scholarly journals Intracellular inactivation, reactivation and dynamic status of prostate androgen receptors

1982 ◽  
Vol 208 (2) ◽  
pp. 383-392 ◽  
Author(s):  
Gian Paolo Rossini ◽  
Shutsung Liao
Keyword(s):  
Protein Synthesis ◽  
Androgen Receptor ◽  
Half Life ◽  
Ventral Prostate ◽  
Receptor Complex ◽  
Nuclear Chromatin ◽  
Cytosol Fraction ◽  
Receptor Complexes ◽  
Rat Ventral Prostate ◽  
Energy Dependent

The dynamic status of the androgen receptor in prostate cells was studied by incubation of rat ventral prostate with radioactive 17β-hydroxy-5α-androstan-3-one (5α-dihydrotestosterone) in the presence and absence of respiratory poisons and inhibitors of protein and RNA synthesis and also by isotope chasing of the radioactive androgen–receptor complexes. The androgen receptor in the prostate appears to go through a dynamic process of recycling between the cytoplasm and the nucleus as well as an inactivation process. The radioactive androgen–receptor complex, however, is maintained at a constant level for at least 2h during incubation at 37°C, even in the absence of new protein synthesis, suggesting that early androgen actions may not require a depletion of a major portion of cellular receptor. In the presence of 2,4-dinitrophenol, the androgen receptor is rapidly deactivated (half life, 2min). The inactive receptor can be reactivated efficiently by an energy-dependent process, even in the absence of protein synthesis. Receptor binding of androgen and nuclear chromatin binding of the androgen–receptor complex are fast processes; half-maximum binding can be achieved within 1 and 10min respectively. On the contrary, the overall process of the release of the receptor complex from nuclear chromatin and its reappearance in the cytosol fraction has a long half life of about 70min. This slow process may reflect the involvement of the steroid–receptor complex in a time-consuming mechanism that is essential for hormone responses. Actinomycin D can increase the nuclear receptor level by 50% or more. This increase is not due to a decrease in the rate of receptor release from nuclei or to inhibition of DNA degradation by the antibiotic.

Extraction of androgen–receptor complexes from regions of rat ventral prostate nuclei sensitive or resistant to nucleases

1983 ◽  
Vol 99 (1) ◽  
pp. 51-61 ◽  
Author(s):  
P. Davies
Keyword(s):  
Androgen Receptor ◽  
Binding Sites ◽  
Micrococcal Nuclease ◽  
Ventral Prostate ◽  
Receptor Complex ◽  
Sedimentation Analysis ◽  
Receptor Complexes ◽  
Rat Ventral Prostate ◽  
Profile Characteristic ◽  
Androgen Binding

Rat ventral prostate nuclei contain androgen-binding sites which are susceptible or resistant to excision by endonucleolytic action. Those which were susceptible were associated both with oligonucleosomal and subnucleosomal particles. The sedimentation profile characteristic of a nuclear androgen-receptor complex could be obtained by exhaustive nucleolytic digestion or by treatment of fractions with KCl (0·6 mol/l). Androgen-binding sites resistant to DNAase I were also resistant to KCl, whereas those sites resistant to micrococcal nuclease were partially extractable with KCl. Nuclease-resistant sites could be extracted with heparin (10 mg/ml). Androgen–receptor complexes obtained from nuclease-sensitive or nuclease-resistant regions by extraction with KCl or heparin were indistinguishable by routine sedimentation analysis.

scholarly journals The use of deoxyribonucleic acid–cellulose chromatography and isoelectric focusing for the characterization and partial purification of steroid–receptor complexes

1973 ◽  
Vol 134 (1) ◽  
pp. 113-127 ◽  
Author(s):  
W. I. P. Mainwaring ◽  
R. Irving
Keyword(s):  
Androgen Receptor ◽  
Physicochemical Properties ◽  
Binding Proteins ◽  
Steroid Receptor ◽  
Partial Purification ◽  
Receptor Complex ◽  
Nuclear Chromatin ◽  
Receptor Complexes ◽  
Steroid Binding Proteins ◽  
Steroid Binding

