Transposition of IS10R in Lactococcus lactis

Aims:  To characterize the transposition mechanism of the IS-element IS 10 R and study how this element is involved in gene disruption in Lactococcus lactis .
Methods and Results:  The gene flciA confers immunity against lactococcin A in lactococci. However, the immunity function was lost when flciA was co-expressed with the regulator gene nisR on a plasmid in L. lactis NZ9000. By PCR and DNA sequencing, it was revealed that flciA in immune-negative transformants was disrupted by the IS-element IS 10 R. Such gene disruption did not occur when flciA was expressed alone nor when the plasmid-located nisR was mutated, suggesting that nisR is directly involved in the transposition. The sequence 5'-CACTTAACC-3', which was found in flciA and at both ends of the inserted IS 10 R, was identified as target site by site-directed mutagenesis.
Conclusions:  IS 10 R transposes in L. lactis NZ9000 in a nisR -dependent fashion and employs the sequence 5'-CACTTAACC-3' as integration site.
Significance and Impact of the Study:  To our knowledge, this is the first time IS 10 R and aspects of its transposition are described in the industrial important bacterium L. lactis . The highly controllable insertion of IS 10 R into a target site might present a great potential as a gene disruption system.     

Hydra: software for tailored processing of H/D exchange data from MS or tandem MS analyses

Background  

Hydrogen/deuterium exchange mass spectrometry (H/DX-MS) experiments implemented to characterize protein interaction and protein folding generate large quantities of data. Organizing, processing and visualizing data requires an automated solution, particularly when accommodating new tandem mass spectrometry modes for H/DX measurement. We sought to develop software that offers flexibility in defining workflows so as to support exploratory treatments of H/DX-MS data, with a particular focus on the analysis of very large protein systems and the mining of tandem mass spectrometry data.     

Attentional Bias in Snus Users: An Experimental Study

The use of nicotine in the form of “snus” is substantial and increasing in some geographic areas, in particular among young people. It has previously been suggested that addictions may operate through a mechanism of attentional bias, in which stimuli representative of the dependent substance increase in salience, thus increasing the addictive behavior. However, this hypothesis has not been tested for the case of snus. The current experiment used a modified Stroop task and a dot-probe task to investigate whether 40 snus users show an attentional bias towards snus-relevant stimuli, compared to 40 non-snus users. There were no significant differences between the two groups on reaction times or accuracy on either Stroop or dot-probe task, thus failing to show an attentional bias towards snus-relevant stimuli for snus users. This could imply that other mechanisms may contribute to maintenance of snus use than for other addictions. However, this is the first experimental study investigating attentional bias in snus users, and more research is warranted.     

血红细胞超氧化物歧化酶(SOD)制备工艺的研究初报

本文报告了本实验室设计的由血红细胞自溶液60℃热变性, 乙醇——氯仿法除血红蛋白,旋转蒸发法减压浓缩抽去氯仿、乙醇,硫酸铵分级盐析法沉降SOD,Sepbadex G-75层析提纯SOD等步骤构成的一条成本低、设计合理、简便实用的分离纯化SOD的工艺路线。     

Is Helicobacter pylori the cause of dyspepsia?

OBJECTIVE--To determine the association between infection with Helicobacter pylori and dyspepsia. DESIGN--Cross sectional study of dyspeptic subjects and age and sex matched controls identified by a questionnaire survey of all inhabitants aged 20-69. (Endoscopy, histological examination, and microbiological examinations of biopsies from the gastric mucosa were performed blind.) SETTING--Population based survey in Sørreisa, Norway. SUBJECTS--All 782 dyspeptic subjects (excluding those with a previous history of peptic ulcer, gall stones or kidney stones, and coronary heart disease) and controls were offered an endoscopy, of whom 309 dyspeptic subjects and 310 controls attended. MAIN OUTCOME MEASURES--Prevalences of endoscopic and histological diagnoses and of cultures positive for H pylori. RESULTS--A high prevalence of positive cultures, increasing with age, was found in both dyspeptic subjects (48%) and non-dyspeptic controls (36%) (p = 0.004). Positive cultures in both dyspeptic subjects and controls were strongly associated with histological gastritis (70%, 95% confidence interval 65.5 to 85.3; 60%, 52.7 to 67.7, respectively) and peptic ulcer (92%, 61.5 to 99.8; 64.1, 9.4 to 99.2, respectively). Only 3% of subjects with a histologically non-inflamed gastric mucosa had this infection (dyspeptic subjects 2%, 0.2 to 7.0; controls 4%; 1.2 to 8.8). CONCLUSIONS--The relation between dyspeptic symptoms and H pylori is dubious; H pylori seems to have a pathogenetic role in gastritis and may be a contributing factor but not a cause of peptic ulcer.     

