Androgenic regulation of elongation of polyribonucleotide chains on rat ventral-prostate chromatin.
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ABSTRACT:
SUBMITTER: Thomas P
PROVIDER: S-EPMC1183886 | biostudies-other | 1978 Feb
REPOSITORIES: biostudies-other
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Project description:The characteristics of initiation of RNA synthesis and the elongation of RNA chains on rat ventral-prostate chromatin by RNA polymerase B were investigated by two methods. 1. Initiation was carried out under low-salt conditions with three ribonucleoside triphosphates, and elongation was begun in the absence of reinitiation by the addition of the fourth ribonucleoside triphosphate and increasing the salt concentration. 2. Stable initiation complexes were formed by preincubation of enzyme with template at 37 degrees C, elongation was started by the addition of all four ribonucleoside triphosphates and reinitiation or spurious RNA synthesis was prevented by rifamycin AF/013. The latter method gave more reliable results. The dependence of those parameters on the androgenic status of the animal was studied. During the first 24h after castration, elongation was mainly affected, whereas after 72h a smaller number of initiation sites for RNA polymerase B on chromatin was evident. Considerable diurnal variations in the various parameters were observed. Changes in the relative concentrations of the chromatin-associated proteins were also observed after castration. In the rat ventral-prostate gland androgenic steroids may not only influence one stage of the transcriptional process, but may affect many factors involved in the control of gene expression.
| S-EPMC1164994 | biostudies-other
Project description:The activity of superhelical-DNA nicking-closing enzyme (NC enzyme) was measured in nuclei from rat ventral prostate by a fluorimetric assay based on the binding of ethidium bromide to supercoiled phage-PM2 DNA. The nuclear concentration of NC-enzyme activity declined rapidly after castration, although this response could be prevented by daily administration of dihydrotestosterone. The low NC-enzyme activity in involuted prostates (10% of normal) was restored to normal after 8-10 days of treatment with androgen. In the regenerating prostate the time course of restoration of NC-enzyme activity was not in phase with that of DNA synthesis. Examination of nucleosome repeat lengths and the arrangement of nucleosomes along the chromatin fibre revealed no differences in the structural organization of chromatin in prostates with high or low NC-enzyme activity. Together, these results suggest that the major role of NC enzyme is related to the onset and maintenance of differentiation in the prostate and that the activity of this enzyme is not expressed through gross alterations in chromatin structure.
| S-EPMC1162982 | biostudies-other
Project description:The restoration of mitosis and growth of the prostate gland of castrated animals by androgens provides a favourable experimental system for studying the hormonal regulation of enzymes engaged in DNA replication. 2. Many DNA polymerase activities were identified in the prostate gland, but only a 9S form with a particular preference for denatured DNA as template was conspicuously enhanced by androgenic stimulation. 3. Thymidine kinase also provided a sensitive indicator of the hormonal regulation of DNA replication, and on electrophoretic criteria, one discrete form of the enzyme appeared precisely with the onset of mitoris. 4. Evidence is presented to support the view that DNA ligase activity is intimately associated in the process of DNA replication in the prostate gland. 5. A spectrum of deoxyribonuclease activities is present in the prostate gland, but only one form (pI7.0) can safely be said to be implicated in the process of DNA replication. 6. Androgenic stimulation of the prostate gland leads to the appearance of a component capable of denaturing or unwinding prostate DNA. This component is seemingly distinct from RNA or DNA polymerase activities on the basis of several distince physicochemical characteristics. 7. The conspicuous feature of all the changes in enzyme activities evoked by androgens in the prostate gland is their acute tissue- and steroid-specificity. Such changes could not be mimicked in liver or spleen and the regulatory role of androgens could not be simulated by other classes of steroid hormones. Particularly on the basis of studies with the anti-androgen cyproterone acetate, it is concluded that the changes are initially mediated by the androgen-receptor system and the high-affinity binding of 5alpha-dihydrotestosterone in the prostate gland. 8. The results are discussed in the context of the mechanism of action of androgens.
