Dynamics of quantitative RNA changes in the Purkinje cells of the rat cerebellum in various functional states

UV cytophotometry was used to study over time the effect of vestibular stimulation on RNA content in Purkinje's cells of the rat cerebellum nodulus. The animals placed in narrow boxes were rotated horizontally (60 r.p.m.) for an hour. To evaluate partially the effects of stress and hypoxia, the content of RNA was examined in the immobilized animals placed in the same boxes without rotation. Rhythmic quantitative changes in RNA in the cytoplasm and nucleus of Purkinje cells were observed upon vestibular stimulation of cerebellum function. In the nucleolus, the changes in the content of RNA were less pronounced. The periodic quantitative changes in the content of RNA in the cytoplasm, nucleus and nucleolus of Purkinje cells were statistically significant in the majority of the animals, but weakly pronounced.     

Protein SH-group concentration and RNA synthesis during the process of induction of proliferation in the stationary phase of cell culture growth]

The dynamics of intracellular protein SH-group (PSH) content was studied cytochemically in the course of stimulation of cell proliferation in stationary cultures of an established Chinese hamster cell line and of human diploid embryo fibroblasts. The results were compared with the pattern of RNA synthesis during the prereplicative period. In Chinese hamster cells immediately after medium changing in stationary cultures there is an augmentation of PSH content in parallel withe the increase in RNA synthesis rate. Later on, the rate of RNA synthesis and PSH content are seen decreasing followed by a new increase in the rate of RNA synthesis correlated with the second rise in PSH content. In stationary cultures of human diploid fibroblasts, there is also an increase in the rate of RNA synthesis and in the content of SH after medium changing, but the second wave of RNA synthesis and the second rise in PSH content are not pronounced. The variation in PSH content reflects the shift in the cell metabolism during the prereplicative period and is not attributed to changes in cell protein content.     

Quantitative cytochemical study of the RNA in tumor cells. II. The characteristics of the cell populatoins]

The quantity of DNA and RNA in tymocytes of AKR mice has been determined by Methylen blue staining. Cell populations of normal and tumor animals were divided into two groups with equal content of DNA to measure RNA content in either of them. The tumor cells had up to 380% RNA, if the normal content be taken for 100%. Dynamics of RNA modifications in cells of normal tymocytes corresponded to the increasing DNA quantity. The RNA quantity in tumor cells increased in the beginning of S-phase.     

RNA dependence in the cell cycle of V79 cells

The cell cycle of V79 Chinese hamster lung cells synchronized by hydroxyurea was investigated by flow cytometry. The metachromatic fluorochrome acridine orange was used to differentially stain DNA and RNA of V79 cells. Green and red fluorescence from individual cells, representing cellular DNA and RNA, respectively, was measured by flow cytometry. Periodic changes of cellular DNA and RNA contents were observed over nine cell cycles. The duration of G1, S, and G2 + M phases of synchronized V79 cells whose RNA content was close to that of the cells in balanced growth was 3, 4.5, and 1.5 hours, respectively. The duration of G1 and S phases of cells containing RNA above a certain threshold was inversely proportional to the RNA content. The RNA content of cells containing RNA above the normal level regressed to normal after a few generations. Coefficients of variation for RNA content were significantly larger than those for DNA. An explanation for the decay of synchrony in a synchronized cell population is proposed.     

Participation of the genetic apparatus of the hippocampal and neocortical cells in the mechanisms of action of desglycine-arginine vasopressin on learning and memory processes in rats

The paper deals with the influence of synthetic vasopressin analogue--desglycine-arginine vasopressin (DG-AVP) on the content of RNA and fractional composition of chromatin proteins in the tissue of neocortex and hippocampus of intact white rats and after establishing of two-way avoidance reflex. Administration of the peptide alone significantly increased RNA content in hippocampal tissue, injection of the peptide 10 min before conditioning did not lead to significant changes in RNA quantity as compared to that in animals in which the conditioned reflex was established against the background of saline administration. In neocortical tissue neither learning itself nor administration of DG-AVP alone was accompanied by significant changes in RNA content, while learning against the background of peptide injection significantly increased RNA in that structure. In hippocampal and neocortical tissues quantitative changes were observed in certain fractions of chromatin proteins in all animal groups studied.     

