Cytoplasmic RNA Content Research Articles

Starvation and hypothyroidism exert an overlapping influence on rat hepatic messenger RNA activity profiles.

To assess the effect of starvation and to explore the potential interrelationship of starvation and thyroid status at the pretranslational level, we have analyzed by two-dimensional gel electrophoresis, the hepatic translational products of starved and fed euthyroid and hypothyroid rats. 5 d of starvation resulted in a statistically significant change in 27 of 240 products visualized, whereas hypothyroidism caused a change in 20, both in comparison with the fed euthyroid state. Of considerable interest was that 68% of all changing messenger (m)RNA sequences were common to the hypothyroid and starved groups and showed the same directional shift. Further, both starvation and hypothyroidism yielded comparable decreases in total hepatic cytoplasmic RNA content. Although it has been well established that the level of circulating triiodothyronine (T3) and the level of hepatic nuclear receptors fall in starvation, this reduction cannot account for the observed decrease of total hepatic RNA nor for all of the alterations in the concentrations of specific mRNA sequences. Thus, administration of T3 to starved animals in a dose designed to occupy all nuclear T3 receptors fails to prevent the fall in total RNA and the majority of starvation-induced changes in the level of mRNA sequences. Moreover, starvation of athyreotic animals results in a further decrease in total RNA and in a further change in the level of individual mRNA species. We conclude, therefore, that although the reduced levels of circulating T3 and the nuclear T3 receptors can contribute to the observed results of starvation, the starvation-induced changes are not exclusively mediated by this factor. The striking overlap in the genomic response between hypothyroid and starved animals raises the possibility that those biochemical mechanisms regulated at a pretranslational level by T3 are either not helpful or injurious to the starving animal. The reduction in circulating T3 and nuclear receptor sites together with T3-independent mechanisms initiated in the starved animal may constitute redundant processes designed to conserve energy and substrate in the nutritionally deprived organism.

Denervation modulated changes in mouse skeletal muscle RNA concentration.

We have examined normal and denervated mouse skeletal muscle for histological evidence of changes in RNA concentration using acridine orange/nucleic acid fluorescence. In denervated muscle, cytoplasmic RNA concentration increases dramatically. The earliest increase is in the subsarcolemmal region, 7 days after denervation. By 28 days after denervation, the increase in RNA is distributed throughout the cytoplasm of the atrophic cells, non-atrophic cells show little or no increase. Prior to the increase in cytoplasmic RNA concentration, many myocyte nuclei exhibit enlarged nucleoli, indicative of active ribosomal RNA synthesis. Such active nucleoli are present throughout the 42 day study period. We conclude that both new RNA synthesis, and preferential accumulation of cytoplasmic RNA are prominent events in denervation atrophy. There is an ordered progression from nucleolar synthesis to subsarcolemmal accumulation to diffuse accumulation in atrophying cells. These observations complement biochemical findings of others which indicate an increase in both RNA and protein synthesis after denervation.

Morphometric measurements and RNA content of axotomized feline cervical motoneurons.

Microspectrophotometric estimates of RNA content and morphometric measurements of cytoplasmic, nuclear and nucleolar areas were made on 30 to 60 motoneurons (somal areas greater than 1000 microns2) ipsilateral and contralateral to brachial plexotomy performed unilaterally on adult cats 2-90 days before sacrifice. Nerve cells of unoperated animals were also assayed. Somal and cytoplasmic areas of axotomized motoneurons were larger than those of the corresponding, contralateral motor nerve cells 4, 6 and 75 days postoperatively. Because of between animal variability, it could not be determined, however, whether this difference was due to an increase in the area of the axotomized motoneurons or to a decrease in the area of the contralateral nerve cells. Nucleolar sizes did not change. In contrast, nuclei of axotomized motoneurons showed a temporary but unequivocal areal decrease. The cytoplasmic RNA content of axotomized motoneurons fell 14-28 days postoperatively but rose thereafter, being increased slightly but significantly 75-90 days after operation. At no postoperative interval, however, did the nucleolar RNA content of the axotomized cells deviate unequivocally from the unoperated or zero day condition. The following points may be emphasized: 1. these results differ from similar measurements of axotomized motoneurons of rodents and lagomorphs; 2. the data do not provide certain evidence of change in either morphometric parameters or RNA content of motoneurons on the side contralateral to surgery, although the possibility of a decrease in the size of these uninjured neurons should be considered; 3. morphometric and RNA measurements on axotomized peripheral (extrinsic) neurons of spinal anterior horn of cat contrast with similar measurements on axotomized central (intrinsic) neurons of cat red nucleus.

