InFiRe -- a novel computational method for the identification of insertion sites in transposon mutagenized bacterial genomes

MOTIVATION: InFiRe, Insertion Finder via Restriction digest, is a novel software tool that allows for the computational identification of transposon insertion sites in known bacterial genome sequences after transposon mutagenesis experiments. The approach is based on the fact that restriction endonuclease digestions of bacterial DNA yield a unique pattern of DNA fragments with defined sizes. Transposon insertion changes the size of the hosting DNA fragment by a known number of base pairs. The exact size of this fragment can be determined by Southern blot hybridization. Subsequently, the position of insertion can be identified with computational analysis. The outlined method provides a solid basis for the establishment of a new high-throughput technology. AVAILABILITY AND IMPLEMENTATION: The software is freely available on our web server at www.infire.tu-bs.de. The algorithm was implemented in the statistical programming language R. For the most flexible use, InFiRe is provided in two different versions. A web interface offers the convenient use in a web browser. In addition, the software and source code is freely available for download as R-packages on our website. CONTACT: m.steinert@tu-bs.de SUPPLEMENTARY INFORMATION: Supplementary data are available at Bioinformatics online.     

Peculiarities of C<Subscript>2</Subscript> metabolism and intensification of the synthesis of surface-active substances in <Emphasis Type="Italic">Rhodococcus erythropolis</Emphasis> EK-1 grown in ethanol

Oxidation of ethanol, acetaldehyde, and acetate in Rhodococcus erythropolis EK-1, producer of surface-active substances (SAS), is catalyzed by N,N-dimethyl-4-nitrosoaniline (DMNA)-dependent alcohol dehydrogenase, NAD⁺/NADP⁺-dependent dehydrogenases (optimum pH 9.5), and acetate kinase/acetyl-CoA-synthetase, respectively. The glyoxylate cycle and complete tricarboxylic acid cycle function in the cells of R. erythropolis EK-1 growing on ethanol; the synthesis of phosphoenolpyruvate (PEP) is provided by the two key enzymes of gluconeogenesis, PEP carboxykinase and PEP synthetase. Introduction of citrate (0.1%) and fumarate (0.2%) into the cultivation medium of R. erythropolis EK-1 containing 2% ethanol resulted in the 1.5-and 3.5-fold increase in the activities of isocitrate lyase and PEP synthetase (the key enzymes of the glyoxylate cycle and gluconeogenesis branch of metabolism, respectively) and of lipid synthesis, as evidenced by the 1.5-fold decrease of isocitrate dehydrogenase activity. In the presence of fumarate and citrate, the indices of SAS synthesis by strain R. erythropolis EK-1 grown on ethanol increased by 40–100%.     

Role of endothelin-1 in the central neural cardiovascular control

In acute experiments on cats, endothelin-1 (ET-1) was injected into the sympathoexcitatory neuronal structures of the ventrolateral medulla (VLM) and the effects of injections on the peripheral mechanisms of circulation control were studied. Effects of ET-1 appeared to be dose-dependent. Injections of 200 pmol ET-1 into the structures of the tested brain zone resulted in considerable regular hypertensive responses due to an increase in the common peripheral vascular resistance (PVR), which corresponds to vasospasm. ET-1 in a smaller dose (100 pmol) could cause either hypertensive or, in some cases, hypotensive responses. Hemodynamic responses induced by injections of 100 pmol ET-1 into the sites of localization of vasomotor neurons in the VLM were based on less expressed shifts in the peripheral vascular resistance, as compared with the effects of ET-1 in the dose of 200 pmol. The effects were related to either facilitation or suppression (in the cases of hypertensive or hypotensive responses, respectively) of descending sympathoexcitatory influences from the VLM on different vascular pools. The threshold for excitation of vasomotor neurons in the tested brain area is likely to be considerably lower as compared with that for medullary “cardiac” neurons, because injections of 200 pmol ET-1 in the site of localization of cardiac neurons were followed by either enhancement, or suppression of the contractile myocardiac activity, while the heart rate usually decreased. The effects of ET-1 were facilitated after blocking of GABAa receptors with bicuculline and attenuated after preliminary application of GABA to the ventral surface of the medulla. The results indicate the possibility of interaction ET-1 and GABA on the neurons in the VLM. Such interaction is likely to be directed toward providing an optimum mode of regulation of the blood circulation.     

Effects of serotonin and stress on the state of the gastric mucosa in rats with the intact and transsected vagus nerve

In experiments on vagotomized and intact rats with the use of two models of experimental gastric ulceration (injection of serotonin and stress) it was demonstrated that the inhibitory action of vagotomy on haemorrhagic gastric effectiveness was more pronounced in stress than after serotonin application. Vagotomy decreased stress-induced erosive lesions but increased serotonin-induced erosions that may be a result of the increase of gastric tissue sensitivity to this amine which developed simultaneously with significant decrease of its level in gastric wall after vagotomy. Serotonin-antagonist peritol decreased stress-induced gastric disturbances in vagotomized rats more significantly than in intact rats; this suggested the great role of serotonin in anti-ulcerogenic effect of vagotomy.     

