Comparison of Barium and Arsenic Concentrations in Well Drinking Water and in Human Body Samples and a Novel Remediation System for These Elements in Well Drinking Water

Health risk for well drinking water is a worldwide problem. Our recent studies showed increased toxicity by exposure to barium alone (≤700 µg/L) and coexposure to barium (137 µg/L) and arsenic (225 µg/L). The present edition of WHO health-based guidelines for drinking water revised in 2011 has maintained the values of arsenic (10 µg/L) and barium (700 µg/L), but not elements such as manganese, iron and zinc. Nevertheless, there have been very few studies on barium in drinking water and human samples. This study showed significant correlations between levels of arsenic and barium, but not its homologous elements (magnesium, calcium and strontium), in urine, toenail and hair samples obtained from residents of Jessore, Bangladesh. Significant correlation between levels of arsenic and barium in well drinking water and levels in human urine, toenail and hair samples were also observed. Based on these results, a high-performance and low-cost adsorbent composed of a hydrotalcite-like compound for barium and arsenic was developed. The adsorbent reduced levels of barium and arsenic from well water in Bangladesh and Vietnam to <7 µg/L within 1 min. Thus, we have showed levels of arsenic and barium in humans and propose a novel remediation system.     

Role of Musclin in the Pathogenesis of Hypertension in Rat

Musclin is a novel skeletal muscle-derived secretory factor found in the signal sequence trap of mouse skeletal muscle cDNAs. Musclin possesses a region homologous to the natriuretic peptide family. Thus, musclin is found to bind with the natriuretic peptide clearance receptors. However, the role of musclin in vascular regulation remains unclear. In this study, we aim to investigate the direct effect of musclin on vascular tone and to analyze its role in hypertension using the spontaneously hypertensive rats (SHR). In aortic strips isolated from SHR, musclin induced contractions in a dose-dependent manner. We found that the musclin-induced vasoconstriction was more marked in SHR than in normal rats (WKY). Moreover, this contraction was reduced by blockade of natriuretic peptide receptor C using the ab14355 antibody. Therefore, mediation of the natriuretic peptide receptor in musclin-induced vasoconstriction can be considered. In addition, similar to the natriuretic peptide receptor, expression of the musclin gene in blood vessels was higher in SHR than in WKY. Injection of musclin markedly increased the blood pressure in rats that can be inhibited by anti-musclin antibodies. Musclin-induced vasoconstriction was more pronounced in SHR than in WKY as in its expression. Taken together, these results suggest that musclin is involved in blood pressure regulation. The higher expression of musclin in hypertension indicates that musclin could be used as a new target for the treatment of hypertension in the future.     

Effect of morph types, body size and prior residence on food-site holding by males of the male-dimorphic stag beetle Prosopocoilus inclinatus (Coleoptera: Lucanidae)

Mate-securing tactics of small males in male-polymorphic species exhibiting male–male combat is an important issue in behavioral ecology. While most studies have focused on the outcomes of such combat encounters, the holding of a mating resource like a feeding site has a greater impact for obtaining reproductive success. We examined the effects of the prior residence at a feeding site on resource acquisition in the male-dimorphic stag beetle, Prosopocoilus inclinatus. More than 70 % of encounters did not result in combat. While larger males tended to occupy a food site after a combat, smaller males with prior residence tended to occupy food sites when no fighting occurred. Morph types or body size have no effect on the occurrence of combat, meaning that small males do not hesitate to fight with large males. These findings show that, under experimental conditions, the prior residence has a positive effect to hold food site in P. inclinatus.     

Fibronectin‐binding protein,FbpA, is the adhesin responsible for pathogenesis of Listeria monocytogenes infection

The role of fibronectin binding protein A (FbpA) in Listeria monocytogenes infection and its pathogenesis were studied in vivo and in vitro by constructing a fbpA‐deficient mutant of L. monocytogenes (ΔfbpA). In vivo, ΔfbpA was less pathogenic in mutant mice than was wild‐type L. monocytogenes. FbpA did not affect the amounts of various virulence‐determining factors, including internalin B and listeriolysin O. However, adherence to, and invasion of, mouse hepatocytes by the ΔfbpA mutant were reduced. In contrast, adherence to, but not invasion of, the ΔfbpA mutant to macrophages was attenuated. Fibronectin contributed to the efficient adherence and invasion of wild‐type L. monocytogenes, but not to those of the ΔfbpA mutant. Attenuation of adhesion and uptake of the ΔfbpA mutant were reversed by overexpression of FbpA in it. FbpA was not involved in intracellular growth, autophagy induction or actin tail formation. Thus, the present findings clearly show that FbpA acts as an important adhesion molecule of L. monocytogenes, especially regarding hepatocytes, without modulating the expression of other virulence factors that have been implicated in the pathogenesis of L. monocytogenes infection.     

