Eicosapentaenoic acid modifies lipid composition in caveolae and induces translocation of endothelial nitric oxide synthase

Endothelial nitric oxide synthase (eNOS) plays a crucial role in the regulation of a variety of cardiovascular functions. Many studies have shown that dietary n-3 polyunsaturated fatty acids (PUFAs) have beneficial effects on coronary atherosclerosis. However, the mechanisms of n-3 PUFAs regulation in eNOS activation remain unknown. In the present study we investigated the effects of eicosapentaenoic acid (EPA, 20:5 n-3) on subcellular distribution of eNOS and lipid composition of caveolae. We demonstrated for the first time that EPA treatment profoundly altered lipid composition and fatty acyl substitutions of phospholipids in caveolae. We found that caveolin-1 was solely located in caveolae fractions in control cells, and EPA treatment displaced caveolin-1 from caveolae. eNOS was detected in the caveolin-enriched fractions and noncaveolae fractions in control cells. EPA treatment induced the translocation of eNOS from caveolae fractions to soluble fractions. P-eNOS was also distributed in both fractions. After EPA treatment, the level of p-eNOS in each fraction was increased but the distribution of which was unaffected. Moreover, the results of immunofluorescence confirmed that EPA could redistribute caveolin-1 and eNOS in plasma membrane. eNOS activity in HUVEC cells was increased after EPA treatment, which was in a dose dependent manner. And incubation with 50 microM EPA had the maximum effect on eNOS activity. Our results suggested that eNOS translocation was paralleled by a stimulated capacity for NO production in the cells. We found that total Akt and p-Akt were primarily presented in heavy membranes in control cells, and the relative level of p-Akt increased but the distribution did not change after EPA treatment. The distribution of CaM was slightly changed after EPA treatment. Our results indicated that n-3 PUFAs profoundly altered caveolae microenvironment, thereby modifying location and function of proteins in caveolae. EPA-induced alterations of lipid and proteins in caveolae may be an important mechanism in the pathophysiologic process of atherosclerosis.     

Heat stress in grapevine: the pros and cons of acclimation

Heat stress is a major limiting factor of grapevine production and quality. Acclimation and recovery are essential to ensure plant survival, and the recovery mechanisms can be independent of the heat response mechanisms. An experimental set up with and without acclimation to heat followed by recovery [stepwise acclimation and recovery (SAR) and stepwise recovery (SR), respectively] was applied to two grapevine varieties, Touriga Nacional (TN), and Trincadeira (TR), with different tolerance to abiotic stress. Major differences were found between leaves of SAR and SR, especially after recovery; in SAR, almost all parameters returned to basal levels while in SR they remained altered. Acclimation led to a swifter and short‐term antioxidative response, affecting the plant to a lesser extent than SR. Significant differences were found among varieties: upon stress, TN significantly increased ascorbate and glutathione reduction levels, boosting the cell's redox‐buffering capacity, while TR needed to synthesize both metabolites, its response being insufficient to keep the redox state at working levels. TR was affected by stress for a longer period and the up‐regulation pattern of antioxidative stress genes was more obvious. In TN, heat shock proteins were significantly induced, but the canonical heat‐stress gene signature was not evident probably because no shutdown of the housekeeping metabolism was needed.     

Comparative kinetics of cofactor association and dissociation for the human and trypanosomal S-adenosylhomocysteine hydrolases. 1. Basic features of the association and dissociation processes