Transcription, processing, and turnover of RNA from the Drosophila mobile genetic element copia

We have investigated the transcription, processing, and size distribution of copia nuclear RNA in Drosophila cultured cells in order to characterize the steps which result in the high concentration of cytoplasmic copia RNA. Ninety-five percent of pulse-labeled (5 min) copia nuclear RNA is 5 kb or shorter. This is consistent with the previously determined absence of intervening sequence(s) in the copia gene and indicates that there is little transcription of contiguous regions 5′ or 3′ to the copia sequence. In pulse-chase experiments, we observed that only 10% or less of the labeled copia nuclear RNA could be chased into the cytoplasm. Continuous labeling experiments, analyzed by taking into account changing nucleotide precursor pool specific activities during labeling and fitting the data simultaneously to a series of differential equations, gave results consistent with the pulse-chase experiments. The data, whether analyzed according to a one-step or a two-step processing model, indicated that total copia nuclear RNA is processed to poly(A) + cytoplasmic RNA with an efficiency of only 7%. In the fit to the two-step processing model, only 7% of the poly(A) + copia nuclear RNA is exported to the cytoplasm; the remainder turns over in the nucleus. The poly(A) + copia nuclear RNA is discrete in size, in contrast to poly(A) − copia nuclear RNA, which is heterogeneous in size. We discuss the implications of these observations for the mechanism of intranuclear processing and decay of copia RNA.

Effect of monaural and binaural stimulation on cytoplasmic RNA content in cells of the central nucleus of the cat inferior colliculus.

A cytophotometric study of sections stained with gallocyanin and chrome alum showed that monaural stimulation for 2 h and binaural stimulation for 1.5 h with rhythmic noise signals led to a marked increase in the cytoplasmic RNA content per cell in the principal and large multipolar neurons of the dorsal and ventral parts of the ventrolateral region of the central nucleus of the inferior colliculus. The increase in cytoplasmic RNA content in the principal cells of the ipsi- and contralateral parts of this nucleus relative to the stimulated ear in the case of monaural stimulation and the increase in RNA content in response to binaural stimulation suggests a uniform distribution of bilaterally converging connections from the lower nuclei of the auditory system on the principal cells. The increase in cytoplasmic RNA in the large multipolar cells of the contralateral central nucleus in response to monaural stimulation is evidence of the predominantly contralateral projection to these cells. The results are evidence of convergence of binaural influences on the principal and large multipolar cells of the central nucleus of the inferior colliculus.

Alterations in protein synthetic capability of nerve cells in Alzheimer's disease.

Cytoplasmic RNA content, nuclear and nucleolar volume are all significantly reduced in nerve cells of the temporal cortex in cases of Alzheimer's disease, examined both at diagnostic craniotomy and post mortem, when compared with non=demented control cases of similar age. On average, at necropsy, all three parameters are equally reduced by about 40-50%, but in biopsy cases, nuclear volume is decreased by the greatest amount (43%), followed by nucleolar volume is decreased by the greatest amount (43%), followed by nucleolar volume (36%), and cytoplasmic RNA (26%). These findings indicate that a change in protein synthesis occurs daily in the course of Alzheimer's disease which may result from a primary alteration within the nuclear apparatus.

Cytophotometric and morphometric studies of postnatal rat hippocampal neurons and perineuronal gliocytes after electroconvulsive shock

Electroconvulsive shock was evoked by transcorneal electric current (50 Hz, 50 mA; 3 times by 0.2s with 10-min pauses) in 10-day and 20-day old rats. 2 hours after the end of the convulsions, the total cytoplasmic RNA and protein content per cell in CA3 hippocampal neurons has been shown cytospectrophotometrically to increase markedly in 10-day old rats without statistically significant changes in 20-day old animals. No changes in the RNA and protein content were found in perineuronal neuroglia cells in the CA3 hippocampus in both age groups of rats. Morphometric determinations revealed a pronounced augmentation of volumes of the hippocampal neuronal bodies, nuclei, and cytoplasm as well as of nucleo-cytoplasmic ratio in 10-day old rats under effect of electroconvulsive shock. In 20-day old rats, the shock led to less marked increase in volumes of the hippocampal neurons, with no changes in nucleo-cytoplasmic ratio.

