Characterisation of the storage protein subunits synthesised in vitro by polyribosomes and RNA from developing pea (Pisum sativum L.)

Evidence is presented to show that legumin, the major storage protein in Pisum, is synthesised in vitro by the wheat germ and reticulocyte lysate systems, from polyribosomes and mRNA isolated from developing pea seeds. While legumin isolated from mature pea seeds consists of 40,000 and 20,000 MW subunits, the in vitro legumin is synthesised as a 60,000 MW precursor consisting of covalently linked 40,000 and 20,000 MW subunits. The implications of these findings are discussed in relationship to studies with other systems.Abbreviations SDS-PAGE SDS-polyacrylamide gel electrophoresis - PBS phosphate buffered saline - MW molecular weight     

Head regeneration inHydra is initiated release of head activator and inhibitor

Summary Hydra regenerating heads release at least two substances into the surrounding medium: one stimulates and one inhibits head formation. The inhibitor is released mainly during the first hour after cutting, the activator is released more slowly with a maximum in the second hour and with substantial release still during the following six hours. The release of both substances seems to be specific for head regeneration: it is not found in animals regenerating feet. The sequential release of these substances leads to the early changes observed at the cellular level during head regeneration inhydra: the inhibitor produces a decrease, the activator an increase in the mitotic activity of interstitial and epithelial cells, if assayed on intact animals. Head regeneration is blocked, if the release of the head activator is prevented. It is therefore suggested that these substances are necessary to initiate head regeneration inhydra.     

A procedure for the isolation and determination of deoxyribose derivatives

A method of determining deoxyribose derivatives in biological material is described. It has high sensitivity, and is particularly useful in that it can be applied to a large range of tissues for which the other available assays are unsuitable. This is because the method is applicable to complex mixtures of nucleotides in which such substances as ribonucleotides are present in very large excess over deoxyribonucleotides, and it is not necessary to equilibrate the nucleotide-precursor pool with radioactive phosphate. The method has mainly been developed with the object of determining deoxyribonucleoside triphosphates, but it can be used to assay ribonucleoside triphosphates, as well as the mono- and diphosphates of both types of nucleoside. The procedure used involves three basic techniques: (1) periodate oxidation and methylamine-induced cleavage of the sugar ring to destroy 2'- and 3'-unsubstituted ribonucleosides; (2) column chromatography to separate the deoxyribonucleotides from each other and from other substances, such as the products of the periodate oxidation; (3) fluorimetric determination of deoxyribose after labilization of the pyrimidine-glycosidic bond by bromination of the heterocyclic ring. Each of these three procedures can be used independently, in conjunction with other analytical procedures.     

Effects of Reproductive Status on Energy Intake,Ingestion Rates,and Dietary Composition of Female <Emphasis Type="BoldItalic">Cebus capucinus</Emphasis> at Santa Rosa,Costa Rica

The limiting factor for fitness in female primates is the acquisition of high-quality food, i.e., food that is high in energy and nutrients, such as protein, fat, and carbohydrates. Reproductive status can influence female feeding patterns, e.g., lactating females in some primate species consume greater quantities of food and specific nutrients than do nonlactating females. We examined the energy intake, ingestion rate, and composition of the diet in female white-faced capuchins in 3 reproductive states: lactating, gestating, and cycling. We observed 12 reproductively active females for 7 mo and compared their energy intake, ingestion rates, and intake rates of nutrients: protein, fat, sugar, and fiber. Lactating females took in significantly more energy per hour while feeding than pregnant and cycling females did. Lactating females also ingested significantly more food items per hour, but they did not spend more time eating than other females did, and they did not differ in the composition of their diet as measured by insect consumption and proportion of protein. Pregnant and cycling females did not differ from each other in any of the measures. We predicted that as the infants aged and began to move independently to forage and play, their mothers’ energy intake rates would decrease in concert with decreased energy demands by the infant. We found a statistically significant relationship between these 2 factors, with infant age explaining 75.4% of variation in mothers’ energy intake, supporting previous studies that found lactation to be the most energetically expensive reproductive state.     

