Sequence analysis of mitochondrial chloramphenicol resistance mutations in Chinese hamster cells

A series of mitochondrially inherited chloramphenicol-resistant (CAP-R) mutants were isolated in Chinese hamster cells. To determine whether the Chinese hamster CAP-R mutations were homologous to those isolated in mouse and human cell culture systems, we determined the nucleotide sequence of the region of the mitochondrial 16S rRNA gene spanning the peptidyl transferase-encoding region for eight CAP-R mutant lines in addition to the parental wildtype line. Three main conclusions are drawn from these studies. (1) Although the region of the gene encoding the peptidyl transferase domain is highly conserved relative to that of mice and rats, the contiguous sequences show less conservation. This sequence divergence not only includes the accumulation of single base pair replacements, but also the presence of small insertions or deletions. (2) For six of the CAP-R mutants, heteroplasmic single base pair changes were detected. These mapped to the same sites within the peptidyl transferase domain as the mutations found previously in mouse and human CAP-R mutants. (3) Two Chinese hamster CAP-R mutants, both with an unusual drug resistance phenotype, did not carry any mutations within the CAP-R peptidyl transferase domain. However, both carried a heteroplasmic mutation at the position corresponding to nucleotide 2505 of the mouse 16S rRNA gene, a site predicted to map within a stem/loop structure attached to this key domain of the ribosome. This is the first evidence for mitochondrial CAP-R mutations that map outside the peptidyl transferase region.     

Acceptance of low-saponin lines of alfalfa with varied phenolic concentrations by pea aphid (Homoptera: Aphididae)

This research aims to examine the effect of phenolics on pea aphid (Acyrthosiphon pisum) (Homoptera: Aphididae) development and feeding behaviour, on leaves of selected low-saponin lines of Radius alfalfa (Medicago sativa). There was a slight, negative correlation (Spearman rank correlation r _s = −0.80) between concentrations of saponins and phenols. Lines with higher concentrations of saponins had less phenolics. Levels of phenolics in low-saponin lines of alfalfa cv. Radius were related to their acceptance by the pea aphid. Our data revealed an inverse relationship between level of phenolics and the aphid abundance and its biology on studied alfalfa lines. Larval development of the pea aphid was longer, reproduction period was shorter, and the fecundity was lower on low-saponin lines with higher level of phenolics. There were observed some tendencies in the pea aphid feeding behaviour on these lines: prolonging the probing of the peripheral tissues (epidermis and mesophyll) and shortening the period of phloem sap ingestion. The better hosts for the pea aphid were low-saponin lines with low levels of phenolic compounds.     

Hepatocytes express abundant surface class I MHC and efficiently use transporter associated with antigen processing, tapasin, and low molecular weight polypeptide proteasome subunit components of antigen processing and presentation pathway

Hepatic expression levels of class I MHC Ags are generally regarded as very low. Because the status of these Ags and their ability to present peptides are important for the understanding of pathogen clearance and tolerogenic properties of the liver, we set out to identify the factors contributing to the reported phenotype. Unexpectedly, we found that the surface densities of K(b) and D(b) on C57BL/6 mouse hepatocytes are nearly as high as on splenocytes, as are the lysate concentrations of mRNA encoding H chain and beta(2)-microglobulin (beta(2)m). In contrast, the components of the peptide-loading pathway are reduced in hepatocytes. Despite the difference in the stoichiometric ratios of H chain/beta(2)m/peptide-loading machineries, both cell types express predominantly thermostable class I and are critically dependent on TAP and tapasin for display of surface Ags. Minor differences in the expression patterns in tapasin(-/-) background suggest cell specificity in class I assembly. Under immunostimulatory conditions, such as exposure to IFN-gamma or Listeria monocytogenes, hepatocytes respond with a vigorous mRNA synthesis of the components of the Ag presentation pathway (up to 10-fold enhancement) but up-regulate H chain and beta(2)m to a lesser degree (<2-fold). This type of response should promote rapid influx of newly generated peptides into the endoplasmic reticulum and preferential presentation of foreign/induced Ag by hepatic class I.     