Circadian changes in the RNA content of rat hypothalamic nuclei in relation to food intake

The ribonucleic acid (RNA) and protein content of the ventromedial hypothalamic nuclei (VMH), the lateral hypothalamic areas (LHA) and the cortex (C) of adult male Wistar rats followed free access to food and water were determined at 6-hour intervals (at 6.00, 12.00, 18.00 and 24.00 h); the RNA and protein content of the suprachiasmatic nuclei (SCN) were determined at 12.00 and 24.00 h. The RNA content of the VMH, LHA and SCN differed at various intervals; in the cortex it did not alter significantly. The highest RNA concentration in the VMH was recorded at 18.00 h and in the LHA at 24.00 h, after which, in both areas, it fell. The lowest RNA content in the LHA and the VMH was found at 12.00 h. In the SCN, the RNA concentration at 24.00 h was significantly higher than at 12.00 h. The protein content did not alter significantly during the circadian cycle in any of the structures examined.     

Globin-coding sequences in the nuclear 28S pre-mRNA and in the cytoplasmic RNA of pigeon erythroid cells

The steady-state content of globin-coding sequences in nuclear and cytoplasmic RNA of pigeon erythroid cells was estimated by hybridization in the excess of nuclear 28S RNA and cytoplasmic poly(A) + RNA with [3H]DNA, synthesized on globin mRNA. Sequences of 9S globin mRNA are found in 0.06% of molecules of non-ribosomal 28S nuclear RNA (pre-mRNA) of erythroblasts and in 0.5% of molecules of non-ribosomal 28S nuclear RNA of reticulocytes. The content of globin mRNA in erythroblast cytoplasm is, respectively lower than in that of reticulocytes.     

Increase in the concentration of RNA in the blood erythrocytes of women in the ovulation period]

It was revealed that there was an increase (by an average of 58%) of the RNA concentration in the blood erythrocytes of women during ovulation. Measurement of the RNA content in erythrocytes can serve as an ovulation test.     

Changes in nuclear, nucleolar and cytoplasmic RNA content during growth and differentiation of root parenchyma cells in plant species with different dynamics of DNA endoreplication

Using cytophotometric method, after staining preparations with gallocyanin RNA content was examined in nucleus, nucleolus and cytoplasm of six species of angiospermal plants in successive (1-7 mm) segments of root representing successive zones of differentiation. During the cell cycle, RNA content duplicates in the nucleus, nucleolus and cytoplasm of meristematic cells. On the other hand, during growth and differentiation of parenchyma cells in species with endoreplication the content of nucleolar RNA does not increase in proportion with DNA content. High level of endoreplication is connected with high nucleolar RNA content and low cytoplasmic RNA content. In species without endoreplication at low nucleolar RNA content, a considerable growth of cytoplasmic RNA content takes place.     

Dynamics of T- and B-lymphocytes in the infectious process according to quantitative cytochemical findings]

Cytophotometric analysis with the use of autologous erythrocytes showed that T lymphocytes had the lowest RNA content and poorly differentiated lymphocytes had the highest RNA content, while in B cells an intermediate RNA level was observed, thus, the relative content of RNA could indicate to which particular system different peripheral blood cells belonged. In active infectious process, irrespective of the etiology of the disease, a decrease in both relative and absolute amounts of T lymphocytes and an increase in the amounts of poorly differentiated and B lymphocytes were observed.     

ppGpp is the major source of growth rate control in E. coli

It is widely accepted that the DNA, RNA and protein content of Enterobacteriaceae is regulated as a function of exponential growth rates; macromolecular content increases with faster growth regardless of specific composition of the growth medium. This phenomenon, called growth rate control, primarily involves regulation of ribosomal RNA and ribosomal protein synthesis. However, it was uncertain whether the global regulator ppGpp is the major determinant for growth rate control. Therefore, here we re-evaluate the effect of ppGpp on macromolecular content for different balanced growth rates in defined media. We find that when ppGpp is absent, RNA/protein and RNA/DNA ratios are equivalent in fast and slow growing cells. Moreover, slow growing ppGpp-deficient cells with increased RNA content, display a normal ribosomal subunit composition although polysome content is reduced when compared with fast growing wild-type cells. From this we conclude that growth rate control does not occur in the absence of ppGpp. Also, artificial elevation of ppGpp or introduction of stringent RNA polymerase mutants in ppGpp-deficient cells restores this control. We believe these findings strongly argue in favour of ppGpp and against redundant regulation of growth rate control by other factors in Escherichia coli and other enteric bacteria.     