Changes in cytoplasmic RNA content of sympathetic neurons after synaptic block

Cytophotometry of RNA was carried out in neurons of the cranial cervical sympathetic ganglion of mature rabbits treated with different doses of the ganglion-blocker Dimecoline. After injection of the drug sharp fluctuations in the RNA content were observed in the cytoplasm of the neurons. The course of the fluctuations differed significantly depending on the dose of the drug injected, but it was similar in mono- and binuclear neurons. It is concluded from analysis of the character of differences found in the investigation that the results support the hypothesis of the leading role of synaptic processes in the formation of quantitative changes in the RNA content in the postsynaptic neuron. The results are evidence of increased functional activity of some ganglionic neurons after administration of different doses of Dimecoline both during and after the end of the block.

Uncoupling of RNA and DNA synthesis after plasma stimulation of G 0-arrested [formula omitted] cells

The addition of whole serum to G 0-arrested, confluent Balb c-3 T3 cells induces them to progress through G 1 and synthesize DNA after a 12-h lag period. Prior to the onset of DNA synthesis, RNA is synthesized and RNA content increases. Serum has been fractionated into two sets of growth factors: a platelet-derived growth factor present in heat-treated (100°C) platelet extracts and platelet-poor plasma. Addition of whole serum, platelet-derived growth factor or platelet-poor plasma induces quiescent cells to increase their cytoplasmic RNA content, but the cells treated with platelet-poor plasma do not synthesize DNA. Messenger RNA content increases within 2 h after stimulation with whole serum or platelet-poor plasma, and after 18 h, mRNA has accumulated to a greater degree than rRNA.

Effect of monaural and binaural stimulation on cytoplasmic RNA content in cells of the central nucleus of the cat inferior colliculus

Effect of monaural and binaural stimulation on cytoplasmic RNA content in cells of the central nucleus of the cat inferior colliculus

The relationship between lipofuscin pigment and ageing in the human nervous system

The cytoplasmic RNA content, nucleolar volume and lipofuscin content of nerve cells of the inferior olivary and dentate nuclei, Purkinje cells and pyramidal cells of the hippocampus has been measured in 82 persons of age range 2–91 years, who were free from overt neurological disease at the time of death. All 4 cell types accumulate lipofuscin in a linear manner throughout life, but to markedly differing extents. Both the RNA content and nucleolar volume of dentate, olivary and hippocampal cells decrease, with advancing age, in a similar way in all of these 3 cell types, despite the widely differing extents of pigment accumulation, with losses of RNA and nucleolar volume at 90 years of age of 15 and 30% respectively. Olivary cells which contain most pigment show a different pattern of change with losses of RNA and nucleolar volume of about 60%. It seems, therefore, that, if there is a causal relationship between lipofuscin accumulation and reductions in RNA and nucleolar volume, it is rather obscure. It cannot simply be a matter of the absolute amounts of pigment present in any one cell type, but rather might be that there is a critical concentration of such material in the cytoplasm.

Reactivity patterns of nerve cell-glia-complexes in mice during pentylenetrazole-induced seizures. Cytometric-cytophotometric studies.

The relationships between nerve cell-glia-complexes (NGCs) and surrounding capillaries were studied by combined cytometric-cytophotometric methods in male mice up to 60 min after injection of 70 milligrams/kilograms of pentylenetetrazole (PTZ). The nuclear area in neurons from cortical layers IV and V of the motoric area 8 increased by 13%, and in neurons from the hypoglossal nucleus there was an increase of 23%, associated with phasic changes in the nuclear area of their glial satellites. The number of perfused capillaries increased in both brain regions and there was a 20%-elongation of erythrocyte-filled capillary segments. The cytoplasmic RNA content enlarged in the hypoglossal neurons but decreased in cortical neurons from area 8. Peptidergic neurons and their glial satellites (supraoptic nucleus) showed nuclear shrinkage. Fifty percent of these peptidergic neurons reacted by reduction of their cytoplasmic RNA content. The measured cell reactions seem to be specific for different groups of NGCs which were defined by their histotopography, their neuronal interconnections, and a prevalence of secretory or nonsecretory neurons.

Regulation of malic enzyme synthesis by insulin triiodothyronine, and glucagon in liver cells in culture.

Cells isolated from the livers of 17- to 19-day-old chick embryos were maintained in a chemically defined culture medium. During 3 days in culture the activity of malic enzyme, a measure of its concentration, was stimulated 2-, 23-, or 77-fold by insulin, triiodothyronine, or insulin plus triiodothyronine, respectively. Glucagon blocked the stimulation caused by insulin plus triiodothyronine. Changes in the relative synthesis of immunologically isolated malic enzyme were similar in magnitude and direction to the changes in enzyme activity. Degradation of malic enzyme was unaffected by the three hormones. Both soluble protein and malic enzyme were degraded with a t1/2 of about 30 hours. In cells preincubated for 2 days with insulin, synthesis of malic enzyme was stimulated 4.5-fold within 3 hours after adding triiodothyronine and reached an apparent new steady state after 24 to 30 hours. If the rate of enzyme synthesis was dependent on the concentration of cytoplasmic malic enzyme messenger RNA, then this messenger RNA appeared to have a t1/2 of about 18 hours. Glucagon rapidly and specifically inhibited synthesis of malic enzyme in preinduced cells, suggesting an action at the level of translation or cytoplasmic messenger RNA processing.

