A highly conserved nuclear gene for low-level phylogenetics: elongation factor-1 alpha recovers morphology-based tree for heliothine moths

Molecular systematists need increased access to nuclear genes. Highly conserved, low copy number protein-encoding nuclear genes have attractive features for phylogenetic inference but have heretofore been applied mostly to very ancient divergences. By virtue of their synonymous substitutions, such genes should contain a wealth of information about lower-level taxonomic relationships as well, with the advantage that amino acid conservatism makes both alignment and primer definition straightforward. We tested this postulate for the elongation factor-1 alpha (EF-1 alpha) gene in the noctuid moth subfamily Heliothinae, which has probably diversified since the middle Tertiary. We sequenced 1,240 bp in 18 taxa representing heliothine groupings strongly supported by previous morphological and allozyme studies. The single most parsimonious gene tree and the neighbor-joining tree for all nucleotides show almost complete concordance with the morphological tree. Homoplasy and pairwise divergence levels are low, transition/transversion ratios are high, and phylogenetic information is spread evenly across gene regions. The EF-1 alpha gene and presumably other highly conserved genes hold much promise for phylogenetics of Tertiary age eukaryote groups.      

Plant virus infection development as affected by heavy metal stress

The work was focused on the investigation of possible dependencies between the development of viral infection in plants and the presence of high heavy metal concentrations in soil. Field experiments have been conducted in order to study the development of systemic tobacco mosaic virus (TMV) infection in Lycopersicon esculentum L. cv. Miliana plants under effect of separate salts of heavy metals Cu, Zn and Pb deposited in soil. As it is shown, simultaneous effect of viral infection and heavy metals in tenfold maximum permissible concentration leads to decrease of total chlorophyll content in experiment plants mainly due to the degradation of chlorophyll a. The reduction of chlorophyll concentration under the combined influence of both stress factors was more serious comparing to the separate effect of every single factor. Plants' treatment with toxic concentrations of lead and zinc leaded to slight delay in the development of systemic TMV infection together with more than twofold increase of virus content in plants that may be an evidence of synergism between these heavy metal's and virus' effects. Contrary, copper although decreased total chlorophyll content but showed protective properties and significantly reduced amount of virus in plants.     

The effects of amino acid supplementation on muscular performance during resistance training overreaching

The purpose of this study was to examine the effects of amino acid supplementation on muscular strength, power, and high-intensity endurance during short-term resistance training overreaching. Seventeen resistance-trained men were randomly assigned to either an amino acid (AA) or placebo (P) group and underwent 4 weeks of total-body resistance training consisting of two 2-week phases of overreaching (phase 1: 3 x 8-12 repetitions maximum [RM], 8 exercises; phase 2: 5 x 3-5 RM, 5 exercises). Muscle strength, power, and high-intensity endurance were determined before (T1) and at the end of each training week (T2-T5). One repetition maximum squat and bench press decreased at T2 in P (5.2 and 3.4 kg, respectively) but not in AA, and significant increases in 1 RM squat and bench press were observed at T3-T5 in both groups. A decrease in the ballistic bench press peak power was observed at T3 in P but not AA. The fatigue index during the 20-repetition jump squat assessment did not change in the P group at T3 and T5 (fatigue index = 18.6 and 18.3%, respectively) whereas a trend for reduction was observed in the AA group (p = 0.06) at T3 (12.8%) but not T5 (15.2%; p = 0.12). These results indicate that the initial impact of high-volume resistance training overreaching reduces muscle strength and power, and it appears that these reductions are attenuated with amino acid supplementation. In addition, an initial high-volume, moderate-intensity phase of overreaching followed by a higher intensity, moderate-volume phase appears to be very effective for enhancing muscle strength in resistance-trained men.     

