RNA-DNA Ratio Research Articles

Recent Growth, Biochemical Composition and Prey Field of Larval Haddock (Melanogrammus aeglefinus) and Atlantic Cod (Gadus morhua) on Georges Bank

A transect across southern Georges Bank in May 1983 showed higher levels of available prey for haddock (Melanogrammus aeglefinus) and cod (Gadus morhua) larvae at two stratified sites than at a well-mixed site. At the stratified sites, prey biomass was high (30–300 μg dry wt∙L−1) near the surface above the thermocline; values were lower and more uniform with depth (10–30 μg dry wt∙L−1) at the well-mixed site. Larval population centers generally coincided with prey biomass vertically. Recent growth in dry weight of haddock larvae as estimated by RNA–DNA ratio analysis was higher at the stratified sites (8–13%∙d−1) than at the well-mixed site (7%∙d−1). Larvae appeared to be in excellent condition at the stratified sites, but up to 50% of haddock larvae from the well-mixed site had RNA–DNA ratios in the range observed for starved larvae in the laboratory. Cod collected at the same site were in better condition and growing faster than haddock. The data support the hypotheses that (1) stratified conditions in the spring favor good growth and survival of haddock larvae and (2) cod larvae are better adapted to grow and survive in well-mixed waters at lower levels of available food than haddock larvae.

Correlated Measurements of DNA, RNA, and Protein in Individual Cells by Flow Cytometry

A cytochemical method was developed to differentially stain cellular DNA, RNA, and proteins with fluorochromes Hoechst 33342, pyronin Y, and fluorescein isothiocyanate, respectively. The fluorescence intensities, reflecting the DNA, RNA, and protein content of individual cells, were measured in a flow cytometer after sequential excitation by three lasers tuned to different excitation wavelengths. The method offers rapid analysis of changes in the cellular content of RNA and protein as well as in the RNA-protein, RNA-DNA, and protein-DNA ratios in relation to cell cycle position for large cell populations. An analysis of cycling cell populations (exponentially growing CHO cultures) and noncycling CHO cells arrested in the G1 phase by growth in isoleucine-free medium demonstrated the potential of the technique.

RNA-DNA ratio: an index of larval fish growth in the sea

Data on water temperature, RNA-DNA ratio, and growth of eight species of temperate marine fish larvae reared in the laboratory were fit to the equation: $$G_{pi} = 0.93{\text{ }}\operatorname{T} + 4.75{\text{ RNA - DNA}} - 18.18$$ where Gpi is the protein growth rate in % d-1 and T is the water temperature. Water temperature and larval RNA-DNA ratio explained 92% of the variability in growth rate of laboratory-reared larvae. The model is useful over the entire range of feeding levels (starvation to excess), temperatures (2° to 20°C) and fish species studied. Estimates of recent growth of larval cod, haddock, and sand lance caught at sea based on water temperature and RNA-DNA ratio ranged from negative to 26% d-1. These data demonstrate the importance of food availability in larval fish mortality and suggest that short-term growth under favorable conditions may be considerably higher than expected from long-term indicators. RNA-DNA ratio analysis offers new possibilities for understanding larval growth and mortality, and their relation to environmental variability.

The influence of the ratio of protein energy to total energy in the feed on the activity of protein synthesis in vitro, the level of ribosomal RNA and the RNA-DNA ratio in white trunk muscle of atlantic cod ( Gadus morhua)

1. 1. Cod ( Gadus morhua) were fed diets containing protein energy to total energy levels (PE/TE) of 10.0, 20.6, 29.6, 38.4, 56.2 and 74.1% for 21 days. 2. 2. Ribosomes were isolated from the white trunk muscle tissue, the capacity for protein synthesis in vitro determined and related to muscle tissue wet weight, rRNA and DNA. 3. 3. Protein concentrations of less than 47.4% PE/TE in the diets reduce the ribosomal capacity for protein synthesis per g wet weight and per mg DNA, and the tissue contents of rRNA and ratio of rRNA/DNA. 4. 4. The capacity for muscle protein synthesis in vitro is a significant and sensitive parameter of protein inadequacy in fish diets.

