scholarly journals Ribonucleic acid synthesis in vitro in primary spermatocytes isolated from rat testis

1977 ◽  
Vol 168 (1) ◽  
pp. 23-31 ◽  
Author(s):  
J. Anton Grootegoed ◽  
Anne H. Grollé-Hey ◽  
Focko F. G. Rommerts ◽  
Henk J. Van Der Molen
Keyword(s):  
Electrophoretic Mobility ◽  
Rna Synthesis ◽  
Actinomycin D ◽  
Marked Change ◽  
Acid Synthesis ◽  
Specific Pattern ◽  
28S Rrna ◽  
Phenol Extraction

The incorporation of [3H]uridine into RNA was studied quantitatively (by incorporation of [3H]uridine into acid-precipitable material) and qualitatively (by phenol extraction and electrophoretic separation of RNA in polyacrylamide gels) in preparations enriched in primary spermatocytes, obtained from testes of rats 26 or 32 days old. The rate of incorporation of [3H]uridine into RNA of isolated spermatocytes was constant during the first 8h of incubation, after which it decreased, but the decreased rate of incorporation was not reflected in a marked change in electrophoretic profiles of labelled RNA. In isolated spermatocytes, [3H]uridine was incorporated mainly into heterogeneous RNA with a low electrophoretic mobility. Most of this RNA was labile, as shown when further RNA synthesis was inhibited with actinomycin D. Spermatocytes in vivo also synthesized heterogeneous RNA with a low electrophoretic mobility. A low rate of incorporation of [3H]uridine into rRNA of isolated spermatocytes was observed. The cleavage of 32S precursor rRNA to 28S rRNA was probably retarded in spermatocytes in vitro as well as in vivo. RNA synthesis by preparations enriched in early spermatids or Sertoli cells was qualitatatively different from RNA synthesis by the spermatocyte preparations. It is concluded that isolated primary spermatocytes maintain a specific pattern of RNA synthesis, which resembles RNA synthesis in spermatocytes in vivo. Therefore isolated spermatocytes of the rat can be used for studying the possible regulation of RNA synthesis during the meiotic prophase.

Synthesis of acid-soluble spore proteins by Bacillus subtilis

1982 ◽  
Vol 152 (3) ◽  
pp. 1117-1125
Author(s):  
J M Leventhal ◽  
G H Chambliss
Keyword(s):  
Bacillus Subtilis ◽  
Maximum Rate ◽  
Rna Synthesis ◽  
Actinomycin D ◽  
Sodium Dodecyl ◽  
Molecular Weights ◽  
Spore Proteins ◽  
Major Acid

The major acid-soluble spore proteins (ASSPs) of Bacillus subtilis were detected by immunoprecipitation of radioactively labeled in vitro- and in vivo-synthesized proteins. ASSP synthesis in vivo began 2 h after the initiation of sporulation (t2) and reached its maximum rate at t7. This corresponded to the time of synthesis of mRNA that stimulated the maximum rate of ASSP synthesis in vitro. Under the set of conditions used in these experiments, protease synthesis began near t0, alkaline phosphatase synthesis began at about t2, and refractile spores were first observed between t7 and t8. In vivo- and in vitro-synthesized ASSPs comigrated in sodium dodecyl sulfate-polyacrylamide gels. Their molecular weights were 4,600 (alpha and beta) and 11,000 (gamma). The average half-life of the ASSP messages was 11 min when either rifampin (10 micrograms/ml) or actinomycin D (1 microgram/ml) was used to inhibit RNA synthesis.

Ribonucleic acid synthesis inPlasmodium knowlesimaintained bothin vivoandin vitro

Parasitology ◽  
1975 ◽  
Vol 71 (2) ◽  
pp. 199-209 ◽  
Author(s):  
P. I. Trigg ◽  
P. G. Shakespeare ◽  
Susan J. Burt ◽  
Sally I. Kyd
Keyword(s):  
Ribosomal Rna ◽  
Rna Synthesis ◽  
Nuclear Division ◽  
Plasmodium Knowlesi ◽  
Acid Synthesis ◽  
Agarose Gel Electrophoresis ◽  
Trophozoite Stage ◽  
Late Trophozoite

RNA extracted from purified parasites ofPlasmodium knowlesiwas fractionated using agarose gel electrophoresis. Preparations from parasites grown bothin vivoandin vitrocontained species of RNA with sedimentation coefficients of 4·0S, 5·0S, 16·6S, 24·2S, 31·4S, 38·0S and 48·3S. There was less RNA present in parasites grownin vitrothan the equivalent stage parasites grownin vivobut the proportional amounts of the various species of RNA was similar in both cases. It is suggested that the 24·2S and 16·6S species of RNA are ribosomal and that the high molecular weight 31·4S, 38·0S and 48·0S species are ribosomal precursors. Ribosomal RNA synthesis occurs throughout the cell cycle during growth from the ring to the schizont stage; maximum incorporation of [H3]-adenosine occurs at the late trophozoite stage before nuclear division.

