Functional profiling of recombinant NS3 proteases from all four serotypes of dengue virus using tetrapeptide and octapeptide substrate libraries

Regulated proteolysis by the two-component NS2B/NS3 protease of dengue virus is essential for virus replication and the maturation of infectious virions. The functional similarity between the NS2B/NS3 proteases from the four genetically and antigenically distinct serotypes was addressed by characterizing the differences in their substrate specificity using tetrapeptide and octapeptide libraries in a positional scanning format, each containing 130,321 substrates. The proteases from different serotypes were shown to be functionally homologous based on the similarity of their substrate cleavage preferences. A strong preference for basic amino acid residues (Arg/Lys) at the P1 positions was observed, whereas the preferences for the P2-4 sites were in the order of Arg > Thr > Gln/Asn/Lys for P2, Lys > Arg > Asn for P3, and Nle > Leu > Lys > Xaa for P4. The prime site substrate specificity was for small and polar amino acids in P1' and P3'. In contrast, the P2' and P4' substrate positions showed minimal activity. The influence of the P2 and P3 amino acids on ground state binding and the P4 position for transition state stabilization was identified through single substrate kinetics with optimal and suboptimal substrate sequences. The specificities observed for dengue NS2B/NS3 have features in common with the physiological cleavage sites in the dengue polyprotein; however, all sites reveal previously unrecognized suboptimal sequences.     

Meta‐analysis on the potential for increasing nitrogen losses from intensifying tropical agriculture

Fertilized temperate croplands export large amounts of reactive nitrogen (N), which degrades water and air quality and contributes to climate change. Fertilizer use is poised to increase in the tropics, where widespread food insecurity persists and increased agricultural productivity will be needed, but much less is known about the potential consequences of increased tropical N fertilizer application. We conducted a meta‐analysis of tropical field studies of nitrate leaching, nitrous oxide emissions, nitric oxide emissions, and ammonia volatilization totaling more than 1,000 observations. We found that the relationship between N inputs and losses differed little between temperate and tropical croplands, although total nitric oxide losses were higher in the tropics. Among the potential drivers we studied, the N input rate controlled all N losses, but soil texture and water inputs also controlled hydrological N losses. Irrigated systems had significantly higher losses of ammonia, and pasture agroecosystems had higher nitric oxide losses. Tripling of fertilizer N inputs to tropical croplands from 50 to 150 kg N ha^?1 year^?1 would have substantial environmental implications and would lead to increases in nitrate leaching (+30%), nitrous oxide emissions (+30%), nitric oxide (+66%) emissions, and ammonia volatilization (+74%), bringing tropical agricultural nitrate, nitrous oxide, and ammonia losses in line with temperate losses and raising nitric oxide losses above them.     

Organic enrichment of sediments reduces arbuscular mycorrhizal fungi in oligotrophic lake plants

1. Arbuscular mycorrhizal fungi (AMF) commonly colonise isoetid species inhabiting oxygenated sediments in oligotrophic lakes but are usually absent in other submerged plants. We hypothesised that organic enrichment of oligotrophic lake sediments reduces AMF colonisation and hyphal growth because of sediment O₂ depletion and low carbon supply from stressed host plants. 2. We added organic matter to sediments inhabited by isoetids and measured pore‐water chemistry (dissolved O₂, inorganic carbon, Fe²⁺ and ), colonisation intensity of roots and hyphal density after 135 days of exposure. 3. Addition of organic matter reduced AMF colonisation of roots of both Lobelia dortmanna and Littorella uniflora, and high additions stressed the plants. Even small additions of organic matter almost stopped AMF colonisation of initially un‐colonised L. uniflora, though without reducing plant growth. Mean hyphal density in sediments was high (6 and 15 m cm^?3) and comparable with that in terrestrial soils (2–40 m cm^?3). Hyphal density was low in the upper 1 cm of isoetid sediments, high in the main root zone between 1 and 8 cm and positively related to root density. Hyphal surface area exceeded root surface area by 1.7–3.2 times. 4. We conclude that AMF efficiently colonise isoetids in oligotrophic sediments and form extensive hyphal networks. Small additions of organic matter to sediments induce sediment anoxia and reduce AMF colonisation of roots but cause no apparent plant stress. High organic addition induces night‐time anoxia in both the sediment and the plant tissue. Tissue anoxia reduces root growth and AMF colonisation, probably because of restricted translocation of nutrient ions and organic solutes between roots and leaves. Isoetids should rely on AMF for P uptake on nutrient‐poor mineral sediments but are capable of growing without AMF on organic sediments.     