Effects of temperature and food availability on growth, survival, and RNA-DNA ratio of larval sand lance (Ammodytes americanus)

Effects of temperature and food availability on growth, survival, and RNA-DNA ratio of larval sand lance (Ammodytes americanus)

Effects of steroids and steroid antagonists on growth, gonadal development, and [formula omitted] ratios in juvenile steelhead trout

Steroids incroporated into Oregon Moist Pellets were fed to steelhead trout ( Salmo gairdneri) to determine if growth could be enhanced. In 1976, steelhead trout were treated with 17 α-methyltestosterone, diethylstilbesterol, diethylstilbesterol plus clomiphene citrate, 17 α-methyltestosterone plus flutamide (4′-nitro-3′ trifluoromethyliosbutranilide), or flutamide from the onset of exogenous feeding. In 1977, steelhead trout were started on treated diets 1 month after the onset of exogenous feeding and received testosterone propionate, testolactone, methylandrostenolone, 17 α-methyltestosterone, flutamide, estradiol, or estradiol plus progesterone. Fish treated with flutamide (20 μg/g food) or diethylstilbesterol (5 μg/g) plus clomiphene citrate (15 μg/g) from the onset of exogenous feeding showed an increase in weight compared with controls. The androgens and other estrogens tested did not enhance the growth of steelhead trout. All compounds (except flutamide) fed to the fish from the onset of feeding caused abnormal sex ratios. Androgens induced more males while estrogens induced more females. 17 α-methyltestosterone (1, 5, 15, or 35 μg/g), 17 α-methyltestosterone (5 μg/g) plus flutamide (20 μg/g), testolactone (10 μg/g), and methylandrostenolone (10 μg/g) advanced spermatogenesis. 17 α-methyltestosterone (2 or 35 μg/g), testolactone (10 μg/g), methylandrostenolone (10 μg/g), or flutamide (20 μg/g) caused hypertrophied granulosa cells or atretic oocytes in some ovaries. Intersexes containing both testicular and ovarian tissue in the gonads were found in some fish treated with 17 α-methyltestosterone (5 μg/g), 17 α-methyltestosterone (5 μg/g) plus diethylstilbesterol (5 μg/g), 17 α-methyltestosterone (5 μg/g) plus flutamide (20 μg/g), or diethylstiblesterol (5 μg/g) plus clomiphene citrate (5 μg/g). 17 α-methyltestosterone at 1, 5, 15, or 35 μg/g caused significant linear dose-dependent increases in epidermal thickness. There were no differences in the RNA DNA ratio between treatments. We conclude from these studies that steelhead trout do not respond positively in terms of growth enhancement from steroid treatment as has been shown in other salmonids. However, the androgen antagonist, flutamide, may be a potential growth enhancer in steelhead trout and may also be used as an antiandrogenic compound when administered with an androgen.

Protein synthesis in the whole body, liver, skeletal muscle and kidney cortex of lambs infected by the nematode Trichostrongylus colubriformis

Jones W. O. and Symons L. E. A. 1982. Protein synthesis in the whole body, liver, skeletal muscle and kidney cortex of lambs infected by the nematode Trichostrongylus colubriformis. International Journal for Parasitology 12: 295–301. Tyrosine flux and the synthesis of protein in the whole body, liver, skeletal muscle and kidney cortex and of albumin in lambs infected with Trichostrongylus colubriformis and uninfected lambs fed ad libitum or pair-fed with the infected group, were measured by constant infusion of 14C- l-tyrosine. Live weight gain was lower in the infected than in pairfed lambs, but rates of whole body protein synthesis were similar in both groups. On the other hand, compared with control lambs, there was a faster rate of protein synthesis per unit of protein consumed in infected but not in pair-fed lambs. Rates of protein synthesis per unit of body weight in infected were higher than in pair-fed lambs, but similar to the rate in control lambs. The fractional synthetic rates (FSR) of albumin and liver proteins and the amount of liver protein synthesized per day were increased by infection. The FSR and amount of protein synthesized per day were depressed in skeletal muscle and kidney cortex. Anorexia did not explain any of these changes. Infection caused a loss of protein from each of these tissues, but this loss was due to anorexia in only the liver. There was generally good correlation between concentration of RNA per g fresh weight or per mg nitrogen and the FSR of protein. However, although the RNA DNA ratio correlated well with synthesis in skeletal muscle, it was poorly correlated for liver proteins. The relationship between the rate of growth and protein synthesis in infected lambs is discussed.

