The properties of the complex between ribosomal protein L2 and tRNA

Escherichia coli ribosomal protein L2 interacts with fMet-tRNA^Met and NacPhe-tRNA^Phe in solution, protecting their 3'-ends from enzymatic degradation. At the same time L2 enhances the rate of spontaneous hydrolysis of the ester bonds between terminal riboses and amino acyl moieties of these two peptidyl-tRNA analogues. L2 has, however, only a slight effect on the rate of spontaneous deacylation of aminoacyltRNAs. We suggest that the role of L2 is in the fixation of the aminoacyl stem of tRNA to the ribosome at its P-site, and speculate that this protein is directly involved in the peptidyl transferase (PT) reaction. Peptidyl transferase Protein L2 tRNA-protein complex     

Electron transport across the plasmalemma of Lemna gibba G1

Lemna gibba L., grown in the presence or absence of Fe, reduced extracellular ferricyanide with a V _max of 3.09 mol · g^-1 fresh weight · h^-1 and a K _m of 115 M. However, Fe³⁺-ethylenediaminetetraacetic acid (EDTA) was reduced only after Fe-starvation. External electron acceptors such as ferricyanide, Fe³⁺-EDTA, 2,6-dichlorophenol indophenol or methylene blue induced a membrane depolarization of up to 100 mV, but electron donors such as ferrocyanide or NADH had no effect. Light or glucose enhanced ferricyanide reduction while the concomitant membrane depolarization was much smaller. Under anaerobic conditions, ferricyanide had no effect on electrical membrane potential difference (E_m). Ferricyanide reduction induced H⁺ and K⁺ release in a ratio of 1.16 H⁺+1 K⁺/2 e^- (in +Fe plants) and 1.28 H⁺+0.8 K⁺/2 e^- (in -Fe plants). Anion uptake was inhibited by ferricyanide reduction. It is concluded that the steady-state transfer of electrons and protons proceeds by separate mechanisms, by a redox system and by a H⁺-ATPase.Abbreviations E _m electrical membrane potential difference - EDTA ethylenediaminetetraacetic acid - DCPIP dichlorophenol indophenol - +Fe control plant - -Fe iron-deficient plant - FW fresh weight - H⁺ electrochemical proton gradient     

Cholesteryl ester hydrolase activity is abolished in HSL macrophages but unchanged in macrophages lacking KIAA1363

Cholesteryl ester (CE) accumulation in macrophages represents a crucial event during foam cell formation, a hallmark of atherogenesis. Here we investigated the role of two previously described CE hydrolases, hormone-sensitive lipase (HSL) and KIAA1363, in macrophage CE hydrolysis. HSL and KIAA1363 exhibited marked differences in their abilities to hydrolyze CE, triacylglycerol (TG), diacylglycerol (DG), and 2-acetyl monoalkylglycerol ether (AcMAGE), a precursor for biosynthesis of platelet-activating factor (PAF). HSL efficiently cleaved all four substrates, whereas KIAA1363 hydrolyzed only AcMAGE. This contradicts previous studies suggesting that KIAA1363 is a neutral CE hydrolase. Macrophages of KIAA1363^−/− and wild-type mice exhibited identical neutral CE hydrolase activity, which was almost abolished in tissues and macrophages of HSL^−/− mice. Conversely, AcMAGE hydrolase activity was diminished in macrophages and some tissues of KIAA1363^−/− but unchanged in HSL^−/− mice. CE turnover was unaffected in macrophages lacking KIAA1363 and HSL, whereas cAMP-dependent cholesterol efflux was influenced by HSL but not by KIAA1363. Despite decreased CE hydrolase activities, HSL^−/− macrophages exhibited CE accumulation similar to wild-type (WT) macrophages. We conclude that additional enzymes must exist that cooperate with HSL to regulate CE levels in macrophages. KIAA1363 affects AcMAGE hydrolase activity but is of minor importance as a direct CE hydrolase in macrophages.     

Cardiac-specific overexpression of perilipin 5 provokes severe cardiac steatosis via the formation of a lipolytic barrier

Cardiac triacylglycerol (TG) catabolism critically depends on the TG hydrolytic activity of adipose triglyceride lipase (ATGL). Perilipin 5 (Plin5) is expressed in cardiac muscle (CM) and has been shown to interact with ATGL and its coactivator comparative gene identification-58 (CGI-58). Furthermore, ectopic Plin5 expression increases cellular TG content and Plin5-deficient mice exhibit reduced cardiac TG levels. In this study we show that mice with cardiac muscle-specific overexpression of perilipin 5 (CM-Plin5) massively accumulate TG in CM, which is accompanied by moderately reduced fatty acid (FA) oxidizing gene expression levels. Cardiac lipid droplet (LD) preparations from CM of CM-Plin5 mice showed reduced ATGL- and hormone-sensitive lipase-mediated TG mobilization implying that Plin5 overexpression restricts cardiac lipolysis via the formation of a lipolytic barrier. To test this hypothesis, we analyzed TG hydrolytic activities in preparations of Plin5-, ATGL-, and CGI-58-transfected cells. In vitro ATGL-mediated TG hydrolysis of an artificial micellar TG substrate was not inhibited by the presence of Plin5, whereas Plin5-coated LDs were resistant toward ATGL-mediated TG catabolism. These findings strongly suggest that Plin5 functions as a lipolytic barrier to protect the cardiac TG pool from uncontrolled TG mobilization and the excessive release of free FAs.     