ACTION OF KI, THYROXINE AND CYCLIC AMP ON [3H]URIDINE INCORPORATION INTO THE RNA OF THYROID SLICES

1976 ◽  
Vol 83 (2) ◽  
pp. 313-320 ◽  
Author(s):  
Mario A. Pisarev ◽  
Leonardo O. Aiello ◽  
Diana L. Kleiman de Pisarev
Keyword(s):  
Cyclic Amp ◽  
Inhibitory Action ◽  
Rna Synthesis ◽  
Actinomycin D ◽  
Protein Biosynthesis ◽  
Rna Degradation ◽  
Precursor Pool ◽  
Rna Precursor

ABSTRACT Potassium iodide (KI) has been shown to impair thyroid protein biosynthesis both in vivo and in vitro. The present study was performed in order to clarify its mechanism of action. Ribonucleic acid (RNA) synthesis was studied in beef thyroid slices with either [32P] or [3H]-uridine as labelled precursors. Both KI and thyroxine (T4) at 10−5 m significantly decreased RNA labelling under our conditions. In other experiments RNA degradation was examined in pulse-labelled and actinomycin D-treated slices. KI did not modify the degradation of the [3H]-RNA thus indicating that it interferes with the biosynthesis rather than with the degradation of RNA. Taking the perchloric acid soluble radioactivity as a rough index of the precursor pool the present results would indicate an action at this level. Both KClO4 and methylmercapto-imidazole relieved the gland from the inhibitory action of KI, supporting the view that an intracellular and organified form of iodine is responsible for this action. Since T4 also reproduced the effects of KI on RNA synthesis we would like to propose iodothyronines as the intermediates of this action. Cyclic AMP has been shown to stimulate thyroid protein biosynthesis. The present results demonstrate an action at the RNA level. Cyclic AMP increased both the PCA-soluble and RNA-linked radioactivity, thus suggesting an effect at the RNA precursor pool. KI at 10−5 m blocked the action of 2 mm cyclic AMP.

scholarly journals Ribonucleic acid synthesis in the renal cortex at the initiation of compensatory growth

1976 ◽  
Vol 158 (2) ◽  
pp. 457-470 ◽  
Author(s):  
P Cortes ◽  
N W Levin ◽  
P R Martin
Keyword(s):  
Rna Synthesis ◽  
Compensatory Growth ◽  
Renal Cortex ◽  
Acid Synthesis ◽  
Sham Operation ◽  
Precursor Pool ◽  
Cortex Slices ◽  
Endogenous Precursor

The mechanisms responsible for the increase in RNA per cell during the first 48h of renal compensatory growth were studied in the renal cortex. Unilaterally nephrectomized, sham-operated or non-operated rats were used. Incorporation into RNA of labelled precursors was studied in vivo and in vitro. Sham-operation produced significant changes in precursor incorporation, absolute amounts of UTP and RNA, and the rate of RNA synthesis. At 6h after surgery, the amount of RNA decreased in sham-operated controls, whereas that in growing cortex remained unchanged. Incorporation into RNA in vivo was greater in the growing cortex, although the rate of RNA synthesis was not increased. At 24h, precursor incorporation into RNA and UTP and RNA synthesis were all increased in the growing cortex. In contrast with results obtained in vivo, slices of growing cortex incorporated less labelled precursor into RNA than did cortex slices from sham-operated controls, from 3 to 48h. Maximal differences were found from 6 to 24h. An attempt was made to equalize endogenous precursor pool sizes by increasing the concentration of unlabelled uridine in the media; incorporation differences were narrowed significantly. Serum from nephrectomized animals did not increase precursor incorporation into RNA in vitro. An increase in RNA synthesis is an important factor in RNA accretion in the renal cortex beyond 12h of compensatory growth. This is accompanied by increased UTP content and preceded by expansion of other pools. The amount of labelled precursor incorporated into RNA is greatly influenced by its delivery rate to the growing kidney in vivo and by intracellular dilution of expanded precursor pools in vitro.