Seasonal variations in RNA–DNA ratios and in indicators of feeding, reproduction, energy storage, and condition in a population of bluegill, Lepomis macrochirus Rafinesque

Liver and epaxial muscle RNA concentrations and RNA‐DNA ratios (RNA/DNA) of bluegill Lepomis macrochirus from a central Tennessee lake were maximum in the spring, low during the summer, high again in the fall, and low during the winter. Liver‐somatic indexes and DNA concentrations indicated that liver cell volume and energy storage varied seasonally and were lowest during late spring and summer. Seasonal variations in gonosomatic indexes were typical of this species, and RNA/DNA decreased as gonads matured during the spring. RNA/DNA and energy storage in the liver decreased during the major spawning season. Seasonal variations in food consumption may have effected seasonal growth and energy storage. However, a summer depression in RNA/DNA may have been more closely associated with the effects of thermal stratification and dissolved oxygen stress.

Subcellular compartmentalization of estrogen receptors in human breast cancer cells

In MCF-7 human breast cancer cells about 75% of the cell's total estrogen receptor sites were unexpectedly found in nuclei not bound with estrogens [13]. These findings were based on studies with crude nuclei and it was originally proposed that these unfilled nuclear estrogen receptors might be biologically active in the absence of estrogen [23]. In order to examine the subcellular distribution of receptor more critically, we have prepared highly purified MCF-7 nuclei, assessed their purity by electron microscopy RNA DNA and DNA protein ratios, and compared their estrogen receptor content with less pure subcellular fractions. In estrogen-withdrawn cells the percentage of the total unfilled receptors found in nuclear fractions (Rn) decreased dramatically from 30% (1.81 pmol/mg DNA) in the crudest preparation to only 3% (0.19 pmol/mg DNA) in highly purified nuclei. (This decrease is not due to an irreversible loss of receptor sites, as receptors lost from nuclei were recovered in the cytosol.) By contrast, after 1 h incubation in vivo with estradiol, the number of estrogen-filled receptor sites (RnE) found in the crudest nuclear preparation (3.28 pmol/mg DNA) and in the most highly purified nuclei (2.66 pmol/mg DNA) were nearly the same. It is very likely that the high level of unfilled receptors found in crude nuclei is a contamination of the preparation with cytoplasmic receptor. 1. 1. Electron microscopy shows gross contamination of crude nuclei with whole cells and cytoplasmic attachments. Purification of nuclei removes attaching cytoplasm which is parallelled by a dramatic decrease in unfilled receptor sites contained in the nuclear preparations. 2. 2. Multiple extractions with Tris buffer (without KCl) releases as much as 72% of the receptor contained in crude nuclei and treatment with 0.2% Triton X-100 extracts more than half. 3. 3. Tris-extractable receptor from crude nuclei sediments identically with cytosol receptor as a single 8S peak on low salt sucrose gradients. 4. 4. Unfilled receptor does not bind in vitro to purified nuclei. Although we cannot entirely exclude the possibility that unfilled receptors are nuclear in origin but may leak from nuclei during cell fractionation, these results are consistent with the localization of unfilled receptor largely in the cytoplasm of MCF-7 cells, as found in normal target tissues such as the rat uterus.

Seasonal patterns of muscle RNA-DNA ratio and growth in black crappie, Pomoxis nigromaculatus

Black crappie (Pomoxis nigromaculatus) were collected weekly from a natural lake during the period mid-April to mid-September. The fish were weighed, state of maturity determined and RNA-DNA ratio of white muscle was measured. Water temperature and primary production were measured in the lake.