Hydrophobic substances induce water stress in microbial cells

Ubiquitous noxious hydrophobic substances, such as hydrocarbons, pesticides and diverse industrial chemicals, stress biological systems and thereby affect their ability to mediate biosphere functions like element and energy cycling vital to biosphere health. Such chemically diverse compounds may have distinct toxic activities for cellular systems; they may also share a common mechanism of stress induction mediated by their hydrophobicity. We hypothesized that the stressful effects of, and cellular adaptations to, hydrophobic stressors operate at the level of water : macromolecule interactions. Here, we present evidence that: (i) hydrocarbons reduce structural interactions within and between cellular macromolecules, (ii) organic compatible solutes – metabolites that protect against osmotic and chaotrope-induced stresses – ameliorate this effect, (iii) toxic hydrophobic substances induce a potent form of water stress in macromolecular and cellular systems, and (iv) the stress mechanism of, and cellular responses to, hydrophobic substances are remarkably similar to those associated with chaotrope-induced water stress. These findings suggest that it may be possible to devise new interventions for microbial processes in both natural environments and industrial reactors to expand microbial tolerance of hydrophobic substances, and hence the biotic windows for such processes.     

Diurnal Cycling in Root Resistance to Water Movement

The occurrence of diurnal changes in root resistance of cotton was studied by measuring the flow of water through 35-to70-day-old root systems under a pressure of 3.10 bars or a vacuum of 0.88 bar. The volume of exudate obtained under constant pressure or constant vacuum was 2 to 3 times greater near midday than near midnight indicating that the root resistance apparently was 2 to 3 times greater at night than during the day. The salt concentration of the exudate also cycled; the concentration was lowest at midday and highest at night, hence there was little diurnal variation in the total amount of salt moved per hour. The cycle for volume of exduate, salt concentration, and apparent root resistance had a period of 22 to 26 hours at 24°C. The cycle gradually died away 2 to 3 days after removal of the shoots. The diurnal variations appeared to be controlled by signals from the shoots because the phase of the cycles could be reset by changing the light-dark cycle under which the plants were grown. Cycling was eliminated by exposure to 8 or more days of continuous light before removing the shoots, and cycling could not be entrained by a 6 hour light-6hour dark cycle. Bubbling nitrogen gas through the nutrient medium stopped cycling. A possible role of ion or growth regulator action is discussed.     

环介导等温扩增核酸技术及其在食品安全检测领域的应用

环介导等温扩增(Loop-mediated isothermal amplification,简称LAMP)是利用能识别靶序列上6个位点的4个特殊设计的引物和一种具有链置换活性的DNA聚合酶,在恒温条件下,特异、高效、快速地扩增核酸的新技术。该技术在1h内扩增效率可达到109-1010个数量级,扩增产物是一系列反向重复的靶序列构成的茎环结构和多环花椰菜样结构的DNA片段混合物,电泳后在凝胶上显现出由不同大小的区带组成的阶梯式图谱。近年来LAMP技术以其特异性强、等温灵敏、操作简单、产物易检测等优点已经应用于食品安全检测领域的多个方面。     

Bacterial mutagenicity testing of 49 food ingredients gives very few positive results

49 substances permitted for use in food in the United States was tested for mutagenicity in the Ames Salmonella typhimurium assay and in Escherichia coli strain WP2. Four of these substances caused increases in revertant counts in S. typhimurium. Two of these four (papain and pepsin) were found to contain histidine, and therefore the results of the tests on these two substances could not be taken as demonstrating mutagenicity. The other two substances causing increases in revertant counts (hydrogen peroxide and potassium nitrite) were mutagenic. The results on one chemical, beta-carotene, were evaluated as inconclusive or questionable. The remaining 44 substances were nonmutagenic in the test systems used. It is concluded that, for those generally physiologically innocuous chemicals tested, there are very few 'false positives' in the bacterial test systems used.     

How do plant growth substances work?