Histological and immunohistochemical effects of Curcuma longa on activation of rat hepatic stellate cells after cadmium induced hepatotoxicity

The liver is a target for toxic chemicals such as cadmium (Cd). When the liver is damaged, hepatic stellate cells (HSC) are activated and transformed into myofibroblast-like cells, which are responsible for liver fibrosis. Curcuma longa has been reported to exert a hepato-protective effect under various pathological conditions. We investigated the effects of C. longa administration on HSC activation in response to Cd induced hepatotoxicity. Forty adult male albino rats were divided into: group 1 (control), group 2 (Cd treated), group 3 (C. longa treated) and group 4 (Cd and C. longa treated). After 6 weeks, liver specimens were prepared for light and electron microscopy examination of histological changes and immunohistochemical localization of alpha smooth muscle actin (αSMA) as a specific marker for activated HSC. Activated HSC with a positive αSMA immune reaction were not detected in groups 1 and 3. Large numbers of activated HSC with αSMA immune reactions were observed in group 2 in addition to Cd induced hepatotoxic changes including excess collagen deposition in thickened portal triads, interlobular septa with hepatic lobulation, inflammatory cell infiltration, a significant increase in Kupffer cells and degenerated hepatocytes. In group 4, we observed a significant decrease in HSC that expressed αSMA with amelioration of the hepatotoxic changes. C. longa administration decreased HSC activation and ameliorated hepatotoxic changes caused by Cd in adult rats.     

Equilibrium and dynamic structure of large, unilamellar, unsaturated acyl chain phosphatidylcholine vesicles. Higher order analysis of 1,6-diphenyl-1,3,5-hexatriene and 1-[4-(trimethylammonio)phenyl]- 6-phenyl-1,3,5-hexatriene anisotropy decay

Equilibrium and dynamic structural properties of minimally to highly unsaturated acyl chain, large, unilamellar phosphatidylcholine (PC) vesicles have been characterized by the dynamic fluorescence properties of 1,6-diphenyl-1,3,5-hexatriene (DPH) and 1-[4-(trimethylammonio)phenyl]-6-phenyl-1,3,5-hexatriene (TMA-DPH). Fluorescence lifetimes and equilibrium and dynamic rotational properties of these probes were analyzed by limited-frequency phase-modulation fluorometry in egg PC, palmitoyloleoyl-PC (POPC), dioleoyl-PC (DOPC), palmitoylarachidonoyl-PC (PAPC), and palmitoyldocosahexaenoyl-PC (P-22:6-PC) vesicles over a temperature range from 5 to 37 degrees C. DPH equilibrium orientational distributions were derived according to a model permitting bimodal orientational distributions in which the parallel probability maximum was aligned parallel to the bilayer normal and the orthogonal probability maximum was oriented parallel to the plane of the bilayer. TMA-DPH orientational distributions were derived according to the same model except that all probability was constrained to the parallel orientation. TMA-DPH fluorescence lifetimes were much more sensitive than those of DPH to variations in acyl chain composition and temperature although the same qualitative behavior was generally observed with both probes. Greater acyl chain unsaturation and higher sample temperatures each gave rise to shorter lifetimes consistent with increased water penetrability into the bilayers. Equilibrium order of the hydrocarbon core (as probed by DPH) and of the interfacial and head group regions of the bilayers (as probed by TMA-DPH) was reduced by increasing levels of unsaturation and by higher sample temperatures.(ABSTRACT TRUNCATED AT 250 WORDS)     

Quorum-sensing based bacteriocin production is down-regulated by N-terminally truncated species of gene activators

Down-regulation of quorum-sensing based pathways is an important but yet poorly understood process in bacterial gene regulation. In this study, we show that the gene regulator plnC not only acts as an activator gene in the quorum-sensing based bacteriocin production in Lactobacillus plantarum C11, but it also concurrently codes for truncated forms that were shown to repress bacteriocin production. By amino acid N-terminal sequencing and DNA sequence analysis, the truncated species of PlnC are believed to be translated from alternative start codons located in the so-called receiver domain of the regulator. To analyse the structure–function relationship of truncated species of PlnC, we performed a series of systematic truncation mutations: ten in the receiver domain, one in the hinge region and two in the C-terminal DNA-binding domain. It was revealed that any truncation mutation containing a disrupted receiver domain together with an intact DNA-binding domain displayed a repressive effect on bacteriocin production. Such a gene repression mechanism mediated by truncated regulators was also found in two other quorum-sensing based bacteriocin systems (spp in L. sakei LTH673 and NC8-pln in L. plantarum NC8), suggesting that this mode of repression might represent a common means applied by bacteria to down-regulate certain quorum-sensing based pathways. Electronic supplementary material The online version of this article (doi:) contains supplementary material, which is available to authorized users.     

Chronomics of pressure overload-induced cardiac hypertrophy in mice reveals altered day/night gene expression and biomarkers of heart disease

There is critical demand in contemporary medicine for gene expression markers in all areas of human disease, for early detection of disease, classification, prognosis, and response to therapy. The integrity of circadian gene expression underlies cardiovascular health and disease; however time-of-day profiling in heart disease has never been examined. We hypothesized that a time-of-day chronomic approach using samples collected across 24-h cycles and analyzed by microarrays and bioinformatics advances contemporary approaches, because it includes sleep-time and/or wake-time molecular responses. As proof of concept, we demonstrate the value of this approach in cardiovascular disease using a murine Transverse Aortic Constriction (TAC) model of pressure overload-induced cardiac hypertrophy in mice. First, microarrays and a novel algorithm termed DeltaGene were used to identify time-of-day differences in gene expression in cardiac hypertrophy 8 wks post-TAC. The top 300 candidates were further analyzed using knowledge-based platforms, paring the list to 20 candidates, which were then validated by real-time polymerase chain reaction (RTPCR). Next, we tested whether the time-of-day gene expression profiles could be indicative of disease progression by comparing the 1- vs. 8-wk TAC. Lastly, since protein expression is functionally relevant, we monitored time-of-day cycling for the analogous cardiac proteins. This approach is generally applicable and can lead to new understanding of disease.