Relationships Between RNA–DNA Ratio, Prey Density, and Growth Rate in Atlantic Cod (Gadus morhua) Larvae

The protein, DNA, and RNA content of larvae maintained at 1.0 plankter/mL increased at the rates of 9.3, 9.9, and 9.8% per day, respectively, for the 5 wk after hatching. Protein reserves of larvae held at 0 or 0.2 plankters/mL were depleted by 45 and 35%, respectively, prior to death 12–13 d after hatching. Starved larvae had similar protein concentrations (percent of dry weight), lower RNA concentrations, and higher DNA concentrations than fed larvae. Larvae held at higher plankton densities had higher RNA–DNA ratios and faster growth rates than larvae held at lower plankton densities. The RNA–DNA ratio was significantly correlated (P < 0.01) with the protein growth rate. The RNA–DNA ratio appears to be a useful index of nutritional status in larval Atlantic cod (Gadus morhua) and may be useful for determining if cod larvae were in a period of rapid or slow growth at the time of capture. Key words: RNA–DNA ratio, starvation, protein, nucleic acids, growth, larval fish, Atlantic cod

Effects of trace metals on growth of yellow perch (Perca flavescens) as measured by RNA-DNA ratios

Relationships between sublethal concentrations of cadmium and zinc in natural water and metal uptake by and growth of fish were investigated. RNA-DNA ratios and weight gain were used to assess seasonal growth differences between yellow perch populations from contaminated and control sites. Whole-body concentrations of cadmium and zinc in young-of-the-year perch (Perca flavescens) were significantly different between sites. Measurable growth differences did occur and were significantly correlated with cadmium levels. Growth differences that were prominent during mid-summer were reduced by late summer. RNA-DNA ratios were sensitive indicators of fish growth.

Effeects of hypophysectomy and beef growth hormone replacement therapy, over time and at various dosages, on body weight and total RNA-DNA levels in the black bullhead ( Ictalurus melas)

Experiments were conducted on the bullhead ( Ictalurus melas) at 24°: first, to define the effects of hypophysectomy and beef growth hormone (bGH) replacement therapy, over time, on body weight and the concentrations and ratios of RNA and DNA in skeletal muscle and liver; second, to evaluate tissue RNA concentrations and RNA-DNA ratios as biochemical indicators of growth; and third, to assess the effect of various bTH doses on the hypophysectomized bullhead. Hypophysectomy arrested growth in weight by the 12th day of a 26-day experiment. At the same time, the concentration of RNA and the RNA DNA ratio in muscle were reduced relative to the levels seen in sham-operated controls. In the latter, muscle RNA concentration, after an initial increase, remained constant throughout the experiment, whereas muscle RNA DNA rose progressively, due to a decline in the concentration of DNA. The livers of hypophysectomized bullheads had lower RNA and RNA DNA levels than the livers of controls by the fourth day after surgery. In animals that had been hypophysectomized for 2 weeks, bGH (at 40 mIU/fish, every second day) stimulated significant increases in body weight and hepatic RNA concentration within 4 days, after only two injections. Muscle RNA and RNA DNA levels were also elevated by bGH injection, but the effects were not statistically meaningful until the sixth day of treatment. After 8 days, linear relationships were demonstrated for percentage gain in body weight and muscle RNA concentration, each plotted against the logarithm of bGH dose (range tested, 5.7 to 320 mIU/fish, every second day). Collectively, the data indicate that GH has a controlling influence over nucleic acid metabolism in the bullhead; that hypophysectomy and GH replacement therapy can exert their effects rather quickly in fish; and that biochemical as well as morphometric responses can probably be used in assaying for teleost GHs.

Isolation and purification of myotube and myoblast nuclei from cultures of embryonic chick skeletal muscle

Abstract Methods are described for the preparation of purified myotubes from embryonic chick skeletal muscle cultures and the preparation of purified nuclei from both myotubes and myoblasts. Myotubes are released from the culture dish by digestion of their collagen substratum with collagenase, and purified by sucrose density gradient sedimentation. Nuclei are prepared from the isolated myotubes by controlled homogenization in Ca2+-free medium and sedimentation through 2.1 M sucrose. Nuclei are prepared from cultured myoblasts in a similar fashion, with the inclusion of the non-ionic detergent NP-40 in the homogenization medium and sedimentation through 2.4 M sucrose. Phase contrast microscopic examination showed that the nuclear preparations are free of visible cytoplasmic contamination, and are morphologically similar to nuclei observed in situ. Biochemical assays (protein/DNA and RNA DNA ratios) confirm the purity of the nuclear preparations. Both nuclear preparations have been used to prepare purified chromatin which has spectral and chemical properties similar to those reported for chromatin purified directly from several chick tissues.

