Climate change as a possible driver of invasion and differential in HSP70 expression in two genetically distinct populations of the invasive killer shrimp, <Emphasis Type="Italic">Dikerogammarus villosus</Emphasis>

Global climate change is known to affect physiological processes in charge of cellular stress response. That often results in forcing many organisms to shift their biogeographic distribution ranges. It also holds true for euryoecious and highly invasive species like the killer shrimp, Dikerogammarus villosus. In this study we compare the level of response to thermal stress in two genetically diversified populations of the amphipod D. villosus on the cellular level, namely HSP70 expression. The results show clear difference in HSP70 expression, that can be a direct consequence of the different climatic conditions both populations faced along their invasion routes. We conclude that the eastern population of D. villosus is more sensitive to thermal stress than the western population, hence its invasion potential may be lower than that of the latter. Considering the thermal tolerance of both populations and global warming, we can make some predictions about further spread of D. villosus, including the possibility of an emergence of the super-invader that may arise after cross-breeding of both populations, imposing even larger threat to the freshwater ecosystems.     

Dormancy removal of apple seeds by cold stratification is associated with fluctuation in H2O2, NO production and protein carbonylation level

Reactive oxygen (ROS) and nitrogen (RNS) species play a signaling role in seed dormancy alleviation and germination. Their action may be described by the oxidative/nitrosative “window/door”. ROS accumulation in embryos could lead to oxidative modification of protein through carbonylation. Mature apple (Malus domestica Borkh.) seeds are dormant and do not germinate. Their dormancy may be overcome by 70–90 days long cold stratification. The aim of this work was to analyze the relationship between germinability of embryos isolated from cold (5 °C) or warm (25 °C) stratified apple seeds and ROS or nitric oxide (NO) production and accumulation of protein carbonyl groups. A biphasic pattern of variation in H₂O₂ concentration in the embryos during cold stratification was detected. H₂O₂ content increased markedly after 7 days of seeds imbibition at 5 °C. After an additional two months of cold stratification, the H₂O₂ concentration in embryos reached the maximum. NO production by the embryos was low during entire period of stratification, but increased significantly in germination sensu stricto (i.e. phase II of the germination process). The highest content of protein carbonyl groups was detected after 6 weeks of cold stratification treatment. Fluctuation of H₂O₂ and protein carbonylation seems to play a pivotal role in seed dormancy alleviation by cold stratification, while NO appears to be necessary for seed germination.     

Single nucleotide polymorphisms of NR3C1 gene and recurrent depressive disorder in population of Poland

Depressive disorder is a disease characterized by disturbances in the hypothalamo–pituitary–adrenal axis. Abnormalities include the increased level of glucocorticoids (GC) and changes in sensitivity to these hormones. The changes are related to glucocorticoid receptors gene (NR3C1) variants. The NR3C1 gene is suggested to be a candidate gene affecting depressive disorder risk and management. The aim of this study was to investigate polymorphisms within the NR3C1 gene and their role in the susceptibility to recurrent depressive disorder (rDD). 181 depressive patients and 149 healthy ethnically matched controls were included in the study. Single nucleotide polymorphisms were assessed using polymerase chain reaction/restriction fragment length polymorphism method. Statistical significance between rDD patients and controls was observed for the allele and genotype frequencies at three loci: BclI, N363S, and ER22/23EK. The presence of C allele, CC, and GC genotype of BclI polymorphism, G allele and GA genotype for N363S and ER22/23EK variants respectively were associated with increased rDD risk. Two haplotypes indicated higher susceptibility for rDD, while haplotype GAG played a protective role with OR_dis 0.29 [95 % confidence interval (CI) = 0.13–0.64]. Data generated from this study support the earlier results that genetic variants of the NR3C1 gene are associated with rDD and suggest further consideration on the possible involvement of these variants in etiology of the disease.     

Co-translational Processing of Glycoprotein 3 from Equine Arteritis Virus: N-GLYCOSYLATION ADJACENT TO THE SIGNAL PEPTIDE PREVENTS CLEAVAGE*

Signal peptide cleavage and N-glycosylation of proteins are co-translational processes, but little is known about their interplay if they compete for adjacent sites. Here we report two unique findings for processing of glycoprotein 3 of equine arteritis virus. Glycoprotein 3 (Gp3) contains an N-terminal signal peptide, which is not removed, although bioinformatics predicts cleavage with high probability. There is an overlapping sequon, NNTT, adjacent to the signal peptide that we show to be glycosylated at both asparagines. Exchanging the overlapping sequon and blocking glycosylation allows signal peptide cleavage, indicating that carbohydrate attachment inhibits processing of a potentially cleavable signal peptide. Bioinformatics analyses suggest that a similar processing scheme may exist for some cellular proteins. Membrane fractionation and secretion experiments revealed that the signal peptide of Gp3 does not act as a membrane anchor, indicating that it is completely translocated into the lumen of the endoplasmic reticulum. Membrane attachment is caused by the hydrophobic C terminus of Gp3, which, however, does not span the membrane but rather attaches the protein peripherally to endoplasmic reticulum membranes.     