Structural and mechanistic diversity of secondary transporters

Recent reports on the three-dimensional structure of secondary transporters have dramatically increased our knowledge of the translocation mechanism of ions and solutes. The structures of five transporters at atomic resolution have yielded four different folds and as many different translocation mechanisms. The structure of the glutamate transporter homologue GltPh confirmed the role of pore-loop structures as essential parts of the translocation mechanism in one family of secondary transporters. Biochemical evidence for pore-loop structures in several other families suggest that they might be common in secondary transporters, adding to the structural and mechanistic diversity of secondary transporters.     

Sex, Lies and fMRI-Gender Differences in Neural Basis of Deception

Deception has always been a part of human communication as it helps to promote self-presentation. Although both men and women are equally prone to try to manage their appearance, their strategies, motivation and eagerness may be different. Here, we asked if lying could be influenced by gender on both the behavioral and neural levels. To test whether the hypothesized gender differences in brain activity related to deceptive responses were caused by differential socialization in men and women, we administered the Gender Identity Inventory probing the participants' subjective social sex role. In an fMRI session, participants were instructed either to lie or to tell the truth while answering a questionnaire focusing on general and personal information. Only for personal information, we found differences in neural responses during instructed deception in men and women. The women vs. men direct contrast revealed no significant differences in areas of activation, but men showed higher BOLD signal compared to women in the left middle frontal gyrus (MFG). Moreover, this effect remained unchanged when self-reported psychological gender was controlled for. Thus, our study showed that gender differences in the neural processes engaged during falsifying personal information might be independent from socialization.     

Vernalization and photoperiod-related changes in the DNA methylation state in winter and spring rapeseed

Vernalization-induced flowering is an effect of the epigenetic regulation of gene expression through DNA methylation and histone modifications. Vernalization-mediated silencing of a floral repressor through histone modifications was shown in Arabidopsis thaliana. However, for Brassica napus L., the mechanism underlying vernalization is unclear, and the roles of DNA methylation and histone modifications have not been established. This study revealed the profiles of changes in the DNA methylation state during vernalization (after 14, 35, 56 days) and the subsequent growth in long- or short-day photoperiods (after 2, 7, 14 days) in the winter and spring rapeseed using TLC and MSAP techniques. TLC analysis showed a significant decrease in the amount of 5-methylcytosine (m⁵C) in genomic DNA in both cultivars at the beginning of vernalization, but upon its termination, the winter rape showed a reduced level of m⁵C contrary to a significantly increased level in the spring rape. MSAP analysis revealed that winter and spring rapeseed differed in the MSAP loci which were demethylated/methylated in the course of the experiment and presented diverse profiles of changes in the methylation state. The winter rape showed permanent demethylations at 69.2 % of MSAP loci in the course of vernalization that were mostly preserved upon its termination. The spring rape showed similar numbers of demethylations and methylations that were mainly transient. The study provides evidence of the role of DNA methylation in vernalization for rapeseed and for the significant prevalence of demethylations at the beginning of vernalization, which is necessary for the transition to reproductive growth.     

Homocysteine Levels in Patients with Schizophrenia on Clozapine Monotherapy

We tested the hypothesis that homocysteine levels are higher in blood of schizophrenic subjects on clozapine monotherapy than in healthy controls and they correlate with anthropometric measurements, laboratory tests and results of bioimpedance analysis of body composition. Data for 24 subjects with schizophrenia treated with clozapine and 24 age- and sex-matched healthy volunteers was analyzed. Regarding the whole group, homocysteine levels were significantly higher in men (17.0 ± 3.4 vs. 12.1 ± 4.0 μmol/L, p = 0.009). Homocysteine levels correlated with waist circumference (R = 0.58, p = 0.003), waist-to-hip ratio (R = 0.57, p = 0.003), basal metabolic rate (R = 0.48, p = 0.01), lean body mass [kg] (R = 0.53, p = 0.008), body water [L] (R = 0.53, p = 0.008) and triglycerides (R = 0.57, p = 0.003). There were no significant differences of homocysteine levels for impaired fasting glucose, abdominal obesity, obesity/overweight, and dyslipidemia. Homocysteine levels did not correlate with age, treatment duration, clozapine dose, weight, body mass index, abdominal circumference, blood pressure, total body fat, cholesterol, high density lipoproteins, low density lipoproteins, uric acid, calcium, glucose, insulin, homoeostasis model assessment of insulin resistance 1, and homoeostasis model assessment of insulin resistance 2. We did not find significant differences in blood homocysteine levels between subjects with schizophrenia and controls. Association with waist circumference may support homocysteine role as an important cardiovascular risk factor. Association with lean weight may explain why men have higher levels of homocysteine than women.     