Abstract. Despite intensive research effort, the role of growth substances in the life of the intact growing plant is far from clear. Two reasons are suggested which may account for the lack of progress. The first is the failure to recognize the unique aspects of plant development. These which are expressed by the regenerative, organizational and developmental plasticity of the meristem probably result from the possession of growth substances. The second is the concept of growth substances as hormones. This represents the main conceptual thrust of research and is considered critically, starting with the historical system, the coleoptile and dealing with other major growth substance systems in turn. It is concluded that a hormonal concept which includes control by changes in growth substance concentration fails to explain the developmental phenomena under examination. A role for growth substances as integrating agents is suggested and the notion of quantitative tissue sensitivity variation is developed to explain the major growth patterns of developing shoots.     

Enzymes as versatile labels and signal amplifiers for monitoring immunochemical reactions

Summary Enzymes have proven to be sensitive and versatile labels for immunochemical assays. The sensitivity of an enzyme label stems from its extraordinary catalytic power which in turn provides a great amplification of signals. Its versatility, however, stems from the fact that enzyme activity can be modulated by a number of substances.Enzyme labeled immunoassays can be divided into two categories: (a) heterogeneous and (b) homogeneous (non-separation). In the heterogeneous systems, the quantitation of the antibody bound and unbound fractions requires a physical separation of these two fractions, whereas the homogeneous or non-separation systems do not require such a separation. In the homogeneous systems, the unbound and antibody bound fractions can be distinguished functionally.A total of 11 unique principles used in the development of enzyme labeled immunoassays are described. The advantages, disadvantages and limitations of them are considered, as well as the future paths for research and developments.     

A Drosophila protein-interaction map centered on cell-cycle regulators

Background  

Maps depicting binary interactions between proteins can be powerful starting points for understanding biological systems. A proven technology for generating such maps is high-throughput yeast two-hybrid screening. In the most extensive screen to date, a Gal4-based two-hybrid system was used recently to detect over 20,000 interactions among Drosophila proteins. Although these data are a valuable resource for insights into protein networks, they cover only a fraction of the expected number of interactions.     

Major environmental drivers determining life and death of cold-water corals through time

Cold-water corals (CWCs) are the engineers of complex ecosystems forming unique biodiversity hotspots in the deep sea. They are expected to suffer dramatically from future environmental changes in the oceans such as ocean warming, food depletion, deoxygenation, and acidification. However, over the last decades of intense deep-sea research, no extinction event of a CWC ecosystem is documented, leaving quite some uncertainty on their sensitivity to these environmental parameters. Paleoceanographic reconstructions offer the opportunity to align the on- and offsets of CWC proliferation to environmental parameters. Here, we present the synthesis of 6 case studies from the North Atlantic Ocean and the Mediterranean Sea, revealing that food supply controlled by export production and turbulent hydrodynamics at the seabed exerted the strongest impact on coral vitality during the past 20,000 years, whereas locally low oxygen concentrations in the bottom water can act as an additional relevant stressor. The fate of CWCs in a changing ocean will largely depend on how these oceanographic processes will be modulated. Future ocean deoxygenation may be compensated regionally where the food delivery and food quality are optimal.

Cold-water corals are the engineers of complex ecosystems forming unique biodiversity hot spots in the deep sea, but there is substantial uncertainty regarding their sensitivity to environmental parameters. A synthesis of six paleoceanographic reconstructions reveals that food supply exerted the strongest impact on coral vitality during the past 20,000 years, whereas locally low oxygen concentrations can act as a stressor.     

Cycling of sensitivity to physical dormancy-break in seeds of Ipomoea lacunosa (Convolvulaceae) and ecological significance