Supraglottic airway pressure-flow relationships during oronasal airflow partitioning in dogs

Amis, T. C., N. O'Neill, T. Van der Touw, A. Tully, and A. Brancatisano. Supraglottic airway pressure-flow relationships during oronasal airflow partitioning in dogs. J. Appl.Physiol. 81(5): 1958-1964, 1996.We studiedpressure-flow relationships in the supraglottic airway of eight pronemouth-open anesthetized (intravenous chloralose or pentobarbitalsodium) crossbred dogs (weight 15-26 kg) during increasingrespiratory drive (CO₂administration; n = 4) and duringgraded-voltage electrical stimulation (SV;n = 4) of the soft palate muscles.During increased respiratory drive, inspiratory airflow occurred viaboth the nose (n) and mouth(m), with the ratio of n tom[%(n/m)]decreasing maximally from 16.0 ± 7.0 (SD) to 2.4 ± 1.6%(P < 0.05). Simultaneously, oralairway resistance at peak inspiratory flow decreased from 2.1 ± 1.0 to 0.4 ± 0.4 cmH₂O(P < 0.05), whereas nasal airway resistance did not change (14.4 ± 7.2 to 13.1 ± 5.4 cmH₂O;P = 0.29). Inspiratory pressure-flowplots of the oral airway were inversely curvilinear or more complex innature. Nasal pathway plots, however, demonstrated a positive linearrelationship in all animals (r = 0.87 ± 0.11; all P < 0.001). Duringelectrical stimulation of soft palate muscle contraction accompanied bygraded constant-inspiratory airflows of 45-385 ml/s through anisolated upper airway, %(n/m)decreased from 69 ± 50 to 10 ± 13% at a SV of 84 ± 3% ofmaximal SV (P < 0.001). At a SV of85 ± 1% of maximum, normalized oral airway resistance (expressedas percent baseline) fell to 5 ± 3%, whereas normalized nasalresistance was 80 ± 9% (both P < 0.03). Thus control of oronasal airflow partitioning in dogsappears mediated more by alterations in oral route geometry than byclosure of the nasopharyngeal airway.

     

Saltwater intrusion affects nitrogen,phosphorus and iron transformations under oxic and anoxic conditions: an incubation experiment

Biogeochemistry - Low-lying coastal ecosystems are rapidly salinizing due to sea level rise and associated saltwater intrusion (SWI). In agricultural soils, SWI can alter biogeochemical cycling of...     

Heritability of cortisol response to confinement stress in European sea bass dicentrarchus labrax

Background

In fish, the most studied production traits in terms of heritability are body weight or growth, stress or disease resistance, while heritability of cortisol levels, widely used as a measure of response to stress, is less studied. In this study, we have estimated heritabilities of two growth traits (body weight and length) and of cortisol response to confinement stress in the European sea bass.

Findings

The F1 progeny analysed (n = 922) belonged to a small effective breeding population with contributions from an unbalanced family structure of just 10 males and 2 females. Heritability values ranged from 0.54 (±0.21) for body weight to 0.65 (±0.22) for standard body length and were low for cortisol response i.e. 0.08 (±0.06). Genetic correlations were positive (0.94) between standard body length and body weight and negative between cortisol and body weight and between cortisol and standard body length (−0.60 and −0.55, respectively).

Conclusion

This study confirms that in European sea bass, heritability of growth-related traits is high and that selection on such traits has potential. However, heritability of cortisol response to stress is low in European sea bass and since it is known to vary greatly among species, further studies are necessary to understand the reasons for these differences.     

The retinoic acid binding protein CRABP2 is increased in murine models of degenerative joint disease

Introduction  

Osteoarthritis (OA) is a debilitating disease with poorly defined aetiology. Multiple signals are involved in directing the formation of cartilage during development and the vitamin A derivatives, the retinoids, figure prominently in embryonic cartilage formation. In the present study, we examined the expression of a retinoid-regulated gene in murine models of OA.