Lsamp–/– mice display lower sensitivity to amphetamine and have elevated 5-HT turnover

In mice, the limbic system-associated membrane protein (Lsamp) gene has been implicated in locomotion, anxiety, fear reaction, learning, social behaviour and adaptation. Human data links the LSAMP gene to several psychiatric disorders and completed suicide. Here, we investigated changes in major monoamine systems in mice lacking the Lsamp gene. First, the locomotor and rewarding effects of amphetamine were studied in Lsamp^–/– mice and Lsamp^+/+ mice. Second, monoamine levels in major brain regions in response to saline and amphetamine injections were measured and, third, the expression levels of dopamine system-related genes in the brain were studied in these mice. Lsamp^–/– mice displayed lower sensitivity to amphetamine in the motility box. Likewise, in the place preference test, the rewarding effect of amphetamine was absent in Lsamp^–/– mice. In all brain regions studied, Lsamp^–/– mice displayed lower serotonin (5-HT) baseline levels, but a greater 5-HT turnover rate, and amphetamine increased the level of 5-HT and lowered 5-HT turnover to a greater extent in Lsamp^–/– mice. Finally, Lsamp^–/– mice had lower level of dopamine transporter (DAT) mRNA in the mesencephalon. In conclusion, Lsamp-deficiency leads to increased endogenous 5-HT-ergic tone and enhanced 5-HT release in response to amphetamine. Elevated 5-HT function and reduced activity of DAT are the probable reasons for the blunted effects of amphetamine in these mice. Lsamp^–/– mice are a promising model to study the neurobiological mechanisms of deviant social behaviour and adaptation impairment observed in many psychiatric disorders.     

Wolf population genetics in Europe: a systematic review,meta‐analysis and suggestions for conservation and management

The grey wolf (Canis lupus) is an iconic large carnivore that has increasingly been recognized as an apex predator with intrinsic value and a keystone species. However, wolves have also long represented a primary source of human–carnivore conflict, which has led to long‐term persecution of wolves, resulting in a significant decrease in their numbers, genetic diversity and gene flow between populations. For more effective protection and management of wolf populations in Europe, robust scientific evidence is crucial. This review serves as an analytical summary of the main findings from wolf population genetic studies in Europe, covering major studies from the ‘pre‐genomic era’ and the first insights of the ‘genomics era’. We analyse, summarize and discuss findings derived from analyses of three compartments of the mammalian genome with different inheritance modes: maternal (mitochondrial DNA), paternal (Y chromosome) and biparental [autosomal microsatellites and single nucleotide polymorphisms (SNPs)]. To describe large‐scale trends and patterns of genetic variation in European wolf populations, we conducted a meta‐analysis based on the results of previous microsatellite studies and also included new data, covering all 19 European countries for which wolf genetic information is available: Norway, Sweden, Finland, Estonia, Latvia, Lithuania, Poland, Czech Republic, Slovakia, Germany, Belarus, Russia, Italy, Croatia, Bulgaria, Bosnia and Herzegovina, Greece, Spain and Portugal. We compared different indices of genetic diversity in wolf populations and found a significant spatial trend in heterozygosity across Europe from south‐west (lowest genetic diversity) to north‐east (highest). The range of spatial autocorrelation calculated on the basis of three characteristics of genetic diversity was 650?850 km, suggesting that the genetic diversity of a given wolf population can be influenced by populations up to 850 km away. As an important outcome of this synthesis, we discuss the most pressing issues threatening wolf populations in Europe, highlight important gaps in current knowledge, suggest solutions to overcome these limitations, and provide recommendations for science‐based wolf conservation and management at regional and Europe‐wide scales.     

Monoclonal antibodies modify acetylcholine-induced ionic channel properties in cultured chick myoballs

Summary Monoclonal antibodies directed against the cholinergic binding site of the acetylcholine receptor were found to alter the ion channel properties in cultured chick myoballs. Time and dose dependent reduction in acetylcholine sensitivity was observed. Noise analysis experiments indicated a decrease in the mean single channel conductance and an increase in the mean single channel open time.     

Vascular NAD(P)H oxidases: specific features,expression, and regulation

The importance of reactive oxygen species (ROS) in vascular physiology and pathology is becoming increasingly evident. All cell types in the vascular wall produce ROS derived from superoxide-generating protein complexes similar to the leukocyte NADPH oxidase. Specific features of the vascular enzymes include constitutive and inducible activities, substrate specificity, and intracellular superoxide production. Most phagocyte enzyme subunits are found in vascular cells, including the catalytic gp91phox (aka, nox2), which was the earliest member of the newly discovered nox family. However, smooth muscle frequently expresses nox1 rather than gp91phox, and nox4 is additionally present in all cell types. In cell culture, agonists increase ROS production by activating multiple signals, including protein kinase C and Rac, and by upregulating oxidase subunits. The oxidases are also upregulated in vascular disease and are involved in the development of atherosclerosis and a significant part of angiotensin II-induced hypertension, possibly via nox1 and nox4. Likewise, enhanced vascular oxidase activity is associated with diabetes. Therefore, members of this enzyme family appear to be important in vascular biology and disease and constitute promising targets for future therapeutic interventions.