Comparisons of Two Bluegill Populations by Means of the RNA-DNA Ratio and Liver-Somatic Index

Abstract Bluegills (Lepomis macrochirus Rafinesque) from two central Tennessee lakes revealed monthly and seasonal variations of liver RNA-DNA ratios (liver RNA/DNA) and liver-somatic indexes (LSI's) that were associated with gonad maturation, spawning, and temperature. Liver RNA/DNA indicated a longer summer growth depression in fish collected from the smaller, shallower lake. Although LSI's were consistently higher in the smaller, shallower lake, liver RNA/DNA and scale analyses showed faster growth rate in the larger, deeper lake. Thus the LSI may be a useful indicator of relative food intake-energy storage levels within a single lake, but it does not necessarily reflect absolute growth rate of the fish. Liver RNA/DNA provided an indication of immediate or short-term growth rates.

Isolation and characterization of rDNA-containing chromatin from nucleoli

Fractions corresponding to ‘euchromatin’ and ‘heterochromatin’ were isolated from Novikoff hepatoma nucleoli by a modification of the DNase II method of Gottesfeld et al. [18, 19]. Nucleolar chromatin [36] was digested with DNase II, and ‘euchromatin’ was obtained as a soluble fraction following differential precipitation of ‘heterochromatin’ with Mg 2+. In this ‘euchromatin’ fraction, the rDNA was enriched approx. 8.5-fold over the unfractionated nucleolar chromatin, as determined by DNA-RNA hybridization. On 8–80% glycerol gradients, the ‘euchromatin’ separated into three discrete subfractions as with euchromatin obtained from the bulk chromatin [18, 19]. Two subfractions which sedimented at 14S and 20S, respectively, were consistently found after DNase II digestion (to 15 min) which contained all histone fractions. All euchromatin subfractions were enriched in rDNA and had a high protein/DNA ratio, a high RNA-DNA ratio, and a high template activity with exogenous RNA polymerase I. Fraction B with a sedimentation of 14S was most enriched in the parameters related to rRNA synthesis. The Mg 2+-precipitable fractions reported to be ‘heterochromatin’ [18, 19] not only contained substantial amounts of ribosomal genes, but also had higher template activities than unsheared chromatin. These results imply (1) nucleolar ‘euchromatin’ has a structural heterogeneity similar to that of the euchromatin containing active single copy sequences; (2) at least a portion of rDNA in ‘euchromatin’ is protected by proteins, including histones, which does not preclude active transcription of rDNA; (3) none of the currently used criteria for ‘euchromatin’ alone defines the chromatin containing rDNA; and (4) an appreciable enrichment of rDNA occurs in isolated euchromatin but not all the rDNA was separated into ‘euchromatin’ by the DNase II method.

A comparison of certain aspects of nitrogen metabolism in non-diapausing and diapausing boll weevils, Anthonomus grandis

Both DNA and RNA were lower in males and females of a diapausing strain of boll weevils, Anthonomus grandis 10 to 15 days after emergence than in newly-emerged insects of the same strain. Protein content remained relatively constant in the strain of diapausing weevils, but increased substantially in the 10 to 15-day boll weevils of a non-diapausing strain; the relationship of lipoprotein to protein generally remained about the same in both strains for weevils of different ages of the same strain. Since the RNA-DNA ratios differed materially only in the 10 to 15-day diapausing males (2.2), it might be used as a corroborative indicator of diapause. The hypothesis that the boll weevil stores nitrogen as uric acid during diapause was disproven; rather the amount of stored uric acid diminished at the onset of diapause.

Isolation and characterization of different-sized nucleoli from Ehrlich ascites tumor cells: Evidence of different nucleolar ribosomal cistrons

