scholarly journals Changes in the ribonucleic acid metabolism of aging mouse tissues with particular reference to the prostate gland

1968 ◽  
Vol 110 (1) ◽  
pp. 79-86 ◽  
Author(s):  
W. I. P. Mainwaring
Keyword(s):  
Ribosomal Rna ◽  
Ribonucleic Acid ◽  
Messenger Rna ◽  
Rna Synthesis ◽  
Acid Metabolism ◽  
Prostate Gland ◽  
Mouse Tissues ◽  
Aging Mouse ◽  
Tissue Homogenates ◽  
Increased Susceptibility

1. The synthesis of RNA in purified nuclei of tissues from aged mice is substantially diminished. 2. As judged by the recovery of RNA in 105000g supernatants of tissue homogenates, the synthesis of ribosomal RNA may be particularly affected. 3. The decrease in RNA synthesis may be due to changes in the composition of the nuclear-associated protein with age, which result in changes in the priming ability of tissue chromatin. 4. Aging is accompanied by marked changes in the sedimentation profiles of ribonucleoprotein particles. These can be attributed to an age-associated depletion in messenger RNA or to an increased susceptibility to enzymic disaggregation.

scholarly journals Deoxyribonucleic acid–ribonucleic acid hybridization. Annealing and quantitative recovery of intact ribosomal ribonucleic acid molecules from hybrids

1969 ◽  
Vol 115 (2) ◽  
pp. 287-294 ◽  
Author(s):  
Michael Fry ◽  
Michael Artman
Keyword(s):  
Ribosomal Rna ◽  
Ribonucleic Acid ◽  
Messenger Rna ◽  
Deoxyribonucleic Acid ◽  
Cellulose Nitrate ◽  
Absolute Size ◽  
Rna Molecules ◽  
Membrane Filters ◽  
Ribosomal Ribonucleic Acid

A simple and efficient method for hybridization and subsequent recovery of non-fragmented ribosomal RNA from the hybrid is described. The procedure involves annealing of immobilized denatured DNA bound on cellulose nitrate membrane filters to complementary RNA in 50% (v/v) formamide–0·33m-potassium chloride–10mm-tris–hydrochloric acid buffer, pH7·4, at 33° for 3hr. Under these conditions no detectable changes in the sedimentation coefficients of the input RNA were detected. The RNA can subsequently be recovered quantitatively from the hybrid in intact form by incubating the filters in formamide or in 85% (v/v) dimethyl sulphoxide. The applicability of the method for the evaluation of the absolute size of ribosomal RNA cistrons in Escherichia coli DNA and for the determination of the size of messenger RNA molecules is discussed.

scholarly journals Characterization of rapidly labelled ribonucleic acid in Escherichia coli by deoxyribonucleic acid–ribonucleic acid hybridization

1968 ◽  
Vol 110 (2) ◽  
pp. 251-263 ◽  
Author(s):  
G. H. Pigott ◽  
J. E. M. Midgley
Keyword(s):  
Escherichia Coli ◽  
Ribosomal Rna ◽  
Ribonucleic Acid ◽  
Messenger Rna ◽  
Rna Binding ◽  
Hybridization Technique ◽  
E Coli ◽  
K 12 ◽  
Dna 2