Gradient of RNA distribution in the cytoplasm of Acetabularia mediterranea

Radiochemical studies and electrophoresis showed that there exists an apico-basal gradient of RNA concentration in Acetabularia mediterranea cytoplasm. The main contribution to the formation of such a gradient is made by the different rates of RNA turnover in the cytoplasm rather than by the transfer of nuclear RNA. High molecular weight RNA fractions synthesized in the cytoplasm originate from chloroplast ribosomes; their sedimentation constants are close to those of 23S and 16S rRNA fractions of E. coli.

Studies on the Nature of the Abnormality of B Cell Differentiation in Avian Lymphoid Leukosis: Production of Heterogeneous IgM by Tumor Cells

Abstract In chickens with lymphoid leukosis, immunoglobulin production, the distribution of immunoglobulin in tumor cells, the morphology of tumor cells, and the distribution of viruses were examined. Regardless of their location, virtually all of the lymphoma cells had detectable surface IgM; the prevalent distribution pattern produced by the bivalent fluorescein-labeled antibodies to IgM determinants was one of global patching. Variable amounts of cytoplasmic IgM were often detected in tumor cells. Release of IgM from tumor cells was indicated by the finding of high levels of serum IgM in birds with widespread lymphomas. Both high molecular weight and low molecular weight IgM were present in serum from tumorous birds, and both forms were electrophoretically heterogeneous. Lymphomatous birds had elevated titers of agglutinating antibodies to sheep and human erythrocytes but had normal levels of antibodies to Salmonella typhi flagellar antigens and leukosis virus neutralizing antibodies. None of the tumor cells contained or carried on their surface IgG or IgA heavy chain determinants; IgG levels were normal in tumor-bearing birds, but IgA levels were usually elevated. Multiple bursal follicles were involved by the tumor process. The tumor nodules in the bursa and in other organs of individual birds often differed in the size, cytoplasmic RNA content, and amount of cytoplasmic IgM that were characteristic of their constituent cells. Most of the tumor cells appeared not to achieve normal maturation; mature plasma cells were usually sparsely distributed around tumor nodules. The tumor cells were found by electron microscopy to contain an abundance of cytoplasmic ribosomes in clusters and scattered strands of rough endoplasmic reticulum. Virus particles budding from cell membranes of tumor cells were occasionally seen, but cell-associated viruses were much more abundant in a variety of non-tumorous cell types. These observations are interpreted to suggest that the avian leukosis viruses exert their lymphomagenic effect by interrupting the switch from IgM to IgG gene expression that normally occurs within clones of bursal lymphocytes.

A histological and histochemical study of the cotyledons of Phaseolus vulgaris L. during germination.

The cotyledon ofPhaseolus vulgaris L. comprises four tissues: epidermis, abaxial hypodermis, storage parenchyma, and procambium. A complex intercellular space system is present throughout the storage tissue and comprises about 16% of the cotyledon volume. All the cells contain protein bodies, and the hypodermis and storage parenchyma also contain starch grains. The epidermal cells are at the 2 C level of DNA, those of the hypodermis at the 4 C level, and the storage cells vary from 8 C to 32 C. During germination stomata differentiate in the epidermis. Reserve mobilization begins in the cells furthest from the epidermis and from the vascular tissue. Protein is removed from these cells with little or no coalescence of protein bodies. The DNA content of the nuclei decreases. The cell walls swell and then decrease in thickness as material is removed. Finally the nuclei and cytoplasm disappear and the cells collapse. In the cells near vascular bundles the protein bodies coalesce before losing their protein. The DNA content of the nuclei declines but nuclei and cytoplasm are still present at abscission. These cells do not collapse. Cytoplasmic RNA content is highest near the abaxial surface. Most of the RNA is removed during the first three days of germination.

Ultrastructure of the nuclear envelope of amphibian oocytes. IV. Chemical nature of the nuclear pore complex material.