1. Two characteristic properties of the specific high-affinity steroid-binding proteins or receptors, their ability to bind to DNA–cellulose and their relatively acidic isoelectric point, have been exploited as a means of purification. These two fundamental properties distinguish the receptors from the steroid-binding proteins in serum and the non-specific low-affinity steroid-binding proteins in hormone-responsive cells. 2. A significant degree of purification of both cytoplasmic and nuclear steroid–receptor complexes can be achieved with practical facility by these procedures. The purity of the receptor complexes is sufficient to enable studies on their possible control of metabolic processes to be investigated in the future. 3. After extensive purification the physicochemical properties of the cytoplasmic androgen–receptor complex, such as sedimentation coefficient, were unchanged. Further, the purified complex fully retained at least one of its fundamental physiological properties, namely the ability to transfer 5α-dihydrotestosterone (17β-hydroxy-5α-androstan-3-one) into chromatin in vitro. 4. The methods may also be employed for studying the changes in the structure and properties of the receptor complexes that are an essential prerequisite for the transfer of cytoplasmic receptor complexes into nuclear chromatin. The temperature-dependence of the binding of androgen–receptor complexes into chromatin is essentially due to a major change in cytoplasmic receptor complex before its attachment to nuclear chromatin. 5. The resolution of these analytical procedures was sufficient to enable a critical comparison of the receptor proteins from different male accessory glands to be undertaken. From these studies, no substantial evidence in support of the tissue specificity of androgen receptors could be established; rather the receptors from different androgen-dependent glands were remarkably similar in physicochemical properties. 6. Although the methods were initially developed for the partial purification of androgen–receptor complexes, they are equally suitable for the prompt and extensive purification of oestrogen–receptor and progesterone–receptor complexes.

DISTRIBUTION OF ACCEPTOR SITES FOR ANDROGEN-RECEPTOR COMPLEXES BETWEEN TRANSCRIPTIONALLY ACTIVE AND INACTIVE FRACTIONS OF RAT VENTRAL PROSTATE CHROMATIN

1980 ◽  
Vol 87 (2) ◽  
pp. 225-240 ◽  
Author(s):  
P. DAVIES ◽  
P. THOMAS ◽  
N. M. BORTHWICK ◽  
M. G. GILES
Keyword(s):  
Androgen Receptor ◽  
Rna Polymerase ◽  
Transcriptional Activity ◽  
Nuclear Dna ◽  
Intact Cell ◽  
Micrococcal Nuclease ◽  
Ventral Prostate ◽  
Receptor Complexes ◽  
Rat Ventral Prostate ◽  
Nascent Rna

Rat ventral prostate chromatin was separated into two main fractions by controlled digestion with micrococcal nuclease. The soluble fraction obtained after lysis of digested nuclei with EDTA (1 mmol/l), the S2 fraction, represented approximately 17% of the original nuclear DNA, and showed properties consistent with transcriptional activity, i.e. enrichment in nascent RNA, non-histone protein and endogenous RNA polymerase B activity as well as depletion in histones. The fraction sedimented after lysis of nuclei, fraction P, comprised approximately 60% of nuclear DNA, was depleted in nascent RNA, non-histone proteins and endogenous RNA polymerase B activity, but had a higher content of histones. In an attempt to relate the concentration of acceptor sites for androgen-receptor complexes with transcriptional activity, it was shown that the S2 fraction was enriched in these acceptor sites. However, if measurements were based on the intact cell the transcriptionally inactive portion contained 2·5–3 times as many 'acceptor' sites, although these sites had lower affinity for androgen-receptor complexes than had those in the transcriptionally active fraction.

scholarly journals Testosterone regulates the synthesis of major proteins in rat ventral prostate

1978 ◽  
Vol 170 (1) ◽  
pp. 115-121 ◽  
Author(s):  
M G Parker ◽  
G T Scrace ◽  
W I P Mainwaring
Keyword(s):  
Protein Synthesis ◽  
Cyproterone Acetate ◽  
Sodium Dodecyl ◽  
Ventral Prostate ◽  
Cytosol Fraction ◽  
Rat Ventral Prostate ◽  
General Protein Synthesis ◽  
Alpha Beta ◽  
General Protein

The presence of three major proteins alpha, beta and gamma in rat ventral prostate was demonstrated by electrophoresis in polyacrylamide gels containing sodium dodecyl sulphate. Their regulation by androgens was studied by measuring the rates of synthesis of the proteins in minced prostatic tissue by using L-[35S]methionine. The three proteins account for 30-40% of the proteins synthesized in the gland. After castration, their rates of synthesis rapidly decline to about 1% that of normal animals, and this cannot be accounted for by the accompanying decrease in general protein synthesis. Testosterone reverses these changes in castrated animals, so that after 4 days normal synthesis is restored. The regulation is specific for androgens, since cyproterone acetate, an anti-androgen, is inhibitory and oestradiol-17beta and corticosterone are without effect. Preliminary characterization of the proteins indicates that protein alpha (mol.wt. 22000, pI unknown) is a glycoprotein containing glucose and/or mannose residues and occurs in both the mitochondrial and cytosol fractions. Protein beta (mol.wt. 12000, pI5.4) is also a glycoprotein, but is found exclusively in the cytosol fraction. Protein gamma (mol.wt. 8000, pI5.4) is also a glycoprotein, but is found exclusively in the cytosol fraction. Protein gamma (mol.wt. 8000, pI5.4) is also found exclusively in the cytosol fraction.

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