1. Rapidly labelled RNA from Escherichia coli K 12 was characterized by hybridization to denatured E. coli DNA on cellulose nitrate membrane filters. The experiments were designed to show that, if sufficient denatured DNA is offered in a single challenge, practically all the rapidly labelled RNA will hybridize. With the technique employed, 75–80% hybridization efficiency could be obtained as a maximum. Even if an excess of DNA sites were offered, this value could not be improved upon in any single challenge of rapidly labelled RNA with denatured E. coli DNA. 2. It was confirmed that the hybridization technique can separate the rapidly labelled RNA into two fractions. One of these (30% of the total) was efficiently hybridized with the low DNA/RNA ratio (10:1, w/w) used in tests. The other fraction (70% of the total) was hybridized to DNA at low efficiencies with the DNA/RNA ratio 10:1, and was hybridized progressively more effectively as the amount of denatured DNA was increased. A practical maximum of 80% hybridization of all the rapidly labelled RNA was first achieved at a DNA/RNA ratio 210:1 (±10:1). This fraction was fully representative of the rapidly labelled RNA with regard to kind and relative amount of materials hybridized. 3. In competition experiments, where additions were made of unlabelled RNA prepared from E. coli DNA, DNA-dependent RNA polymerase (EC 2.7.7.6) and nucleoside 5′-triphosphates, the rapidly labelled RNA fraction hybridized at a low (10:1) DNA/RNA ratio was shown to be competitive with a product from genes other than those responsible for ribosomal RNA synthesis and thus was presumably messenger RNA. At higher DNA/rapidly labelled RNA ratios (200:1), competition with added unlabelled E. coli ribosomal RNA (without messenger RNA contaminants) lowered the hybridization of the rapidly labelled RNA from its 80% maximum to 23%. This proportion of rapidly labelled RNA was not competitive with E. coli ribosomal RNA even when the latter was in large excess. The ribosomal RNA would also not compete with the 23% rapidly labelled RNA bound to DNA at low DNA/RNA ratios. It was thus demonstrated that the major part of E. coli rapidly labelled RNA (70%) is ribosomal RNA, presumably a precursor to the RNA in mature ribosomes. 4. These studies have shown that, when earlier workers used low DNA/RNA ratios (about 10:1) in the assay of messenger RNA in bacterial rapidly labelled RNA, a reasonable estimate of this fraction was achieved. Criticisms that individual messenger RNA species may be synthesized from single DNA sites in E. coli at rates that lead to low efficiencies of messenger RNA binding at low DNA/RNA ratios are refuted. In accordance with earlier results, estimations of the messenger RNA content of E. coli in both rapidly labelled and randomly labelled RNA show that this fraction is 1·8–1·9% of the total RNA. This shows that, if any messenger RNA of relatively long life exists in E. coli, it does not contribute a measurable weight to that of rapidly labelled messenger RNA.

scholarly journals Synthesis and turnover of mitochondrial ribonucleic acid in HeLa cells: the mature ribosomal and messenger ribonucleic acid species are metabolically unstable.

1981 ◽  
Vol 1 (6) ◽  
pp. 497-511 ◽  
Author(s):  
R Gelfand ◽  
G Attardi
Keyword(s):  
Ribosomal Rna ◽  
Hela Cells ◽  
Ribonucleic Acid ◽  
Half Life ◽  
Messenger Rna ◽  
Specific Activity ◽  
Mitochondrial Rna ◽  
Messenger Rnas ◽  
Precursor Pool ◽  
Messenger Ribonucleic Acid

The synthesis rates and half-lives of the individual mitochondrial ribosomal ribonucleic acid (RNA) and polyadenylic acid-containing RNA species in HeLa cells have been determined by analyzing their kinetics of labeling with [5-3H]-uridine and the changes in specific activity of the mitochondrial nucleotide precursor pools. In one experiment, a novel method for determining the nucleotide precursor pool specific activities, using nascent RNA chains, has been utilized. All mitochondrial RNA species analyzed were found to be metabolically unstable, with half-lives of 2.5 to 3.5 h for the two ribosomal RNA components and between 25 and 90 min for the various putative messenger RNAs. A cordycepin "chase" experiment yielded half-life values for the messenger RNA species which were, in general, larger by a factor of 1.5 to 2.5 than those estimated in the labeling kinetics experiments. On the basis of previous observations, a model is proposed whereby the rate of mitochondrial RNA decay is under feedback control by some mechanism linked to RNA synthesis or processing. A short half-life was determined for five large polyadenylated RNAs, which are probably precursors of mature species. A rate of synthesis of one to two molecules per minute per cell was estimated for the various H-strand-coded messenger RNA species, and a rate of synthesis 50 to 100 times higher was estimated for the ribosomal RNA species. These data indicate that the major portion of the H-strand in each mitochondrial deoxyribonucleic acid molecule is transcribed very infrequently, possibly as rarely as once or twice per cell generation. Furthermore, these results are consistent with a previously proposed model of H-strand transcription in the form of a single polycistronic molecule.

