEAE-cellulose and phosphocellulose chromatography, adsorption on DNA-cellulose and gel filtration in an Agarose A-i1.5 m column. In most cases, A and B receptor proteins were released in equivalent amounts, implying that the cross-linked form was an A-B complex.

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“…More recent findings, nevertheless, have localized a pyridoxal 5'-phosphate binding site to a proteolytic fragment of the receptor (42) which retains the capacity to bind dexamethasone. This receptor fragment has been referred to as the meroreceptor and is defined as the smallest proteolytic fragment of a steroid receptor which retains the capacity to specifically bind steroid (1,38,43). In these studies, rat thymic glucocorticoid mero-receptors were generated by proteolytic cleavage with either trypsin or chymotrypsin.…”

Section: Alteration Of Steroid Receptor Structure By B 6 Vitamersmentioning

confidence: 99%

Vitamin B₆and Glucocorticoid Action*

Compton

Cidlowski

1986

Endocrine Reviews

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T HE MECHANISMS by which steroid hormones interact with target tissues to induce endocrine responses have been studied since the early 1960's (for a recent review see Ref. 1), and many contributions have added to our current understanding of steroid hormone action. In essence, it is believed that the lipophilic steroid hormones enter target tissues by passively diffusing across the cell membrane. Once inside the cell, steroids bind reversibly to cytosolic receptor proteins in a stereospecific fashion to form steroid-receptor complexes. The actual localization of these receptors in the cytoplasm of target cells is a controversial subject (2); however, for the purposes of this discussion, it will be assumed that these receptors are available for steroid binding in the cytoplasmic compartment of the cell. This complex then undergoes a physicochemical transformation, termed activation, which enables the complex to bind tightly to appropriate chromatin acceptor sites. The phenomenon of steroid receptor activation is an incompletely understood process, but it can be simulated in vitro by a variety of techniques including warming of the receptor preparation from 0-25 C, exposure of the receptor complex to a high ionic strength environment, or dilution of the cytosol. After activation, the steroidreceptor complex associates tightly with the nucleus and binds to nuclear acceptor sites which probably consist of DNA and/or protein. This interaction between the receptor complex and chromatin results in the transcription of specific messenger RNAs that are subsequently translated into steroid-induced proteins.In the case of glucocorticoids, as well as other classes of steroids, hormonal regulation of gene expression is dependent upon a number of factors. The action of steroid hormones can be modulated by the nutrient status of the cell (3), cellular differentiation events (4, 5), and the stage of the cell cycle (6-8), as well as by factors

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Multiple forms of oviduct progesterone receptors analyzed by ion exchange filtration and gel electrophoresis

Cited by 99 publications

References 29 publications

Estrogen-Binding Sites of Chicken Liver. Preliminary Characterization of Nuclear Components

Estrogen-Binding Sites of Chicken Liver. Preliminary Characterization of Nuclear Components

Chemical cross-linking of chick oviduct progesterone-receptor subunits by using a reversible bifunctional cross-linking agent

Vitamin B₆and Glucocorticoid Action*

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Multiple forms of oviduct progesterone receptors analyzed by ion exchange filtration and gel electrophoresis

Cited by 99 publications

References 29 publications

Estrogen-Binding Sites of Chicken Liver. Preliminary Characterization of Nuclear Components

Estrogen-Binding Sites of Chicken Liver. Preliminary Characterization of Nuclear Components

Chemical cross-linking of chick oviduct progesterone-receptor subunits by using a reversible bifunctional cross-linking agent

Vitamin B6and Glucocorticoid Action*

Contact Info

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About

Vitamin B₆and Glucocorticoid Action*