Running economy and maximal oxygen consumption after 4 weeks of oral Echinacea supplementation

The purpose of this investigation was to determine the effects of 4 weeks of oral Echinacea (ECH) supplementation on erythropoietin (EPO), red blood cell (RBC) count, running economy (RE), and VO2max. Twenty-four men aged 24.9 ± 4.2 years, height 178.9 ± 7.9 cm, weight 87.9 ± 14.6 kg, body fat 19.3 ± 6.5% were grouped using a double-blind design and self-administered an 8,000-mg·d(-1) dosage of either ECH or placebo (PLA) in 5 × 400 mg × 4 times per day for 28 days. Blood samples were collected and analyzed for RBCs and EPO using automated flow cytometery and enzyme-linked immunosorbent assay. Maximal graded exercise tests (GXTs) were administered to measure VO2max, RE, and heart-rate responses. Analysis of variance was used to determine statistically significant differences (P ≤ 0.05). The EPO increased significantly in ECH at 7 days (ECH: 15.75 ± 0.64, PLA: 10.01 ± 0.73 mU·ml(-1)), 14 days (ECH: 18.88 ± 0.71, PLA: 11.02 ± 0.69 mU·ml(-1)), and 21 days (ECH: 16.06 ± 0.55, PLA: 9.20 ± 0.55 mU·ml(-1)). VO2max increased significantly in ECH (ECH: 1.47 ± 1.28, PLA: -0.13 ± 0.52%). Running economy improved significantly in ECH as indicated by a decrease in submaximal VO2max during the first 2 stages of the GXT (stage 1: ECH -1.50 ± 1.21, PLA 0.60 ± 1.95%; stage 2: ECH -1.67 ± 1.43, PLA 0.01 ± 1.03%). These data suggest that ECH supplementation results in significant increases in EPO, VO2max, and running economy.     

Rehydration with glycerol: endocrine, cardiovascular, and thermoregulatory responses during exercise in the heat.

The impact of rehydration with glycerol on cardiovascular and thermoregulatory responses during exercise in the heat was studied in eight highly trained male cyclists. Each subject completed three dehydration-rehydration experimental trials that differed only in the rehydration treatment, each separated by 7 days. Before each experimental day, subjects dehydrated to -4% of their body weight by exercise and water restriction. The experimental treatments were as follows: no fluid (NF), glycerol bolus (1 g/kg body wt) followed by water (G), and water alone (W). Rehydration (3% body weight) was given over an 80-min period. After rehydration, subjects cycled (74% peak O2 uptake) to exhaustion in a hot and wet (37 degrees C and 48% relative humidity) environment. For G, plasma volume was expanded (P < 0.05) during rehydration and remained higher than W (P < 0.05) during exercise. Exercise time to exhaustion during G (33 +/- 4 min) was longer (P < 0.05) compared with both W (27 +/- 3 min) and NF (19 +/- 3 min). Cutaneous vascular conductance was significantly elevated (P < 0.05) during G, but G provided no other thermoregulatory or cardiovascular benefits compared with W and NF. Fluid-regulating hormones (vasopressin, aldosterone, atriopeptin, and plasma renin activity) decreased during rehydration and increased during exercise (except atriopeptin), but there were no differences between G and W. These data indicated that glycerol had little or no major effect on fluid-regulating factors during rehydration or exercise, and the improved exercise capacity in G was likely due to a greater plasma volume during exercise.     

Evidence for contractile protein translocation in macrophage spreading, phagocytosis, and phagolysosome formation

Macrophage pseudopodia that surround objects during phagocytosis contain a meshwork of actin filaments and exclude organelles. Between these pseudopodia at the base of developing phagosomes, the organelle exclusion ceases, and lysosomes enter the cell periphery to fuse with the phagosomes. Macrophages also extend hyaline pseudopodia on the surface of nylon wool fibers and secrete lysosomal enzymes into the extracellular medium instead of into phagosomes. To analyze biochemically these concurrent alterations in cytoplasmic architecture, we allowed rabbit lung macrophages to spread on nylon wool fibers and then subjected the adherent cells to shear. This procedure caused the selective release of β-glucoronidase into the extracellular medium and yielded two fractions, cell bodies and isolated pseudopod blebs resembling podosomes, which are plasma-lemma-bounded sacs of cortical cytoplasm. Cytoplasmic extracts of the cell bodies eluted from nylon fibers contained two-thirds less actin-binding protein and myosin, and approximately 20 percent less actin and two-thirds of the other two proteins were accounted for in podosomes. The alterations in protein composition correlated with assays of myosin-associated EDTA-activated adenosine triphosphatase activity, and with a diminution in the capacity of extracts of nylon wool fiber-treated cell bodies to gel, a property dependent on the interaction between actin-binding protein and F-actin. However, the capacity of the remaining actin in cell bodies to polymerize did not change. We propose that actin-binding protein and myosin are concentrated in the cell cortex and particularly in pseudopodia where prominent gelation and syneresis of actin occur. Actin in the regions from which actin-binding protein and myosin are displaced disaggregates without depolymerizing, permitting lysosomes to gain access to the plasmalemma. Translocation of contractile proteins could therefore account for the concomitant differences in organelle exclusion that characterize phagocytosis.     