| S-EPMC1172433 | biostudies-other
Project description:The ventral urogenital sinus (UGS) of control male mice has two rows of 3-4 prostatic buds at birth, but how androgens regulate ventral bud (VB) number and patterning is unclear. VBs in both sexes appeared to be a mixture of prostatic and urethral buds. UGSs from Tfm male and antiandrogen (flutamide)-exposed mice had small VBs, suggesting that initiation of some VBs is androgen independent. Tfm male mice are widely considered completely androgen insensitive yet their UGSs were 5alpha-dihydrotestosterone (DHT)- responsive. VBs (6-8) were generally distributed bimodally on the left-right axis at both minimal and normal male androgen signaling. Yet control females and DHT-exposed Tfm males had 13-14 VBs, whose left-right distribution was fairly uniform. These results suggest that VB number and distribution respond biphasically as androgen signaling increases from minimal, and that androgens regulate bud specification. Complete VB agenesis by the selective budding inhibitor 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) required high androgen signaling.
| S-EPMC3041151 | biostudies-literature
Project description:Incidence of prostatic diseases increases dramatically with age which may be related to a decline in androgen support. However, the key mechanisms underlying prostate aging remain unclear. In the present study, we investigated the aging process in the ventral prostate (VP) of Noble rats by identifying differentially expressed prostate proteins between 3- and 16-month-old animals using ICAT and MS. In total, 472 proteins were identified with less than a 1% false positive rate, among which 34 were determined to have a greater than two-fold increase or 1.7-fold decrease in expression in the aged VPs versus their younger counterparts. The majority of the differentially expressed proteins identified have not been previously reported to be associated with prostate aging, and they fall into specific functional categories, including oxidative stress/detoxification, chaperones, protein biosynthesis, vesicle transport, and intracellular trafficking. The expression of GST, ferritin, clusterin, kininogen, oxygen regulated protein 150, spermidine synthase, ADP ribosylation factor, and cyclophilin B was verified by Western blot analyses on samples used for the ICAT study, as well as on those obtained from an independent group of animals comprised of three age groups. To the best of our knowledge, this is the first study on the proteome of the aging rat prostate.
| S-EPMC4482334 | biostudies-literature
Project description:1. The regulation by testosterone of mRNA complexity and mRNA activity was investigated in rat caput and cauda epididymidis. 2. The sequence complexity of cytoplasmic poly(A)-containing RNA from normal rats was determined by homologous hybridization with radiolabelled complementary DNA probes by using RNA in excess. Computer analysis of results suggested that hybridization could best be described by curves composed of two components distinguished by their relative abundance. Thus caput-epididymidal RNA consists of approx. 260 moderately abundant and 16400 scarce sequences, whereas cauda-epididymidal RNA consists of approx. 124 moderately abundant and 13400 scarce sequences. Judging by heterologous-hybridization reactions, castration did not result in appreciable alterations in either sequence complexity or the relative abundance of the two classes of poly(A)-containing RNA. 3. To investigate if individual mRNA sequences were regulated by androgens, mRNA was translated in a cell-free system derived from reticulocyte lysate. Since most of the translation products had a different mobility on sodium dodecyl sulphate/polyacrylamide gels from the authentic proteins synthesized in tissue minces, antibodies were used to identify specific translation products. Antibodies to the two related major proteins (mol.wt. 18500 and 19000) secreted by the caput epididymidis and whose synthesis is stimulated by testosterone both precipitated a single translation product of mol.wt. 21000. That this polypeptide was a precursor to the secreted proteins was suggested by the fact that the addition of microsomal membranes isolated from dog pancreas resulted in the appearance of a polypeptide of mol. wt. 19000. 4. Translation of RNA from the caput epididymidis of rats of different hormonal status showed that mRNA activity for the 21000-dalton polypeptide declined after castration, but could be restored by treating rats with testosterone. 5. It is concluded that testosterone stimulates the synthesis of a major protein secreted by the caput epididymidis by regulating its mRNA activity.