Characterization of a Small Molecule Inhibitor of Plasminogen Activator Inhibitor Type 1 That Accelerates the Transition into the Latent Conformation

A novel class of small molecule inhibitors for plasminogen activator inhibitor type 1 (PAI-1), represented by AZ3976, was identified in a high throughput screening campaign. AZ3976 displayed an IC₅₀ value of 26 μm in an enzymatic chromogenic assay. In a plasma clot lysis assay, the compound was active with an IC₅₀ of 16 μm. Surprisingly, AZ3976 did not bind to active PAI-1 but bound to latent PAI-1 with a K_D of 0.29 μm at 35 °C and a binding stoichiometry of 0.94, as measured by isothermal calorimetry. Reversible binding was confirmed by surface plasmon resonance direct binding experiments. The x-ray structure of AZ3976 in complex with latent PAI-1 was determined at 2.4 Å resolution. The inhibitor was bound in the flexible joint region with the entrance to the cavity located between α-helix D and β-strand 2A. A set of surface plasmon resonance experiments revealed that AZ3976 inhibited PAI-1 by enhancing the latency transition of active PAI-1. Because AZ3976 only had measurable affinity for latent PAI-1, we propose that its mechanism of inhibition is based on binding to a small fraction in equilibrium with active PAI-1, a latent-like prelatent form, from which latent PAI-1 is then generated more rapidly. This mode of action, with induced accelerated latency transition of active PAI-1 may, together with supporting x-ray data, provide improved opportunities for small molecule drug design in the hunt for therapeutically useful PAI-1 inhibitors.     

The Activity of Class I, II, III and IV of Alcohol Dehydrogenase (ADH) Isoenzymes and Aldehyde Dehydrogenase (ALDH) in Brain Cancer

The brain being highly sensitive to the action of alcohol is potentially susceptible to its carcinogenic effects. Alcohol dehydrogenase (ADH) and aldehyde dehydrogenase (ALDH) are the main enzymes involved in ethanol metabolism, which leads to the generation of carcinogenic acetaldehyde. Human brain tissue contains various ADH isoenzymes and possess also ALDH activity. The purpose of this study was to compare the capacity for ethanol metabolism measured by ADH isoenzymes and ALDH activity in cancer tissues and healthy brain cells. The samples were taken from 62 brain cancer patients (36 glioblastoma, 26 meningioma). For the measurement of the activity of class I and II ADH isoenzymes and ALDH activity, the fluorometric methods were used. The total ADH activity and activity of class III and IV isoenzymes were measured by the photometric method. The total activity of ADH, and activity of class I ADH were significantly higher in cancer cells than in healthy tissues. The other tested classes of ADH and ALDH did not show statistically significant differences of activity in cancer and in normal cells. Analysis of the enzymes activity did not show significant differences depending on the location of the tumor. The differences in the activity of total alcohol dehydrogenase, and class I isoenzyme between cancer tissues and healthy brain cells might be a factor for metabolic changes and disturbances in low mature cancer cells and additionally might be a reason for higher level of acetaldehyde which can intensify the carcinogenesis.     

Reputation-based partner choice is an effective alternative to indirect reciprocity in solving social dilemmas

When group interests clash with individual ones, maintaining cooperation poses a problem. However, cooperation can be facilitated by introducing reputational incentives. Through indirect reciprocity, people who cooperate in a social dilemma are more likely to receive cooperative acts from others. Another mechanism that enhances group cooperation is reputation-based partner choice, or competitive altruism. According to this framework, cooperators benefit via increased access to cooperative partners. Our study compared the effectiveness of indirect reciprocity and competitive altruism in re-establishing cooperation after the typical decline found during repeated public goods games. Twenty groups of four participants first played a series of public goods games, which confirmed the expected decline. Subsequently, public goods games were alternated with either indirect reciprocity games (in which participants had an opportunity to give to another individual from whom they would never receive a direct return) or competitive altruism games (in which they could choose partners for directly reciprocal interactions). We found that public goods game contributions increased when interspersed with competitive altruism games; they were also higher than in public goods games interspersed with indirect reciprocity games. Investing in reputation by increasing contributions to public goods was a profitable strategy in that it increased returns in subsequent competitive altruism and indirect reciprocity games. There was also some evidence that these returns were greater under competitive altruism than indirect reciprocity. Our findings indicate that strategic reputation building through competitive altruism provides an effective alternative to indirect reciprocity as a means for restoring cooperation in social dilemmas.     