Identification of polydisperse RNA in the cytoplasm of neurons isolated from the immature brain cortex

The cytoplasmic RNA of 10-day-old rats was studied in bulk-isolated, cortical neurons, obtained under conditions which minimize nuclear contamination. Two RNA fractions, one enriched in rRNA and the other in polydisperse RNA, were obtained by differential extraction with phenol. Gel electrophoresis and pulse labelling with 5-[³H]uridine were used to confirm the delayed appearance of newly synthesized rRNA in the cytoplasm and to demonstrate its stability. Polydisperse RNA appeared in the cytoplasm earlier and had a shorter half-life than rRNA. A wide range of molecular weights for this RNA was found with no predominant individual species. A set of cytoplasmic RNA components of molecular size between 28S and 18S was also present probably reflecting the in vivo degradation of rRNA. The significance of the unexpectedly high amounts of neuronal polydisperse RNA is discussed.     

Effect of the Kinetin-Naphthaleneacetic Acid Interaction Upon Total RNA and Protein in Senescing Detached Leaves

The interaction between kinetin and naphthaleneacetic acid in the regulation of senescence of excised tissue of mature broccoli leaves has been used to examine the extent of synchrony between changes in chlorophyll, RNA, and protein. Kinetin increased the net uptake of (14)C-labeled orotic acid and leucine. Naphthaleneacetic acid decreased the effect of kinetin on net uptake after long treatment, but in short-time treatments the auxin increased the effect of kinetin on net uptake. Results of long (24 hr) treatments indicated a general synchrony between the loss of RNA, protein, and chlorophyll. Naphthaleneacetic acid reduced the stabilizing effect of kinetin upon chlorophyll content and upon the content and synthesis of RNA. In short-time experiments, however, RNA content and synthesis were transiently increased by kinetin, and further increased by kinetin plus naphthaleneacetic acid, while chlorophyll content decreased in the presence of kinetin and decreased further in the presence of kinetin plus naphthaleneacetic acid. Actinomycin-D accelerated the loss of chlorophyll, RNA and protein and strongly depressed the rate of RNA synthesis. In the presence of actinomycin-D the stabilizing effect of kinetin upon RNA was substantially reduced. In contrast, the chlorophyll and protein contents remained higher than in the control. Actinomycin-D did not nullify the basal incorporation of orotic acid into RNA, nor did it negate the effect of kinetin upon incorporation. The failure of synchrony between changes in chlorophyll and RNA does not substantiate the proposal that kinetin regulates senescence by a direct effect upon DNA-dependent RNA synthesis.     

Change in the content and biosynthesis of RNA in the process of development of a Saccharomyces carlsbergensis culture]

The concentration of RNA in the cells of Saccharamyces carlsbergensis was found to vary from 3.51 to 7.54% with aging. A high content of RNA, a high rate of the incorporation of 14C-uridine into RNA and the acid-soluble fraction are typical of the cells in the logarithmic phase of growth. The total RNA can be separated into three zones on Sephadex G-100. No changes in the chromatographic profile were detected with aging.     

Reduction of the nucleic acid content of single-cell protein concentrates

Methods for reducing the content of nucleic acid in protein concentrates from disintegrated yeast and microalgae were investigated. Protein concentrates were prepared by acid precipitation of extracted protein after cell wall separation. The influence of alkaline protein extraction on the content of RNA in isoelectrically precipitated protein concentrates was studied. It was found that when a strong decrease in the RNA content was obtained, this was followed by a decrease in the yield of protein concentrate. Protein concentrates were also prepared without cell wall separation by precipitation with different agents after cell disintegration. In the precipitates from microalgae, a RNA reduction was obtained. Precipitation of yeast, protein gave no essential reduction with the precipitants used. Precipitation of yeast protein by heating at an alkaline pH gave a protein concentrate with a low content of RNA. A slightly lower RNA content was obtained when the precipitation was performed in the presence of NaCl. The yield of amino acid nitrogen was 70–80% and the RNA content was 1–2%. A process with precipitation at alkaline pH for the production of microbial protein concentrates with a low content of nucleic acid is suggested.     