Characterization of Rapidly Labelled Nucleic Acids in Tissues of Plant Seedlings

1966 ◽  
Vol 21 (10) ◽  
pp. 983-992 ◽  
Author(s):  
Vera Hemleben-Vielhaben
Keyword(s):  
Nucleic Acids ◽  
Ribosomal Rna ◽  
Messenger Rna ◽  
Specific Activity ◽  
Supernatant Fraction ◽  
Nucleic Acid Metabolism ◽  
Tissue Homogenates ◽  
Fraction I ◽  
Plant Seedlings

The nucleic acid metabolism of plant tissues was examined by incubating seedlings of Phaseolus, Vicia, Pisum, and Soja or sections of them with 32P for short periods of time. The nucleic acids extracted from this material were fractionated on columns of methylated albumin coated on kieselgur, and the radioactivity and composition of the specific fractions were determined. Application of 32P to intact seedlings or to excised parts of seedlings resulted in the same pattern of labelled nucleic acids in all tissues, but the total amount of incorporated radioactivity was different. In all tissues investigated five rapidly labelled RNA fractions were found associated with the following components: (I) soluble RNAs, (II) DNA, (III—V) ribosomal RNA. Fraction I contained equally high amounts of CMP and GMP thus differing significantly from the soluble RNAs. The composition of fraction (II) which was probably bound to DNA in the form of a complex, was similar to that of fraction I provided the labelling period was short. Fractions III—V were of the ribosomal type. The rapidly labelled DNA fraction had a high guanine-cytosine content (60%) as compared with the bulk DNA (40%). Fractional centrifugation of the tissue homogenates revealed that the labelled RNA of the ribosomal type was partly associated with the ribosomes, partly with larger particles which were sedimented by low speed centrifugation. The RNA associated with the latter had a higher specific activity than the ribosomal RNA in the supernatant. Fraction I and the second component of the soluble RNA (s-2) were also confined to the sediment of larger particles. Actinomycin D (10 µg/ml) inhibited the incorporation of 32P in the nucleic acids. In chase experiments it caused a decrease in the specific activity of all RNA fractions, most prominently, however, in fraction I and II indicating their instability and resemblance to messenger RNA.

Rates of synthesis of polyadenylated messenger RNA and ribosomal RNA during the cell cycle of Schizosaccharomyces pombe. With an appendix: calculation of the pattern of protein accumulation from observed changes in the rate of messenger RNA synthesis

1976 ◽  
Vol 21 (3) ◽  
pp. 497-521
Author(s):  
R.S. Fraser ◽  
F. Moreno
Keyword(s):  
Cell Cycle ◽  
Dna Synthesis ◽  
Schizosaccharomyces Pombe ◽  
Ribosomal Rna ◽  
Messenger Rna ◽  
Rna Synthesis ◽  
Protein Accumulation ◽  
Synchronous Culture ◽  
Synchronous Cultures ◽  
Gene Copies

The rates of polyadenylated messenger RNA and ribosomal RNA synthesis were measured in synchronously dividing cultures of fission yeast (Schizosaccharomyces pombe). Control asynchronous cultures, which had been exposed to the conditions used for preparing synchronous cultures, were investigated to check for effects of the synchronization procedure itself on RNA synthesis. After each period of DNA synthesis in synchronous culture, the rates of messenger and ribosomal RNA synthesis doubled, suggesting that gene number controls the rate of messenger and ribosomal RNA synthesis. This was confirmed by experiments with asynchronous, exponential-phase cultures in which DNA synthesis was inhibited by hydroxyurea. Both synchronous culture and hydroxyurea experiments suggested that there is a delay of 15 min (0-1 of the cell generation time) between replication of the DNA and transcription of both gene copies. A pattern of protein accumulation was calculated from changes in the rate of polyadenylated messenger RNA synthesis during synchronous culture. The simulated pattern indicates that protein is accumulated linearly, with a doubling in the rate of accumulation once per cell cycle. The simulated pattern of protein accumulation is very similar to measurements previously reported by other workers of changes in activities of 3 enzymes in synchronous cultures. It is suggested that the doubling of the rate of messenger RNA synthesis, as a consequence of the replication of the DNA once per cycle, provides the basis of a mechanism for control of the doubling of other cellular constituents during the cell cycle.