| S-EPMC1162125 | biostudies-other
Project description:Little is known about the roles of androgens in the regulation of redox state in the prostate, a cellular process believed to profoundly influence normal and aberrant prostate functions. We demonstrate that castration induced discrete oxidative stress (OS) in the acinar epithelium of rat ventral prostate (VP), as evident from marked increases in 8-hydroxy-2'-deoxy-guanosine and 4-hydroxynonenal protein adducts in the regressing epithelium. Testosterone replacement partially reduced OS in VP epithelia of castrates, but the level remained higher than in intact rats. Quantification of steady-state mRNA levels of 14 genes involved in the anabolism and catabolism of reactive oxygen species (ROS) showed that castration resulted in dramatic increases of three ROS-generating NAD(P)H oxidases (Noxs) including Nox1, gp91(phox), and Nox4, significant reductions of key ROS-detoxifying enzymes (superoxide dismutase 2, glutathione peroxidase 1, thioredoxin, and peroxiredoxin 5), and unchanged levels of catalase, glutathione reductase, gamma-glutamyl transpeptidase, and glutathione synthetase. Testosterone replacement in castrated rats partially reduced expression of Noxs but restored expression of superoxide dismutase 2, glutathione peroxidase 1, thioredoxin, and peroxiredoxin 5 to complete normalcy and induced a compensatory increase in expression of catalase, glutathione reductase, gamma-glutamyl transpeptidase, and glutathione synthetase in the regenerating VP. Expression of superoxide dismutase 1, glutathione S-transferase-pi, and glucose-6-phosphate dehydrogenase was unaffected by castration and testosterone replacement. These findings indicate androgen-deprivation induces OS in the rat VP through elevation of ROS anabolism and diminution of antioxidant detoxification. Androgen replacement partially reduces OS in rat VP to precastration levels. Expression of Noxs remained high amid a broad-based recovery of antioxidant defense mechanism(s). These data might have implications on the use of androgen blockade for prostate cancer prevention and androgen therapy for andropause treatment in elderly men.
| S-EPMC1892368 | biostudies-literature
Project description:Gene expression microarray analysis was performed on ventral prostate from normal adult rats, and adult rats treated BPA for 4 weeks(10 animals per group). Animals of 4 groups were treated with bisphenol-A (0,10, 30, or 90 ug/kg, i.g., daily) for 4 weeks, and the dosing volume is 10 ml/kg body weight. In this experiment, we only selected samples from two doses(0 and 10µg/kg) to analyze.
2014-01-15 | E-MTAB-2181 | biostudies-arrayexpress
Project description:1. Aldolase was selected as a suitable marker for following the androgenic regulation of mRNA synthesis in the prostate gland. 2. Antibodies raised in rabbits against crystalline prostate aldolase were used to monitor the synthesis of this androgen-induced enzyme after hormonal stimulation of castrated animals, by using procedures in vivo and in vitro for the translation of prostate poly(A)-rich mRNA. 3. After androgenic stimulation in vivo the poly(A)-rich mRNA was isolated from the prostate gland and other tissues of castrated rats, and added to a protein-synthesizing system in vitro derived from Krebs II ascites-tumour cells. By using this approach it was found that androgens regulate the synthesis of aldolase mRNA in a highly tissue-specific manner. Stimulation of aldolase mRNA synthesis reached a maximum after 8h of androgenic treatment and then declined. 4. The androgenic control of aldolase mRNA synthesis was also investigated in vivo. After treatment of castrated animals with various steroids in vivo [(35)S]methionine was injected directly into the prostate gland, and labelled aldolase was selectively precipitated from isolated polyribosomes with anti-aldolase serum. The regulation of aldolase mRNA synthesis in the prostate gland was stringently steroid-specific and could only be evoked by androgens. After a single injection of testosterone, aldolase synthesis reached a maximum after 16h of hormonal stimulation and then declined. 5. Although androgens exert significant control over transcriptional processes in the prostate gland, and appear to regulate the synthesis of aldolase mRNA de novo, the possibility exists for additional means of control at the translational level of aldolase synthesis. The results are discussed in the context of the overall mechanism of action of androgens.
| S-EPMC1168510 | biostudies-other
Project description:1. Labelled testosterone, injected directly into the ventral prostate of castrated rats became associated, in part, with a cytoplasmic high-molecular-weight fraction, fraction ;A'. 2. The label present in fraction ;A' was found to be mainly associated with dihydrotestosterone. 3. Unlike fraction ;A' from testosterone-pelleted castrated rats, fraction ;A' obtained from untreated castrated rats, 48h or more after castration, was strongly inhibitory towards Escherichia coli RNA polymerase in vitro. 4. The inhibition of RNA polymerase by fraction ;A' from castrated rats was not changed by the addition of testosterone or dihydrotestosterone in vitro, but pre-heating it to 80 degrees C resulted in a loss of its inhibitory capacity. 5. Fraction ;A' from castrated rats contained ribonuclease activity. The elution profile of ribonuclease activity from Sephadex columns indicated that this activity was responsible for the inhibitory effect on the RNA polymerase assays. 6. It is concluded that, unlike the inhibitor present in the uterus of ovariectomized rats (Talwar, Segal, Evans & Davidson, 1964), no direct connexion exists between the steroid-binding capacity of prostatic fraction ;A' and its effect on E. coli RNA polymerase activity in vitro.
| S-EPMC1178943 | biostudies-other