The Relationship between the Kinetics of Ribonucleic Acid Accumulation and the Morphological Development of the Fern Gametophyte, Dryopteris borreri

Fern gametophytes were grown under blue light with and without the addition of 5-fluorouracil or 8-azaguanine, and under red light. Nucleic acids were extracted by either the detergent-chloroform or the detergent-diethylpyrocarbonate method and analyzed by polyacrylamide gel electrophoresis. No significant differences in the relative distribution of the stable RNA components accompanied the transition to biplanar growth. The RNA content per average cell decreased with growth and also varied between the cultural conditions, yet it was independent of the pattern of morphological development. The falling RNA content per average cell resulted from a progressive reduction of the RNA content of the apical cell, as determined histochemically. Since filamentous growth occurred by division of this apical cell, the rate of cell division was independent of the RNA content of the dividing cell.     

Macromolecular syntheses and the course of cell cycle events in the chlorococcal algaScenedesmus quadricauda under nutrient starvation: Effect of nitrogen starvation

Daughter cells of the chlorococcal algaScenedesmus quadricauda incubated under photosynthesizing conditions in a nitrogen-free medium did not make any progress in the cell cycle. Photosynthetic starch formation continued for a period corresponding to a half of the cell cycle and then levelled off. Protein synthesis was very slow and it did not surpass double the initial amount. RNA content decayed from the start of treatment and approached about 2 pg/cell. When a synchronous population was deprived of nitrogen or of light in the middle of the cell cycle RNA synthesis stopped immediately or very soon afterwards and, in spite ofabundant intracellular nitrogen reserves, RNA content slowly declined. This degradation was much extensive in nitrogen starved cells where, eventually, the RNA content attained about half the starting value. In both experimental variants, DNA replications started at the same time as in control culture, but the final amount of DNA attained only half the control value. Protein synthesis stopped immediately in the dark. In the nitrogen-starved cells, it continued for several hours and protein content increased about 70 % of the amount present at the start of starvation. The number of daughter cells formed was proportional to the final protein content in the nitrogen-and light-deprived cells (corresponding division numbers were 6 and 4, respectively). Upon refeeding of daughter cells formed under nitrogen starvation, RNA synthesis started immediately, while protein synthesis displayed a lag of about 5 h. DNA replications were triggered at the time when the ratio of RNA to DNA content attained the same value as in the control culture.     

Determination of the RNA-content in samples of isolated chromatin by circular dichroism measurements

A method is presented to determine the RNA content in isolated chromatin. An increased elipticity at 260nm–290nm in the circular dichroism spectrum of chromatin is due to RNA released together with chromatin from the cell nuclei. Mixtures of chicken erythrocyte chromatin and pure 5s-RNA were used to reconstruct such spectra. From the obtained data an RNA content higher than 3–5% in samples of isolated chromatin can be determined.     

Involvement of RNA and DNA in the staining of Escherichia coli by SYTO 13

AIMS: To assess the extent to which DNA and RNA bacterial content contributes to fluorescent response of SYTO 13. METHODS AND RESULTS: RNA and DNA of Escherichia coli 536 cells were extracted and fluorimetrically quantified to compare the different contents, throughout a 24 h culture, with their SYTO 13 fluorescence emission when analysed by the cytometer. SYTO 13 fluorescence varied depending on the stage of bacterial growth and in accordance with both DNA and RNA content. RNA content accounted for at least two-thirds of the total fluorescence of a cell. Escherichia coli cells were treated with chloramphenicol to improve their RNA content. With this treatment, both nucleic acids remained constant but there was a clear improvement in fluorescent emission. SYTO 13 fluorescence was also studied in E. coli X-1488 minicells. CONCLUSIONS: Although both nucleic acids are implicated, RNA accounts for a major part of SYTO 13 fluorescence. The fluorescence cannot be considered as a direct reflection of nucleic acid content. Other factors, such as topology or supercoiling, need to be considered. SIGNIFICANCE AND IMPACT OF THE STUDY: The results confirm the efficacy of SYTO 13 for labelling bacteria and for assessing the distinct physiological status. A better knowledge of the parameters implicated in its fluorescence emission has been achieved.     

Ribonuclease Activity and Auxin Effects in the Lens Root

In Lens root tips, a direct proportionality between RNA and auxin levels and inverse proportionality between RNA content and RNase activity were found. IAA treatment of the Lens seedlings causes, in the root, both an increase of RNA and auxin content and a decrease of RNase activity. Addition of IAA to the excised roots produces an inhibition of both the decrease of the RNA levels and an inhibition of the increase of the RNase activity. The action of IAA on growth, related to the control of the RNA metabolism is discussed.