Tetracycline-resistant bacteria as indicators of antimicrobial resistance in protected waters—The example of the Drwęca River Nature Reserve (Poland)

The objective of this study was to determine the suitability of Tet^R tetracycline-resistant bacteria as potential indicators of drug resistance, a parameter of the microbiological quality of river waters in natural reserves which are threatened by man-made pollution. The microbiological assays covered a 15-km long section of the upper reach of the Drw?ca River (Poland), a part of the European Ecological “Natura 2000” Network of nature protected areas. The quality of the investigated ecosystem was affected by surface runoffs from the river's agricultural catchment as well as outflows from three fish farms. The counts of Tet^R bacteria, incubated at 14 °C and 28 °C on TSA medium with sheep blood and tetracycline, were determined in river water samples. The highest counts of both bacterial groups were determined in samples collected from sites behind fish farms. A statistical analysis of the abundance of Tet^R14 °C and Tet^R28 °C bacteria revealed significant differences in the size of Tet^R28 °C populations at the studied sampling sites (p = 0.0011), which is why hemolytic bacteria of this group (HemTet^R28 °C) were selected for further investigations. The predominant strains in the group of 86 HemTet^R28 °C isolates obtained by 16S rRNA gene sequencing were Pseudomonas fluorescens (42 isolates) and Aeromonas hydrophila (28 isolates). Analyses of the identified HemTet^R28 °C strains demonstrated MIC ≥256 μg/ml in more than 50% isolates. The MAR index of HemTet^R28 °C was in the range of 0.67 at the control site to 1 at sites behind fish farms. The results suggest that tetracycline-resistant bacteria, in particular HemTet^R28 °C, are a reliable indicator of antimicrobial resistance and the microbial quality of surface waters polluted due to human activity. The above can be attributed to several factors: (I) the highest percentage share of HemTet^R28 °C among HPC28 °C was noted at sites behind fish farms, (II) tetracycline-resistant bacteria quickly respond to environmental changes, as demonstrated by the high level of resistance to tetracycline and a very high MAR index, and (III) genera/species that are easy to culture under laboratory conditions dominate in the qualitative and quantitative composition of the studied bacteria.     

Responses of grass pea seedlings to salinity stress in in vitro culture conditions

Physiological and molecular mechanisms of adaptation to abiotic stresses of grass pea (Lathyrus sativus L.) are still poorly understood. Responses of four genotypes of grass pea to salinity stress in tissue culture conditions were investigated at early seedling growth stages. Salinity stress was induced in the agar media by adding 0, 50, 100 and 200 mM of NaCl. Germination and seedling emergence percentage was not significantly affected by 50 and 100 mM of NaCl. However, NaCl in 200 mM concentration lowered level of these parameters. Generally, exposure to NaCl stress significantly reduced length of grass pea seedling organs (root and shoot) but did not influence the content of dry weight in shoots and increased it in the roots in two cases. Increasing salt concentration decreased integrity of cellular membranes both in root and shoot tissues. Higher accumulation of phenolic compounds and significant changes in activity of antioxidant enzymes (peroxidase and catalase) were observed in the roots but not in the shoots. Similarly, the content of proline increased mostly in the roots from moderate (100 mM) salinity conditions. Adverse conditions did not resulted in alterations in photosynthetic pigments content of any tested genotypes. The better performance of shoots than roots may result from in vitro conditions in which experiments were conducted.