BACKGROUND AND AIMS: Although a claim has been made that dormancy cycling occurs in seeds of Ipomoea lacunosa (Convolvulaceae) with physical dormancy, this would seem to be impossible since the water gap cannot be closed again after it opens (dormancy break). On the other hand, changes in sensitivity (sensitive <--> non-sensitive) to dormancy-breaking factors have been reported in seeds of Fabaceae with physical dormancy. The primary aim of the present study was to determine if sensitivity cycling also occurs in physically dormant seeds of I. lacunosa. METHODS: Treatments simulating conditions in the natural habitat of I. lacunosa were used to break seed dormancy. Storage of seeds at temperatures simulating those in spring, summer, autumn and winter were tested for their effect on sensitivity change. Seeds made non-dormant were stored dry in different temperature regimes to test for dormancy cycling. In addition, seeds collected on different dates (i.e. matured under different climatic conditions) were used to test for maternal effects on sensitivity to dormancy-breaking factors. KEY RESULTS: Sensitivity was induced by storing seeds under wet conditions and reversed by storing them under dry conditions at low (< or = 5 degrees C) or high (> or = 30 degrees C) temperatures, demonstrating that seeds of I. lacunosa can cycle between sensitive and insensitive states. Sensitive seeds required > or = 2 h at 35 degrees C on moist sand for release of dormancy. However, there is no evidence to support dormancy cycling per se. Conceptual models are proposed for sensitivity cycling and germination phenology of I. lacunosa in the field. CONCLUSIONS: Seasonal germination behaviour of physically dormant I. lacunosa seeds can be explained by sensitivity cycling but not by dormancy cycling per se. Convolvulaceae is only the second of 16 families known to contain species with physical dormancy for which sensitivity cycling has been demonstrated.     

Effect of some hormones on the rate of the triacylglycerol/fatty-acid substrate cycle in adipose tissue of the mouse in vivo

A method is described for the measurement of the rate of the triacylglycerol/fatty-acid cycle in adipose tissue of the mouse in vivo, which depends upon the incor-poration of tritium from [³H]H₂O into the glycerol and fatty-acid moieties of triacyiglycerol. The rate of the cycling is increased two-fold by feeding, an effect that is completely abolished by the β-adrenergic blocker propranolol. The β-adrenergic agonist fenoterol increased the rate of cycling five-fold in white adipose tissue and three-fold in brown adipose tissue. Cold exposure had no effect on the rate of cycling in white adipose tissue but increased the rate almost two-fold in brown adipose tissue. The increased rate of cycling during feeding, which may be due to increased sympathetic nervous activity, is consistent with the view that the role of cycling is to increase sensitivity of metabolic control systems when required.     

Cytotoxic Effects of 6-Hydroxydopamine,Merocyanine-540 and Related Compounds on Human Neuroblastoma-and Hematopoietic Stem Cells

6-Hydroxydopamine(6-OHDA) and Merocyanine-540(MC-540) have been used clinically for purging of neuroblastoma cells prior to autologous bone marrow transplantation. Both substances were found to be more toxic against neuroblastoma cells than against hematopoietic stem cells. The more pronounced cytotoxic effects of 6-OHDA against neuroblastoma cells were not caused by its selective uptake; the rapid autooxidation at physiological pH leads to the formation of H,O, already in the incubation medium. Cytotoxic effects were not detected in short-time test systems (4 hour chromium-51 release assay) but only after longer incubation periods. In contrast, MC-540 proved to be toxic almost equally in short- and long-time test systems. 4-Hydroxynonenal(4-HNE) that may be formed in the plasma membrane subsequently to photoactivation of MC-540 was only slightly more toxic to neuroblastoma cells than to hematopoietic cells. Although the use of 6-OHDA and MC-540 in bone marrow purging has some limitations, the sensitivity of neuroblastoma cells against reactive oxygen compounds may be exploited more generally for therapy of this tumor.     

The phenolic compounds in <Emphasis Type="Italic">Cladonia</Emphasis> lichens are not antimicrobial in soils

According to classic text books on lichen biology, the phenolic secondary chemicals in lichens have antibiotic effects on soil microorganisms and mycorrhizal fungi in ecosystems. However, the experimental evidence for this under natural conditions is still relatively scarce. We examined some of the assumptions behind the concept of antimicrobial effects of lichen secondary substances: (1) the secondary substances of Cladonia stellaris, usnic and perlatolic acids, are leached out from the lichens by rainwater; (2) these substances inhibit the microbial activity of soil, and; (3) since they are extremely resistant to microbial decomposition, the soil underneath a continuous lichen mat is enriched in usnic and perlatolic acids. Our results did not support any of these assumptions. The evidence for the antimicrobial activity of lichen secondary substances seems to be weak in comparison to other suggested functions such as light filtering and herbivore protection. We suggest that it is time to re-evaluate the evidence for the antimicrobial ecological role of lichen secondary substances in natural systems.     