scholarly journals Ribonucleic acid synthesis in porphyric rat liver induced by 3,5-dicarbethoxy-1,4-dihydrocollidine

1973 ◽  
Vol 136 (1) ◽  
pp. 209-215 ◽  
Author(s):  
Hajime Nawata ◽  
Kenichi Kato
Keyword(s):  
Rat Liver ◽  
Liver Cell ◽  
Ribonucleic Acid ◽  
Rna Synthesis ◽  
Actinomycin D ◽  
Acid Synthesis ◽  
Rapid Decrease ◽  
Aminolaevulinic Acid ◽  
Induction Mechanism

1. Administration of 3,5-dicarbethoxy-1,4-dihydrocollidine to rats caused a marked increase in the activity of δ-aminolaevulinic acid synthetase as well as a slight net increase in RNA in the livers. 2. 3,5-Dicarbethoxy-1,4-dihydrocollidine primarily stimulated the synthesis of RNA in the nucleus of the liver cell. 3. The decrease in RNA synthesis after administration of this drug resulted in a rapid decrease in the activity of this induced enzyme. This was also confirmed by treatment with actinomycin D. 4. From kinetic experiments on synthesis of RNA and δ-aminolaevulinic acid synthetase in vivo the induction mechanism of this enzyme was discussed.

scholarly journals INTRACELLULAR SYNTHESIS, TRANSPORT, AND PACKAGING OF PROTEINACEOUS YOLK IN OOCYTES OF ORCONECTES IMMUNIS

1972 ◽  
Vol 52 (2) ◽  
pp. 420-437 ◽  
Author(s):  
L. R. Ganion ◽  
R. G. Kessel
Keyword(s):  
Amino Acids ◽  
Endoplasmic Reticulum ◽  
Rna Synthesis ◽  
Actinomycin D ◽  
Maternal Blood ◽  
Blood Proteins ◽  
Yolk Granules ◽  
Early Phases

The incorporation of leucine-3H into either ovarian or oocyte proteins occurs throughout vitellogenesis, but is at a maximum during early phases of this process. The labeling of ovarian and oocyte proteins is inhibited with cycloheximide. Oocytes are permeable to actinomycin D, and this drug does not affect the incorporation of amino acids into oocyte proteins but does block oocyte RNA synthesis. By means of both light microscope and high resolution radioautography, it has been demonstrated that the initial incorporation of leucine-3H under both in vitro and in vivo conditions occurs in elements of the rough-surfaced endoplasmic reticulum in the oocyte. Under pulse-chase conditions, the label subsequently becomes associated with intracisternal (precursor yolk) granules now aggregated within the cisternae of the connected smooth-surfaced endoplasmic reticulum. By 7 days, mature yolk globules are extensively labeled. The results of experiments designed to assess the possible contribution of maternal blood proteins to yolk deposition indicate that such a contribution is minimal. It is concluded that the crayfish oocyte is programmed for and capable of synthesizing the massive store of proteinaceous yolk present in the egg at the end of oogenesis.

scholarly journals METABOLIC AND ULTRASTRUCTURAL CHANGES IN THE FROG OVARIAN FOLLICLE IN RESPONSE TO PITUITARY STIMULATION

1970 ◽  
Vol 46 (3) ◽  
pp. 491-504 ◽  
Author(s):  
John Walberg Anderson ◽  
Milton B. Yatvin
Keyword(s):  
Epithelial Cells ◽  
Rna Synthesis ◽  
Actinomycin D ◽  
Ovarian Tissue ◽  
Ovarian Follicle ◽  
Ovarian Follicles ◽  
Ultrastructural Changes ◽  
Surface Epithelium