Effects of the semi-dwarfing sdw1/denso gene in barley

Recent advances in cereal genomics have made it possible to analyse the architecture of cereal genomes and their expressed components, leading to an increase in our knowledge of those genes that are associated with the key agronomical traits. Presently, use of a dwarfing gene in breeding process is crucial for the development of modern cultivars. In barley, more than 30 types of dwarfs or semi-dwarfs have been hitherto described. However, only a few of them have been successfully used in barley breeding programs. Both breeding and molecular mapping experiments were undertaken to enhance and evaluate the performance of semi-dwarf barley lines. The semi-dwarfing cultivars had improved lodging resistance and a higher harvest index. There have been a lot of investigations that have contributed new information to our basic understanding of the mechanisms underlying growth regulations in barley. This paper reviews semi-dwarfing genes in barley in general and special attention is paid to mapping of the sdw1/denso locus, changes in protein abundance and associations of the semi-dwarfness with gibberellins.     

Potential tumor biomarkers identified in ovarian cyst fluid by quantitative proteomic analysis,iTRAQ

Background

Epithelial-derived ovarian adenocarcinoma (EOC) is the most deadly gynecologic tumor, and the principle cause of the poor survival rate is diagnosis at a late stage. Screening and diagnostic biomarkers with acceptable specificity and sensitivity are lacking. Ovarian cyst fluid should harbor early ovarian cancer biomarkers because of its closeness to the tumor. We investigated ovarian cyst fluid as a source for discovering biomarkers for use in the diagnosis of EOC.

Results

Using quantitative mass spectrometry, iTRAQ MS, we identified 837 proteins in cyst fluid from benign, EOC stage I, and EOC stage III. Only patients of serous histology were included in the study. Comparing the benign (n = 5) with the malignant (n = 10) group, 87 of the proteins were significantly (p < 0.05) differentially expressed. Two proteins, serum amyloid A-4 (SAA4) and astacin-like metalloendopeptidase (ASTL), were selected for verification of the iTRAQ method and external validation with immunoblot in a larger cohort with mixed histology, in plasma (n = 68), and cyst fluid (n = 68). The protein selections were based on either high significance and high fold change or abundant appearance and several peptide recognitions in the sample sets (p = 0.04, FC = 1.95) and (p < 0.001, FC = 8.48) for SAA4 and ASTL respectively. Both were found to be significantly expressed (p < 0.05), but the methods did not correlate concerning ASTL.

Conclusions

Fluid from ovarian cysts connected directly to the primary tumor harbor many possible new tumor-specific biomarkers. We have identified 87 differentially expressed proteins and validated two candidates to verify the iTRAQ method. However several of the proteins are of interest for validation in a larger setting.     

High quality methylome-wide investigations through next-generation sequencing of DNA from a single archived dry blood spot

The potential importance of DNA methylation in the etiology of complex diseases has led to interest in the development of methylome-wide association studies (MWAS) aimed at interrogating all methylation sites in the human genome. When using blood as biomaterial for a MWAS the DNA is typically extracted directly from fresh or frozen whole blood that was collected via venous puncture. However, DNA extracted from dry blood spots may also be an alternative starting material. In the present study, we apply a methyl-CpG binding domain (MBD) protein enrichment-based technique in combination with next generation sequencing (MBD-seq) to assess the methylation status of the ~27 million CpGs in the human autosomal reference genome. We investigate eight methylomes using DNA from blood spots. This data are compared with 1,500 methylomes previously assayed with the same MBD-seq approach using DNA from whole blood. When investigating the sequence quality and the enrichment profile across biological features, we find that DNA extracted from blood spots gives comparable results with DNA extracted from whole blood. Only if the amount of starting material is ≤ 0.5µg DNA we observe a slight decrease in the assay performance. In conclusion, we show that high quality methylome-wide investigations using MBD-seq can be conducted in DNA extracted from archived dry blood spots without sacrificing quality and without bias in enrichment profile as long as the amount of starting material is sufficient. In general, the amount of DNA extracted from a single blood spot is sufficient for methylome-wide investigations with the MBD-seq approach.