Tracing sources and pathways of dissolved nitrate in forest and river ecosystems using high-resolution isotopic techniques: a review

Nitrogen inputs into stream and river ecosystems, and the factors influencing those inputs, are important for various ecological and environmental concerns. Reliable information on where and how nitrogen compounds flow into aquatic ecosystems is indispensable to understanding the nutrient status of these ecosystems. Such information should include the biogeochemical mechanisms and hydrological controls of nutrient leaching into rivers from terrestrial systems such as forests, agricultural fields, and urbanized areas. Advancements in stable isotopomer measurements over the past two decades have expanded the variety of target substances and the precision with which they can be investigated. The high-throughput microbial denitrifier method allows for simultaneous measurement of nitrogen and oxygen isotope ratios and can provide high-resolution spatiotemporal information on both nitrate sources and biogeochemical processes. Although advanced techniques of stable isotope analysis have been used extensively to detect sources and estimate the relative contributions of multi-source systems in various rivers, there are still new horizons in investigating nitrogen transformations. For example, stable isotopes of oxygen (¹⁸O and ¹⁷O) occurring in nitrate due to atmospheric deposition can be used as natural tracers for evaluating internal nitrogen cycling; these isotopes are distinct from the oxygen within microbially generated nitrate in soils and water bodies. Another future challenge is improved use of nitrous oxide isotopomers in evaluating the relative contributions of nitrification and denitrification. Such analysis could provide insight into the nitrogen transformation that occurs under redox conditions at the boundary between terrestrial and aquatic systems, where nitrification and denitrification often occur simultaneously in soil and aquatic environments.     

Large-scale assessment of the zebrafish embryo as a possible predictive model in toxicity testing

Background

In the drug discovery pipeline, safety pharmacology is a major issue. The zebrafish has been proposed as a model that can bridge the gap in this field between cell assays (which are cost-effective, but low in data content) and rodent assays (which are high in data content, but less cost-efficient). However, zebrafish assays are only likely to be useful if they can be shown to have high predictive power. We examined this issue by assaying 60 water-soluble compounds representing a range of chemical classes and toxicological mechanisms.

Methodology/Principal Findings

Over 20,000 wild-type zebrafish embryos (including controls) were cultured individually in defined buffer in 96-well plates. Embryos were exposed for a 96 hour period starting at 24 hours post fertilization. A logarithmic concentration series was used for range-finding, followed by a narrower geometric series for LC₅₀ determination. Zebrafish embryo LC₅₀ (log mmol/L), and published data on rodent LD₅₀ (log mmol/kg), were found to be strongly correlated (using Kendall''s rank correlation tau and Pearson''s product-moment correlation). The slope of the regression line for the full set of compounds was 0.73403. However, we found that the slope was strongly influenced by compound class. Thus, while most compounds had a similar toxicity level in both species, some compounds were markedly more toxic in zebrafish than in rodents, or vice versa.

Conclusions

For the substances examined here, in aggregate, the zebrafish embryo model has good predictivity for toxicity in rodents. However, the correlation between zebrafish and rodent toxicity varies considerably between individual compounds and compound class. We discuss the strengths and limitations of the zebrafish model in light of these findings.     

Tests with Daphnia magna: A new approach to prescreen toxicity of newly synthesized acetylcholinesterase reactivators

Reactivators of phosphorylated acetylcholinesterase (oximes) are substances used as a human antidotal therapy for organophosphate poisoning. The objective of our study was to examine if juveniles of freshwater microcrustacean Daphnia magna could be employed as test animals in early screen toxicity tests of those substances as a first step for further experiments with daphnids intoxicated by organophosphates. For this purpose, seven different oximes were investigated. It was found that toxicity of all tested oximes increased with time. Mono-quaternary oximes were approximately ten fold (EC₅₀, 14.9 mg.l^{? 1}) more toxic in 24 hour tests and five fold (EC₅₀ was 79.46 mg.l^{? 1}) more toxic in 48 hour tests than bis-quaternary oximes. Tests with daphnids were shown to be easy to carry out at low cost and provided valuable results which could be used as a starting point for further research.