Molecular dynamics simulation to investigate the impact of disulfide bond formation on conformational stability of chicken cystatin I66Q mutant

Chicken cystatin (cC) mutant I66Q is located in the hydrophobic core of the protein and increases the propensity for amyloid formation. Here, we demonstrate that under physiological conditions, the replacement of Ile with the Gln in the I66Q mutant increases the susceptibility for the disulfide bond Cys71–Cys81 to be reduced when compared to the wild type (WT) cC. Molecular dynamics (MD) simulations under conditions favoring cC amyloid fibril formation are in agreement with the experimental results. MD simulations were also performed to investigate the impact of disrupting the Cys71–Cys81 disulfide bond on the conformational stability of cC at the atomic level, and highlighted major disruption to the cC appendant structure. Domain swapping and extensive unfolding has been proposed as one of the possible mechanisms initiating amyloid fibril formation by cystatin. Our in silico studies suggest that disulfide bond formation between residues Cys95 and Cys115 is necessary to maintain conformational stability of the I66Q mutant following breakage of the Cys71–Cys81 disulfide bridge. Subsequent breakage of disulfide bond Cys95–Cys115 resulted in large structural destabilization of the I66Q mutant, which increased the α–β interface distance and expanded the hydrophobic core. These experimental and computational studies provide molecular-level insight into the relationship between disulfide bond formation and progressive unfolding of amyloidogenic cC mutant I66Q.

An animated Interactive 3D Complement (I3DC) is available in Proteopedia at http://proteopedia.org/w/Journal:JBSD:23     

Nitrogen dynamics following field application of biochar in a temperate North American maize-based production system

Background and aims

Biochar additions to tropical soils have been shown to reduce N leaching and increase N use efficiency. No studies exist verifying reduced N leaching in field experiments on temperate agricultural soils or identifying the mechanism for N retention.

Methods

Biochar derived from maize stover was applied to a maize cropping system in central New York State at rates of 0, 1, 3, 12, and 30 t?ha^-1 in 2007. Secondary N fertilizer was added at 100, 90, 70, and 50 % of the recommended rate (108 kg N ha^-1). Nitrogen fertilizer enriched with ¹⁵?N was applied in 2009 to the 0 and 12 t?ha^-1 of biochar at 100 and 50 % secondary N application.

Results

Maize yield and plant N uptake did not change with biochar additions (p?>?0.05; n?=?3). Less N (by 82 %; p?<?0.05) was lost after biochar application through leaching only at 100 %?N fertilization. The reason for an observed 140 % greater retention of applied ¹⁵?N in the topsoil may have been the incorporation of added ¹⁵?N into microbial biomass which increased approximately three-fold which warrants further research. The low leaching of applied fertilizer ¹⁵?N (0.42 % of applied N; p?<?0.05) and comparatively high recovery of applied ¹⁵?N in the soil (39 %) after biochar additions after one cropping season may also indicate greater overall N retention through lower gaseous or erosion N losses with biochar.

Conclusions

Addition of biochar to fertile soil in a temperate climate did not improve crop growth or N use efficiency, but increased retention of fertilizer N in the topsoil.     

Generation of Rhesus Macaque-Tropic HIV-1 Clones That Are Resistant to Major Anti-HIV-1 Restriction Factors

