The Arabidopsis STRESS RESPONSE SUPPRESSOR DEAD‐box RNA helicases are nucleolar‐ and chromocenter‐localized proteins that undergo stress‐mediated relocalization and are involved in epigenetic gene silencing

DEAD‐box RNA helicases are involved in many aspects of RNA metabolism and in diverse biological processes in plants. Arabidopsis thaliana mutants of two DEAD‐box RNA helicases, STRESS RESPONSE SUPPRESSOR1 (STRS1) and STRS2 were previously shown to exhibit tolerance to abiotic stresses and up‐regulated stress‐responsive gene expression. Here, we show that Arabidopsis STRS‐overexpressing lines displayed a less tolerant phenotype and reduced expression of stress‐induced genes confirming the STRSs as attenuators of Arabidopsis stress responses. GFP–STRS fusion proteins exhibited localization to the nucleolus, nucleoplasm and chromocenters and exhibited relocalization in response to abscisic acid (ABA) treatment and various stresses. This relocalization was reversed when stress treatments were removed. The STRS proteins displayed mis‐localization in specific gene‐silencing mutants and exhibited RNA‐dependent ATPase and RNA‐unwinding activities. In particular, STRS2 showed mis‐localization in three out of four mutants of the RNA‐directed DNA methylation (RdDM) pathway while STRS1 was mis‐localized in the hd2c mutant that is defective in histone deacetylase activity. Furthermore, heterochromatic RdDM target loci displayed reduced DNA methylation and increased expression in the strs mutants. Taken together, our findings suggest that the STRS proteins are involved in epigenetic silencing of gene expression to bring about suppression of the Arabidopsis stress response.     

Accumulation of overoxidized Peroxiredoxin III in aged rat liver mitochondria

Overoxidation and subsequent inactivation of Peroxiredoxin III (PrxIII), a mitochondrial H₂O₂ scavenging enzyme, have been reported in oxidative stress conditions. No data are available in the literature about the presence of overoxidized forms of PrxIII in aged tissues. Liver mitochondria from 12-month-old rats and 28-month-old rats were here analyzed by two-dimensional gel electrophoresis. A spot corresponding to the native form of PrxIII was present in adult and old rats with the same volume, whereas an additional, more acidic spot, of the same molecular weight of the native form, accumulated only in old rats. The acidic spot was identified, by MALDI-MS analysis, as a form of PrxIII bearing the cysteine of the catalytic site overoxidized to sulphonic acid. This modified PrxIII form corresponds to the irreversibly inactivated enzyme, here reported, for the first time, in aging. Three groups of 28-month-old rats treated with acetyl-l-carnitine were also examined. Reduced accumulation of the overoxidized PrxIII form was found in all ALCAR-treated groups.     

High resolution preparation of monocyte-derived macrophages (MDM) protein fractions for clinical proteomics

Background  

Macrophages are involved in a number of key physiological processes and complex responses such as inflammatory, immunological, infectious diseases and iron homeostasis. These cells are specialised for iron storage and recycling from senescent erythrocytes so they play a central role in the fine tuning of iron balancing and distribution. The comprehension of the many physiological responses of macrophages implies the study of the related molecular events. To this regard, proteomic analysis, is one of the most powerful tools for the elucidation of the molecular mechanisms, in terms of changes in protein expression levels.     

Effect of moderate UV-B irradiation on Synechocystis PCC 6803 biliproteins

In the present study, we investigated the mechanism of UV-B radiation induced damage to the light harvesting apparatus of the cyanobacterium Synechocystis 6803. Liquid chromatography analysis and spectroscopy investigations performed on phycobilisomes or isolated biliproteins irradiated with moderate UV-B intensity (1.3 W/m(2)) revealed rapid destruction of beta-phycocyanin and a slower damage of the other biliproteins, alpha-phycocyanin and both alpha and beta-allophycocyanin. EPR spin trapping measurements revealed that carbon centered adducts of the spin trap DMPO were formed. This evidence indicates that free radicals produced from bilins probably attack the polypeptide chain of protein inducing its degradation. Our results show that the bilin chromophore is the main target of UV-B irradiation, causing structural changes, which in turn induce reaction of the chromophore with atmospheric oxygen and lead to production of reactive radicals. Our results also demonstrate that beta-phycocyanin is the most affected biliprotein, probably due to the presence of two bilins as chromophore.     

Novel protein phosphorylation site identification in spinach stroma membranes by titanium dioxide microcolumns and tandem mass spectrometry

In this work, spinach stroma membrane, instead of thylakoid, has been investigated for the presence of phosphorylated proteins. We identified seven previously unknown phosphorylation sites by taking advantage of TiO(2) phosphopeptides enrichment coupled to mass spectrometric analysis. Upon illumination at 100 micromol m(-2) s(-1), two novel phosphopeptides belonging to the N-terminal region of Lhcb1 light-harvesting protein were detected: NVSSGS(p)PWYGPDR and T(p)VQSSSPWYGPDR. Moreover, three new threonine residues in CP43 (Thr-6, Thr-8, and Thr-346) and, for the first time, two amino acid residues of the N-terminus of Rieske Fe-S protein of the cytochrome b(6)f complex (Thr-2 and Ser-3) were revealed to be phosphorylated. Since Lhcb1 and CP43 have been reported as mobile proteins, it may be suggested that illumination derived phosphorylation, and consequently the addition of negatively charged groups to the protein, is a necessary condition to induce a significant protein structural change.     