The eukaryotic replisome tolerates leading‐strand base damage by replicase switching

The high‐fidelity replicative DNA polymerases, Pol ε and Pol δ, are generally thought to be poorly equipped to replicate damaged DNA. Direct and complete replication of a damaged template therefore typically requires the activity of low‐fidelity translesion synthesis (TLS) polymerases. Here we show that a yeast replisome, reconstituted with purified proteins, is inherently tolerant of the common oxidative lesion thymine glycol (Tg). Surprisingly, leading‐strand Tg was bypassed efficiently in the presence and absence of the TLS machinery. Our data reveal that following helicase–polymerase uncoupling a switch from Pol ε, the canonical leading‐strand replicase, to the lagging‐strand replicase Pol δ, facilitates rapid, efficient and error‐free lesion bypass at physiological nucleotide levels. This replicase switch mechanism also promotes bypass of the unrelated oxidative lesion, 8‐oxoguanine. We propose that replicase switching may promote continued leading‐strand synthesis whenever the replisome encounters leading‐strand damage that is bypassed more efficiently by Pol δ than by Pol ε.     

Novel and functional regulatory SNPs in the promoter region of <Emphasis Type="Italic">FOXP3</Emphasis> gene in a Gabonese population

Parasites exert a selection pressure on their hosts and are accountable for driving diversity within gene families and immune gene polymorphisms in a host population. The overwhelming response of regulatory T cells during infectious challenges directs the host immune system to lose the ability to mount parasite specific T cell responses. The underlying idea of this study is that regulatory single nucleotide polymorphism (SNPs) can cause significant changes in gene expression in functional immune genes. We identified and investigated regulatory SNPs in the promoter region of the FOXP3 gene in a group of Gabonese individuals exposed to a variety of parasitic infections. We identified two novel and one promoter variants in 40 individual subjects. We further validated these promoter variants for their allelic gene expression using transient transfection assays. Two promoter variants, −794 (C/G) and the other variant −734/−540 (C/T) revealed a significant higher expression of the reporter gene at basal level in comparison to the major allele. The identification of SNPs that modify function in the promoter regions could provide a basis for studying parasite susceptibility in association studies.     

Association of CISH -292A/T genetic variant with hepatitis B virus infection

Cytokine-inducible SRC homology 2 domain protein (CISH) is a suppressor of cytokine signaling that controls interleukin-2 signaling pathway. We investigated the single nucleotide polymorphism (SNP) -292A>T in 473 Vietnamese hepatitis B virus (HBV) carriers and 416 healthy controls. CISH variants at -292A>T were associated to HBV infection (Allelic: OR, 1.22 95% CI, 1–1.49; P = 0.04; Recessive: OR, 1.69 95% CI 1.23–2.54; P = 0.007). A gene dose effect for the risk allele -292T was observed (P = 0.04). The level of interleukin 2 and liver enzymes such as alanine transaminase, aspartate transaminase, total bilirubin, and direct bilirubin were not associated to CISH polymorphism at position -292A>T This study associated the vital role of CISH SNP -292A>T variant to hepatitis B virus infection in a Vietnamese population.     

Zea ribosomal repeat evolution and substitution patterns

Zea and Tripsacum nuclear ribosomal internal transcribed spacer (ITS) sequences were used to evaluate patterns of concerted evolution, rates of substitutions, patterns of methylation-induced deamination, and structural constraints of the ITS. ITS pseudogenes were identified by their phylogenetic position, differences in nucleotide composition, extensive deamination at ancestral methylation sites, and substitutions resulting in low-stability secondary RNA structures. Selection was important in shaping the kinds of polymorphisms and substitutions observed in the ITS. ITS substitution rates were significantly different among the Zea taxa. Deamination of cytosines at methylation sites was a potent mutation source, but selection appeared to maintain high methylation site density throughout the ribosomal repeat except for the gene promoter. Nucleotide divergence statistics identified selectively constrained regions at the 5' ends of the ITS1 and ITS2.