Long non‐coding RNAs in melanoma

Melanoma is the most lethal cutaneous cancer with a highly aggressive and metastatic phenotype. While recent genetic and epigenetic studies have shed new insights into the mechanism of melanoma development, the involvement of regulatory non‐coding RNAs remain unclear. Long non‐coding RNAs (lncRNAs) are a group of endogenous non‐protein‐coding RNAs with the capacity to regulate gene expression at multiple levels. Recent evidences have shown that lncRNAs can regulate many cellular processes, such as cell proliferation, differentiation, migration and invasion. In the melanoma, deregulation of a number of lncRNAs, such as HOTAIR, MALAT1, BANCR, ANRIL, SPRY‐IT1 and SAMMSON, have been reported. Our review summarizes the functional role of lncRNAs in melanoma and their potential clinical application for diagnosis, prognostication and treatment.     

The effect of cultivation conditions on the physicochemical properties of the exopolysaccharide ethapolan

The physicochemical properties of the complex exopolysaccharide ethapolan (EPS) produced by Acinetobacter sp. 12S during growth on media with various C/N ratios and different concentrations of mineral components and phosphate buffer were studied. Irrespective of the cultivation conditions, the concentrations of carbohydrates (38–44%) and pyruvic acid (3.2–3.7%) in the total EPS, as well as in the acylated (AP) and nonacylated (NAP) polysaccharides obtained from them, were practically the same. The EPS, AP, and NAP were also identical in their monosaccharide composition: the molar ratio of glucose, mannose, galactose, and rhamnose was 3: 2: 1: 1. The polysaccharides contained different concentrations of mineral salts (6–28%), uronic acid (3.7–22.0%), and fatty acids (5.8–15.4%); they also differed in the ratio of acetylated and nonacetylated polysaccharides. Due to the differences in the chemical composition and molecular mass (500 kDa–1.5 MDa), the viscosities of the EPS solutions (in the presence of 0.1 M KCl, in the H⁺-form, and in Cu²⁺-glycine system) were different as well. The mechanisms responsible for changes in the physicochemical properties of the total EPS, AP, and NAP synthesized on various media are discussed.     

Molecular genetic analysis of soriz genome (Sorghum oryzoidum)

Molecular-genetic analysis of soriz genotypes (Sorghum oryzoidum), its paternal form Sorghum bicolor (L.) Moench (grain sorghum), possible parents (Sorghum sudanense (Piper.) Stapf. (Sudan grass) and Oryza sativa L. (rice planting)) and the nearest relatives has been carried out using microsatellite (MS) loci of sorghum and rice. Based on these data genetic distances have been calculated. It was shown that soriz do not bear DNA fragments of rice, but contains in its genome DNA fragments belonging to the Sudanese grass indicating that the origin of soriz is associated with Sorghum sudanense.     

Protective effect of bixin on carbon tetrachloride-induced hepatotoxicity in rats

Background

The liver is an important organ for its ability to transform xenobiotics, making the liver tissue a prime target for toxic substances. The carotenoid bixin present in annatto is an antioxidant that can protect cells and tissues against the deleterious effects of free radicals. In this study, we evaluated the protective effect of bixin on liver damage induced by carbon tetrachloride (CCl₄) in rats.

Results

The animals were divided into four groups with six rats in each group. CCl₄ (0.125 mL kg^-1 body wt.) was injected intraperitoneally, and bixin (5.0 mg kg^-1 body wt.) was given by gavage 7 days before the CCl₄ injection. Bixin prevented the liver damage caused by CCl₄, as noted by the significant decrease in serum aminotransferases release. Bixin protected the liver against the oxidizing effects of CCl₄ by preventing a decrease in glutathione reductase activity and the levels of reduced glutathione and NADPH. The peroxidation of membrane lipids and histopathological damage of the liver was significantly prevented by bixin treatment.

Conclusion

Therefore, we can conclude that the protective effect of bixin against hepatotoxicity induced by CCl₄ is related to the antioxidant activity of the compound.     

Simple oxygraphic analysis for the presence of adenylate kinase 1 and 2 in normal and tumor cells

The adenylate kinase (AK) isoforms network plays an important role in the intracellular energy transfer processes, the maintenance of energy homeostasis, and it is a major player in AMP metabolic signaling circuits in some highly-differentiated cells. For this purpose, a rapid and sensitive method was developed that enables to estimate directly and semi-quantitatively the distribution between cytosolic AK1 and mitochondrial AK2 localized in the intermembrane space, both in isolated cells and tissue samples (biopsy material). Experiments were performed on isolated rat mitochondria or permeabilized material, including undifferentiated and differentiated neuroblastoma Neuro-2a cells, HL-1 cells, isolated rat heart cardiomyocytes as well as on human breast cancer postoperative samples. In these samples, the presence of AK1 and AK2 could be detected by high-resolution respirometry due to the functional coupling of these enzymes with ATP synthesis. By eliminating extra-mitochondrial ADP with an excess of pyruvate kinase and its substrate phosphoenolpyruvate, the coupling of the AK reaction with mitochondrial ATP synthesis could be quantified for total AK and mitochondrial AK2 as a specific AK index. In contrast to the creatine kinase pathway, the AK phosphotransfer pathway is up-regulated in murine neuroblastoma and HL-1 sarcoma cells and in these malignant cells expression of AK2 is higher than AK1. Differentiated Neuro-2a neuroblastoma cells exhibited considerably higher OXPHOS capacity than undifferentiated cells, and this was associated with a remarkable decrease in their AK activity. The respirometric method also revealed a considerable difference in mitochondrial affinity for AMP between non-transformed cells and tumor cells.