Pathogen responses to host immunity: the impact of time delays and memory on the evolution of virulence

Current analytical models of the mammalian immune system typically assume a specialist predator-prey relationship between invading pathogens and the active components of the immune response. However, in reality, the specific immune system is not immediately effective following invasion by a novel pathogen. First, there may be an explicit time delay between infection and immune initiation and, second, there may be a gradual build-up in immune efficacy (for instance, during the period of B-cell affinity maturation) during which the immune response develops, before reaching maximal specificity to the pathogen. Here, we use a novel theoretical approach to show that these processes, together with the presence of long-lived immune memory, decouple the immune response from current pathogen levels, greatly changing the dynamics of the pathogen-immune system interaction and the ability of the immune response to eliminate the pathogen. Furthermore, we use this model to show how distributed primary immune responses combine with immune memory to greatly affect the optimal virulence of the pathogen, potentially resulting in the evolution of highly virulent pathogens.     

Genetic history of hepatitis C virus in Venezuela: high diversity and long time of evolution of HCV genotype 2

Background

The subtype diversity of the hepatitis C virus (HCV) genotypes is unknown in Venezuela.

Methodology/Principal Findings

Partial sequencing of the NS5B region was performed in 310 isolates circulating in patients from 1995 to 2007. In the samples collected between 2005 and 2007, HCV genotype 1 (G1) was the most common genotype (63%), composed as expected of mainly G1a and G1b. G2 was the second most common genotype (33%), being G2a almost absent and G2j the most frequent subtype. Sequence analysis of the core region confirmed the subtype assignment performed within the NS5b region in 63 isolates. The complete genome sequence of G2j was obtained. G2j has been described in France, Canada and Burkina Fasso, but it was not found in Martinique, where several subtypes of G2 circulate in the general population. Bayesian coalescence analysis indicated a most recent common ancestor (MRCA) of G2j around 1785, before the introduction of G1b (1869) and G1a (1922). While HCV G1a and G1b experienced a growth reduction since 1990, coincident with the time when blood testing was implemented in Venezuela, HCV G2j did not seem to reach growth equilibrium during this period.

Conclusions/Significance

Assuming the introduction of G2j from Africa during the slave trade, the high frequency of G2j found in Venezuela could suggest: 1- the introduction of African ethnic groups different from the ones introduced to Martinique or 2- the occurrence of a founder effect. This study represents an in-depth analysis of the subtype diversity of HCV in Venezuela, which is still unexplored in the Americas and deserves further studies.     

Testosterone replacement therapy in insulin‐sensitive hypogonadal men restores phosphatidylcholine levels by regulation of arachidonic acid metabolism

Male hypogonadism is notoriously associated with altered lipid metabolism. In this study, we performed an untargeted mass spectrometry–based profiling of plasma lipids from twenty healthy and twenty hypogonadal men before and after testosterone replacement therapy (TRT) for 60 days. Results demonstrated that hypogonadism was associated with a significant increase in sphingomyelin (SM), whereas phosphatidylcholine (PC) was mainly cleaved by activated phospholipase‐A2 into lysophosphatidylcholine (LPC). In hypogonadal patients, arachidonic acid (AA), also produced through the latter cleavage, was prevalently bio‐transformed into leukotriene B4 (LTB4) and not into endoperoxides from which prostaglandins and thromboxanes are derived. Interestingly, upon testosterone treatment SM, PC and LPC returned to levels similar to controls. Also, AA was newly converted into prostaglandin‐A2, thromboxane‐A2 and 5(S)‐hydroxyeicosatetraenoic acid (HETE), suggesting that testosterone probably plays a role in controlling hypogonadal alterations above reported.     

EDTA-enhanced lead phytoremediation in sunflower (Helianthus annuus L.) hydroponic culture

The aim of the present study was to investigate the capability of Sunflower (Helianthus annuus L.) to tolerate and accumulate high amount of lead (Pb) and propose it for soil phytoremediation. To this regard, plants were grown in hydroponics and treated with different Pb concentrations (10 to 160 ??M) and a fixed concentration (500 ??M) EDTA (ethylene diamine tetra acetic acid) for 14 and 28 days (d). Effects on total biomass production, photosynthetic pigments and protein contents as well as the quantities of non protein thiols (NP-SH), glutathione (GSH), phytochelatins (PCs) and activity of glutathione reductase (GR) were estimated. Results revealed that roots (575 ??g g^?1 DW) and shoots (135 ??g g^?1 DW) accumulated Pb after 28 d of exposure, however, addition of EDTA enhanced the Pb accumulation in roots (645 ??g g^?1 DW) and shoots (255 ??g g^?1 DW ). Exposure of Pb (28 d) registered a significant (P?<?0.05) reduction in growth parameters and induction of phytochelatins (P?<?0.05; r?=?0.26) plus some of the important antioxidants (P?<?0.05; r?=?0.42), which were positively correlated to metal accumulation. Sunflower exposed at 40 ??M of Pb for 28 d synthesized higher quantity of PC₂ (18.5 fold) and PC₃ (10.5 fold), as compared to control. However, the results showed that addition of EDTA resulted in low toxicity compared to Pb alone. These data support the capability of H. annuus L. to accumulate and tolerate significant quantity of Pb and its utility for phytoremediation. This is because of the plant has the capacity to combat metal induced oxidative stress via significant synthesis of NP-SH, GSH and high activity of GR, as it would provide sufficient GSH not only for PCs synthesis but also for antioxidant function.