scholarly journals The cyclo-oxygenase inhibitors indomethacin and ibuprofen inhibit the insulin-induced stimulation of ribosomal RNA synthesis in L6 myoblasts

1989 ◽  
Vol 264 (1) ◽  
pp. 101-106 ◽  
Author(s):  
R M Palmer ◽  
G P Campbell ◽  
P F Whitelaw ◽  
D S Brown ◽  
P A Bain ◽  
...  
Keyword(s):  
Arachidonic Acid ◽  
Ribosomal Rna ◽  
Rna Synthesis ◽  
Acid Metabolism ◽  
Arachidonic Acid Metabolism ◽  
High Concentration ◽  
Ribosomal Subunits ◽  
Rrna Species ◽  
Inflammatory Drugs ◽  
Stimulation Of

Insulin stimulated total RNA accretion and the incorporation of [3H]uridine into RNA in L6 skeletal-muscle myoblasts. Incorporation of uridine into the rRNA was measured after either separation of 18 S and 28 S rRNA species by agarose-gel electrophoresis or separation of dissociated 40 S and 60 S ribosomal subunits on sucrose density gradients. Both methods showed a stimulation by insulin of uridine incorporation into the RNA of the two subunits. Two non-steroidal anti-inflammatory drugs, indomethacin and ibuprofen, which inhibit the metabolism of arachidonic acid by the cyclo-oxygenase pathway, inhibited the insulin-induced accretion of total cellular RNA and the incorporation of uridine into the RNA of both ribosomal subunits. The effect of insulin was observed both by using a tracer dose of [3H]uridine (5 microM) and in the presence of a high concentration (1 mM) of uridine to minimize possible changes in intracellular precursor pools. Neither insulin nor indomethacin was found to affect the incorporation of uridine into the total intracellular nucleotide pool, or the conversion of uridine into UTP. The ability of inhibitors of arachidonic acid metabolism to prevent insulin-induced increases in RNA metabolism suggests that a prostaglandin or other eicosanoid is involved in the signal mechanism whereby insulin stimulates RNA synthesis.

scholarly journals The control of ribonucleic acid synthesis in bacteria. Steady-state content of messenger ribonucleic acid in Escherichia coli M.R.E. 600

1970 ◽  
Vol 120 (2) ◽  
pp. 279-288 ◽  
Author(s):  
W. J. H. Gray ◽  
J. E. M. Midgley
Keyword(s):  
Escherichia Coli ◽  
Ribosomal Rna ◽  
Ribonucleic Acid ◽  
Messenger Rna ◽  
Acid Synthesis ◽  
Growth Conditions ◽  
Transfer Rna ◽  
Average Lifetime ◽  
Messenger Ribonucleic Acid ◽  
Wide Range

1. The technique of DNA–RNA hybridization was used to follow changes in the amount and average lifetime of unstable messenger RNA in Escherichia coli M.R.E. 600 over a wide range of different growth conditions. The method of analysis was based on the kinetics of incorporation of exogenous labelled nucleic acid bases into the RNA of steadily growing cultures, as described by Bolton & McCarthy (1962). 2. The ratio of the average lifetime of messenger RNA to the mean generation time of E. coli cultures was constant over the temperature range 25–45°C in a given medium, but the constant varied with the nature of the growth medium. For cultures growing in sodium lactate–salts or glucose–salts media the ratio was 0.046±0.005 and in enriched broth it was 0.087±0.009. Measurements of the amounts of transfer RNA, ribosomal RNA and messenger RNA were also made. The results confirmed earlier reports that the ratio of the amount of messenger RNA to the amount of ribosomes in the cells is virtually constant. On the other hand, the ratio of the amount of transfer RNA to the amount of ribosomal RNA decreased with increasing growth rate at a given temperature. 3. In cultures at temperatures higher than necessary for optimum rates of growth the average lifetime of messenger RNA lengthened in harmony with the increased time required for cell division. It seems that suboptimum growth rates at higher temperatures cannot be explained simply as a combination of increased rates of synthesis and breakdown of messenger RNA with a grossly decreased efficiency of translation. The absolute rate of messenger RNA synthesis was lowered, and its amount in the cells was typical of all other cultures grown at lower temperatures in the same medium. 4. The rate of entry of exogenous labelled uracil into unstable messenger RNA and stable ribosomal RNA was constant in all media at all temperatures in the approximate ratio 1:2. In media supporting a lower rate of growth, e.g. lactate–salts or glucose–salts media, the messenger RNA fraction constituted 2.2±0.3% of the total cellular RNA. In enriched broth 3.6±0.3% of the total RNA was messenger.