Frog ovarian fragments were prevented from ovulating in vitro by the addition of actinomycin D up to 3 hr following pituitary stimulation; but addition of Actinomycin D 6 hr after stimulation was far less effective. Puromycin, on the other hand, effectively inhibited ovulation when added as late as 6 hr after pituitary stimulation. Although actinomycin D reduced uptake of uridine-3H, and puromycin reduced uptake of leucine-3H and lysine-14 by pituitary-stimulated ovarian tissue minus oocytes (OTMO) in vitro, it was found that pituitary stimulation did not significantly increase uptake of these compounds by OTMO. Radioautographs of ovarian follicles fixed 6 hr after the addition of pituitary extract and uridine-3H in vitro revealed increased RNA synthesis in the peritoneal surface epithelium, compared with unstimulated controls, while the ovarian sac epithelium showed no increase. Gross ultrastructural changes occurred in the peritoneal area of ovarian follicles following pituitary stimulation in vivo, including loss of collagen fibrils, and general disorganization of the connective tissue theca. Changes in the rough endoplasmic reticulum of the peritoneal epithelial cells, while frequently encountered, were less pronounced. None of these changes was observed in the ovarian sac area, or in the interfollicular region. The above data are consistent with the hypothesis that pituitary stimulation of the frog ovary results in increased synthesis of RNA and protein by the peritoneal epithelial cells, and that the protein may be collagenase.

Effect of Some Metabolic Inhibitors on Vinblastine-Induced Ribosomal-Granular Material Complexes

1971 ◽  
Vol 29 ◽  
pp. 548-549
Author(s):  
Awtar Krishan ◽  
Dora Hsu
Keyword(s):  
Granular Material ◽  
Rna Synthesis ◽  
Culture Medium ◽  
Actinomycin D ◽  
Metabolic Inhibitors ◽  
Protein And Rna Synthesis ◽  
Apparent Reduction ◽  
Plant Alkaloids ◽  
In Cells

Cells exposed to antitumor plant alkaloids, vinblastine and vincristine sulfate have large proteinacious crystals and complexes of ribosomes, helical polyribosomes and electron-dense granular material (ribosomal complexes) in their cytoplasm, Binding of H3-colchicine by the in vivo crystals shows that they contain microtubular proteins. Association of ribosomal complexes with the crystals suggests that these structures may be interrelated.In the present study cultured human leukemic lymphoblasts (CCRF-CEM), were incubated with protein and RNA-synthesis inhibitors, p. fluorophenylalanine, puromycin, cycloheximide or actinomycin-D before the addition of crystal-inducing doses of vinblastine to the culture medium. None of these compounds could completely prevent the formation of the ribosomal complexes or the crystals. However, in cells pre-incubated with puromycin, cycloheximide, or actinomycin-D, a reduction in the number and size of the ribosomal complexes was seen. Large helical polyribosomes were absent in the ribosomal complexes of cells treated with puromycin, while in cells exposed to cycloheximide, there was an apparent reduction in the number of ribosomes associated with the ribosomal complexes (Fig. 2).

CONTRIBUTION TO THE EXISTENCE OF REGULATORY MECHANISMS AT THE ADRENAL LEVEL

1972 ◽  
Vol 70 (4) ◽  
pp. 741-757
Author(s):  
Otto Linèt
Keyword(s):  
Orotic Acid ◽  
Adrenal Glands ◽  
Actinomycin D ◽  
Delay Period ◽  
Regulatory Mechanisms ◽  
Leucine Incorporation ◽  
Total Period ◽  
Free Media

ABSTRACT Rat adrenal glands atrophied by the administration of cortisol acetate in vivo were used as a model for the study of early metabolic processes occurring in vitro. Atrophied adrenals incubated in the presence of 14C-leucine incorporated subnormal quantities of this amino acid per mg of protein for the first 120 min. When the incubation lasted for a total period of 180 or 240 min a supranormal rise in the 14C-leucine incorporation was observed. Similar changes occurred with some delay with regard to corticosterone production as expressed per 100 mg of tissue. No differences in 14C-leucine incorporation were observed between the control and atrophied adrenals in vivo. Homogenates from atrophied glands incorporated 14C-leucine to a greater extent than the control homogenates. The in vitro incorporation of 14C-orotic acid into the RNA was also higher in atrophied adrenals. The in vitro use of actinomycin D, cycloheximide and amphenone indicated that corticosterone production depended on the incorporation of 14C-leucine. The addition of cortisol to the incubation media markedly decreased the enhancement of 14C-lysine incorporation into the protein of atrophied adrenals. These, as well as additional results suggest rebound phenomena: once atrophic adrenals are transferred to cortisol-free media, reparative processes begin after a delay period. Such phenomena seem to be mediated by regulatory mechanisms at the adrenal level.

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