Human immunodeficiency virus type 1 (HIV-1) replication in macaque cells is restricted mainly by antiviral cellular APOBEC3, TRIM5α/TRIM5CypA, and tetherin proteins. For basic and clinical HIV-1/AIDS studies, efforts to construct macaque-tropic HIV-1 (HIV-1mt) have been made by us and others. Although rhesus macaques are commonly and successfully used as infection models, no HIV-1 derivatives suitable for in vivo rhesus research are available to date. In this study, to obtain novel HIV-1mt clones that are resistant to major restriction factors, we altered Gag and Vpu of our best HIV-1mt clone described previously. First, by sequence- and structure-guided mutagenesis, three amino acid residues in Gag-capsid (CA) (M94L/R98S/G114Q) were found to be responsible for viral growth enhancement in a macaque cell line. Results of in vitro TRIM5α susceptibility testing of HIV-1mt carrying these substitutions correlated well with the increased viral replication potential in macaque peripheral blood mononuclear cells (PBMCs) with different TRIM5 alleles, suggesting that the three amino acids in HIV-1mt CA are involved in the interaction with TRIM5α. Second, we replaced the transmembrane domain of Vpu of this clone with the corresponding region of simian immunodeficiency virus SIVgsn166 Vpu. The resultant clone, MN4/LSDQgtu, was able to antagonize macaque but not human tetherin, and its Vpu effectively functioned during viral replication in a macaque cell line. Notably, MN4/LSDQgtu grew comparably to SIVmac239 and much better than any of our other HIV-1mt clones in rhesus macaque PBMCs. In sum, MN4/LSDQgtu is the first HIV-1 derivative that exhibits resistance to the major restriction factors in rhesus macaque cells.     

Important functional role of residue x of the presenilin GxGD protease active site motif for APP substrate cleavage specificity and substrate selectivity of γ‐secretase

γ‐Secretase plays a central role in the generation of the Alzheimer disease‐causing amyloid β‐peptide (Aβ) from the β‐amyloid precursor protein (APP) and is thus a major Alzheimer′s disease drug target. As several other γ‐secretase substrates including Notch1 and CD44 have crucial signaling functions, an understanding of the mechanism of substrate recognition and cleavage is key for the development of APP selective γ‐secretase‐targeting drugs. The γ‐secretase active site domain in its catalytic subunit presenilin (PS) 1 has been implicated in substrate recognition/docking and cleavage. Highly critical in this process is its GxGD active site motif, whose invariant glycine residues cannot be replaced without causing severe functional losses in substrate selection and/or cleavage efficiency. Here, we have investigated the contribution of the less well characterized residue x of the motif (L383 in PS1) to this function. Extensive mutational analysis showed that processing of APP was overall well‐tolerated over a wide range of hydrophobic and hydrophilic mutations. Interestingly, however, most L383 mutants gave rise to reduced levels of Aβ_37–39 species, and several increased the pathogenic Aβ_42/43 species. Several of the Aβ_42/43‐increasing mutants severely impaired the cleavages of Notch1 and CD44 substrates, which were not affected by any other L383 mutation. Our data thus establish an important, but compared with the glycine residues of the motif, overall less critical functional role for L383. We suggest that L383 and the flanking glycine residues form a spatial arrangement in PS1 that is critical for docking and/or cleavage of different γ‐secretase substrates.     

Inhibition of the First Step in Synthesis of the Mycobacterial Cell Wall Core,Catalyzed by the GlcNAc-1-phosphate Transferase WecA,by the Novel Caprazamycin Derivative CPZEN-45

Because tuberculosis is one of the most prevalent and serious infections, countermeasures against it are urgently required. We isolated the antitubercular agents caprazamycins from the culture of an actinomycete strain and created CPZEN-45 as the most promising derivative of the caprazamycins. Herein, we describe the mode of action of CPZEN-45 first against Bacillus subtilis. Unlike the caprazamycins, CPZEN-45 strongly inhibited incorporation of radiolabeled glycerol into growing cultures and showed antibacterial activity against caprazamycin-resistant strains, including a strain overexpressing translocase-I (MraY, involved in the biosynthesis of peptidoglycan), the target of the caprazamycins. By contrast, CPZEN-45 was not effective against a strain overexpressing undecaprenyl-phosphate–GlcNAc-1-phosphate transferase (TagO, involved in the biosynthesis of teichoic acid), and a mutation was found in the tagO gene of the spontaneous CPZEN-45-resistant strain. This suggested that the primary target of CPZEN-45 in B. subtilis is TagO, which is a different target from that of the parent caprazamycins. This suggestion was confirmed by evaluation of the activities of these enzymes. Finally, we showed that CPZEN-45 was effective against WecA (Rv1302, also called Rfe) of Mycobacterium tuberculosis, the ortholog of TagO and involved in the biosynthesis of the mycolylarabinogalactan of the cell wall of M. tuberculosis. The outlook for WecA as a promising target for the development of antituberculous drugs as a countermeasure of drug resistant tuberculosis is discussed.