scholarly journals Nucleolar 4s ribonucleic acid in dipteran salivary glands in the presence of inhibitor

1968 ◽  
Vol 109 (3) ◽  
pp. 375-387 ◽  
Author(s):  
J. L. Sirlin ◽  
U E Loening
Keyword(s):  
Salivary Glands ◽  
Ribosomal Rna ◽  
Ribonucleic Acid ◽  
Rna Synthesis ◽  
Transfer Rna ◽  
Chromosome 2 ◽  
Insect Larvae ◽  
Nuclear Rna

1. Salivary glands of insect larvae accumulate newly made transfer RNA in the nucleolus when maintained in the presence of nucleoside antagonists that inhibit RNA synthesis preferentially at the chromosome. 2. The nucleus contains precursor transfer RNA, which, on the basis of the general evidence, may originate in the chromosome and then be methylated in the nucleolus. 3. The maturation of precursor ribosomal RNA is blocked in the nucleolus during inhibition. 4. The transport of nuclear RNA to cytoplasm is also blocked. 5. It is suggested that, if the transfer RNA accumulated in the nucleolus does indeed originate in the chromosome, the accumulation may result from the blockage of an obligatory transient association of the RNA with the nucleolus.

scholarly journals Synthesis and turnover of mitochondrial ribonucleic acid in HeLa cells: the mature ribosomal and messenger ribonucleic acid species are metabolically unstable

1981 ◽  
Vol 1 (6) ◽  
pp. 497-511
Author(s):  
R Gelfand ◽  
G Attardi
Keyword(s):  
Ribosomal Rna ◽  
Hela Cells ◽  
Ribonucleic Acid ◽  
Half Life ◽  
Messenger Rna ◽  
Specific Activity ◽  
Mitochondrial Rna ◽  
Messenger Rnas ◽  
Precursor Pool ◽  
Messenger Ribonucleic Acid

The synthesis rates and half-lives of the individual mitochondrial ribosomal ribonucleic acid (RNA) and polyadenylic acid-containing RNA species in HeLa cells have been determined by analyzing their kinetics of labeling with [5-3H]-uridine and the changes in specific activity of the mitochondrial nucleotide precursor pools. In one experiment, a novel method for determining the nucleotide precursor pool specific activities, using nascent RNA chains, has been utilized. All mitochondrial RNA species analyzed were found to be metabolically unstable, with half-lives of 2.5 to 3.5 h for the two ribosomal RNA components and between 25 and 90 min for the various putative messenger RNAs. A cordycepin "chase" experiment yielded half-life values for the messenger RNA species which were, in general, larger by a factor of 1.5 to 2.5 than those estimated in the labeling kinetics experiments. On the basis of previous observations, a model is proposed whereby the rate of mitochondrial RNA decay is under feedback control by some mechanism linked to RNA synthesis or processing. A short half-life was determined for five large polyadenylated RNAs, which are probably precursors of mature species. A rate of synthesis of one to two molecules per minute per cell was estimated for the various H-strand-coded messenger RNA species, and a rate of synthesis 50 to 100 times higher was estimated for the ribosomal RNA species. These data indicate that the major portion of the H-strand in each mitochondrial deoxyribonucleic acid molecule is transcribed very infrequently, possibly as rarely as once or twice per cell generation. Furthermore, these results are consistent with a previously proposed model of H-strand transcription in the form of a single polycistronic molecule.

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