In order to investigate the chemical composition of the nuclear pore complexes isolated nuclei from matureXenopus laevis oocytes were manually fractioned into nucleoplasmic aggregates and the nuclear envelopes. The whole isolation procedure takes no more than 60–90 sec, and the pore complexes of the isolated envelopes are well preserved as demonstrated by electron microscopy. Minor nucleoplasmic and cytoplasmic contaminations associated with the isolated nuclear envelopes were determined with electron microscopic morphometry and were found to be quantitatively negligible as far as their mass and nucleic acid content is concerned. The RNA content of the fractions was determined by direct phosphorus analysis after differential alkaline hydrolysis. Approximately 9% of the total nuclear RNA of the matureXenopus egg was found to be attached to the nuclear envelope. The nonmembranous elements of one pore complex contain 0.41×10−16 g RNA. This value agrees well with the content estimated from morphometric data. The RNA package density in the pore complexes (270×10−15 g/μ3) is compared with the nucleolar, nucleoplasmic and cytoplasmic RNA concentration and is discussed in context with the importance of the pore complexes for the nucleo-cytoplasmic transport of RNA-containing macromolecules.

Androgen-stimulated restoration in rat seminal vesicle and prostate epithelial cells.

Between 1 and 2 days after daily 150 μg injections of testosterone have begun the nuclear RNA content reaches peak values in both seminal vesicle and prostate epithelial cells of castrate rats. Between 2 and 6 days the nuclear RNA content decreased to normal control levels, and the cytoplasmic RNA content increased to normal control levels. DNA synthesis began between 1 and 2 days of testosterone injectionsinjections and was complete after 6 days. At the time of maximal mitotic rate, after 3 days of testosterone injections, marked increases were seen in cytoplasmic and nuclear protein-bound sulfhydryl and disulfide concentrations. In spite of cytoplasmic and nuclear enlargement, protein concentrations were maintained relatively constant throughout the experiment. (Endocrinology 84: 844, 1969)

Increased cytoplasmic RNA reflecting an early step of inductive tissue interaction

Cytoplasmic RNA concentration was measured with ultraviolet microspectrophotometry in metanephrogenic mesenchyme cells during morphological differentiation in vitro induced by spinal cord tissue. After 24 h in vitro, prior to any morphological sign of differentiation, cells from induced mesenchyme were shown to contain higher concentrations of cytoplasmic RNA than cells from non-induced tissue. A similar difference was found after 40 h in vitro. A gradual increase in cytoplasmic RNA concentration was found in undifferentiated cells from induced tissue during the first 48 h in vitro. Values for 48 and 72 h were the same. From 72 to 120 h there was a decrease. After 36 or 48 h of “induction” cells with early signs of specific differentiation, i.e. cells from “condensations” had slightly higher cytoplasmic RNA concentration than undifferentiated cells, while the values for tubular cells were slightly lower than for “condensate” -cells and after prolonged cultivation tubular cells had lower cytoplasmic RNA concentrations than undifferentiated cells. During induction, after initial morphological signs of differentiation, the mean cytoplasmic ultraviolet absorption maximum shifted slightly towards longer wavelength. It has earlier been shown that in this system induction is completed within 24 h. Apparently the nucleic acid synthesis has already been triggered by this period and may constitute the basis for subsequent autonomous cytodifferentiation.

The regulation of protein synthesis in the liver of rats. Mechanisms of dietary amino acid control in the immature animal

Weanling (23-day-old) rats were fed either on an amino acid-deficient diet (6% of casein, which in effect represents an ;amino acid-deficient' diet) or on a diet containing an adequate amount of protein (18% of casein) for 28 days. The hepatic cells from the animals fed on the low-protein diet were characterized by low amino acid content, almost complete inhibition of cell proliferation and a marked decrease in cell volume, protein content and concentration of cytoplasmic RNA compared with cells from control rats. The lower concentration of cytoplasmic RNA was correlated with a decreased ribosomal-RNA content, of which a larger proportion was in the form of free ribosomes. The protein-synthetic competence and messenger-RNA content of isolated ribosomes from liver cells of protein-deprived animals were 40-50% of those noted in controls. At 1hr. after an injection of radioactive uridine, the specific radioactivity of liver total RNA was greater in the group fed on the low-protein diet, but the amount of label that was associated with cytoplasmic RNA or ribosomes was significantly less than that noted in control animals. From these data it was concluded that dietary amino acids regulate hepatic protein synthesis (1) by affecting the ability of polyribosomes to synthesize protein and (2) by influencing the concentration of cytoplasmic ribosomes. It is also tentatively hypothesized that the former process may be directly related to the concentration of cellular free amino acids, whereas the latter could be correlated with the ability of newly synthesized ribosomal sub-units to leave the nucleus.