A procedure was developed for isolation of variously sized nucleoli in order to study the mechanism of nucleolar formation from multiple nucleolar organizers and to compare the compositions of different-sized nucleoli from Ehrlich ascites tumor cells. Relatively small nucleoli and large nucleoli from Ehrlich ascites tumor cells were separated by centrifugation at 400 g for 5 min in a layer of 0.34 M sucrose over 0.88 M sucrose. Small nucleoli remained in the 0.34 M sucrose layer while the large nucleoli accumulated in the 0.88 M sucrose. Three fractions, provisionally named small, intermediate and large nucleoli, containing 0.33, 0.41 and 0.84 pg DNA/nucleolus, respectively, were separated. Unfractionated nucleoli contained 0.59 pg DNA/nucleolus. The RNA content also increased with the size of the nucleolus and no significant difference was observed in the RNA DNA ratios in the three fractions. Large nucleoli incorporated more [ 3H]uridine and [ 32P]orthophosphate into RNA than did small nucleoli, but the base compositions of the RNAs extracted from the different-sized nucleoli were similar. No significant fragmentation occurred on sonication of large nucleoli for 3 min, so the observed difference in the DNA contents was not due to mechanical damage of the nucleoli. The DNAs of these different-sized nucleoli were analysed on CsCl gradients. The nucleoli contained similar percentages of satellite DNA (20–22%) which were also similar to those of total, unfractionated nucleoli. Approx. 10% of the extranucleolar DNA is satellite DNA—thus the nucleolar fractions were probably not appreciably contaminated with extranucleolar DNA. The DNA of small nucleoli contained a slightly lower percentage (0.058%) of ribosomal cistrons than large nucleoli (0.081%). This means that the higher content of DNA in the large nucleoli is not merely due to longer sized chromatin with extra regions of the vicinity of nucleolar organizers. Thus these results suggest that the total content of ribosomal cistrons/nucleolus is roughly proportional to the DNA content of the nucleoli, at least in Ehrlich ascites tumor cells. Namely, the number of ribosomal cistrons per nucleolus for small, intermediate and large nucleoli is 40, 60 and 130, respectively.

Nucleus-associated steroid 5α-reductase activity and androgen responsiveness. a study in various organs and brain regions of rats

1. 1. Purified nuclei were prepared from various organs and brain regions of rats. The preparations were assayed for 5α-reductase activity, and checked for purity by measurement of RNA-DNA ratios. Although prostatic nuclei showed the highest 5α-reductase activity, the other nuclear fractions were also able to reduce testosterone into its active metabolite: 17β-hydroxy-5α-androstan-3-one. In all tissues, however, the nucleus-associated 5α-reductase activity was inversely related to the purity of the nuclear fractions. 2. 2. Prostate, kidney, submaxillary gland, lung and midbrain were completely fractionated. Total 5α-reductase activity varied as follows: lung > prostate > kidney > submaxillary gland > midbrain. The particulate fractions were the major sites of 5α-reductase activity in all tissues except for the submaxillary gland. The nuclear fraction reached 19% of the total 5α-reductase activity in the kidney and 25% in the prostate. 3. 3. The low apparent K m of the 5α-reductase for testosterone reported in prostatic nuclei was also found in lung nuclei (1.56 μM) and kidney nuclei (0.96 μM). 4. 4. It is concluded that the androgen responsiveness of a particular tissue cannot be accounted for by any of the parameters of 5α-reductase activity studied: the occurrence of an enzyme with particularly high affinity for testosterone, the total cellular activity, or the absolute or relative amount of this activity associated with purified nuclei.

A comparison of RNA metabolism in the visual cortex of sighted rats and rats with retinal degeneration

The RNA metabolism in the visual cortex of visually stimulated adult blind and sighted rats has been compared. The blind rats belonged to a strain suffering from an inherited retinal dystrophy. This condition renders them blind after the age of 3–4 weeks. There is no gross difference in the metabolic rate of the blind and sighted strains. No significant differences were seen in the RNA, DNA, and protein content per wet weight of the two types of visual cortex but the RNA DNA ratio was significantly lower in the blind animals. Intraventricularly administered ( 14C) orotic acid was incorporated into RNA to a lesser extent in the visual cortex of the blind animals. This could not be wholly attributed to a difference in the size of the acid soluble pool. Chromatographic studies on RNA hydrolysates and the acid soluble precursor pool indicated that the reduced labeling of RNA in the blind visual cortex was not due to a reduction in the rate of conversion of orotic acid to UTP and thus it would appear that the lowered rate of incorporation of label into RNA is due to a reduced rate of RNA synthesis. This was restricted to the visual cortex, no differences were observed in the frontal cortex. In vitro studies indicated that there were no differences in the total ribonuclease activity in the blind and sighted visual cortex but the RNA polymerase activity (but not the Poly C synthetase activity) was slightly lower in the visual cortex of the blind animals.