The S-adenosyl-l-homocysteine (AdoHcy) hydrolases catalyze the reversible conversion of AdoHcy to adenosine and homocysteine, making use of a catalytic cycle in which a tightly bound NAD+ oxidizes the 3-hydroxyl group of the substrate at the beginning of the cycle, activating the 4-CH bond for elimination of homocysteine, followed by Michael addition of water to the resulting intermediate and a final reduction by the tightly bound NADH to give adenosine. The equilibrium and kinetic properties of the association and dissociation of the cofactor NAD+ from the enzymes of Homo sapiens (Hs-SAHH) and Trypanosoma cruzi (Tc-SAHH) are qualitatively similar but quantitatively distinct. Both enzymes bind NAD+ in a complex scheme. The four active sites of the homotetrameric apoenzyme appear to divide into two numerically equal classes of active sites. One class of sites binds cofactor weakly and generates full activity very rapidly (in less than 1 min). The other class binds cofactor more strongly but generates activity only slowly (>30 min). In the case of Tc-SAHH, the final affinity for NAD+ is roughly micromolar and this affinity persists as the equilibrium affinity. In the case of Hs-SAHH, the slow-binding phase terminates in micromolar affinity also, but over a period of hours, the dissociation rate constant decreases until the final equilibrium affinity is in the nanomolar range. The slow binding of NAD+ by both enzymes exhibits saturation kinetics with respect to the cofactor concentration; however, binding to Hs-SAHH has a maximum rate constant around 0.06 s-1, while the rate constant for binding to Tc-SAHH levels out at 0.006 s-1. In contrast to the complex kinetics of association, both enzymes undergo dissociation of NAD+ from all four sites in a single first-order reaction. The equilibrium affinities of both Hs-SAHH and Tc-SAHH for NADH are in the nanomolar range. The dissociation rate constants and the slow-binding association rate constants for NAD+ show a complex temperature dependence with both enzymes; however, the cofactor always dissociates more rapidly from Tc-SAHH than from Hs-SAHH, the ratio being around 80-fold at 37 degrees C, and the cofactor binds more rapidly to Hs-SAHH than to Tc-SAHH above approximately 16 degrees C. These features present an opening for selective inhibition of Tc-SAHH over Hs-SAHH, demonstrated with the thioamide analogues of NAD+ and NADH. Both analogues bind to Hs-SAHH with approximately 40 nM affinities but much more weakly to Tc-SAHH (0.6-15 microM). Nevertheless, both analogues inactivated Tc-SAHH 60% (NAD+ analogue) or 100% (NADH analogue) within 30 min, while the degree of inhibition of Hs-SAHH approached 30% only after 12 h. The rate of loss of activity is equal to the rate of dissociation of the cofactor and thus 80-fold faster at 37 degrees C for Tc-SAHH.     

不同生境下空心莲子草响应模拟昆虫采食的生长和化学防御策略

外来植物往往可以入侵多种生境并受到多种昆虫的采食,而不同生境条件将可能会影响这些入侵植物对昆虫采食的防御策略。以入侵我国的克隆植物——空心莲子草为研究对象,分别选择生长在水生生境、水陆两栖生境和陆生生境中的无性个体(分株),通过50%去叶处理模拟昆虫采食,分析不同生境下空心莲子草对模拟昆虫采食处理的生长及化学防御响应的差异。模拟昆虫采食处理显著抑制了陆生生境、水陆两栖生境以及水生生境下空心莲子草的根、茎、叶和总生物量,但对3种生境下空心莲子草的生物量分配(根冠比、根生物量分配、茎生物量分配和叶生物量分配)均无显著影响。陆生生境下空心莲子草根、茎和总生物量显著高于水陆两栖生境和水生生境,根冠比显著低于水陆两栖生境和水生生境。模拟昆虫采食处理显著降低了空心莲子草的木质素含量,而对单宁和总酚含量影响不显著。生境对木质素含量无显著影响,但陆生生境下空心莲子草单宁含量显著高于水陆两栖生境和水生生境,且总酚含量显著高于水陆两栖生境,表明陆生生境中空心莲子草具有更强的防御能力。空心莲子草木质素含量与总生物量无显著相关性,但在模拟采食情况下,其总酚含量与总生物量呈显著负相关,而无论模拟昆虫采食处理存在与否,空心莲子草单宁含量与总生物量均呈显著正相关。因此,空心莲子草存在昆虫介导的生长和化学防御之间的权衡,在昆虫采食的情况下可通过减少生长来增加对化学防御物质的投入,但生境对空心莲子草这种生长-防御权衡的影响十分有限。     

Richness and diversity of Leguminosae in an altitudinal gradient in the tropical semi‐arid zone of Brazil

Abstract Many studies are based on the premise that, on a local scale, diversity is the result of ecological processes, whereas on a regional scale factors such as the topography, geology, hydrology, and historical and evolutionary events would influence this control. The Baturité Mountain Range (Ceará state), located in the Brazilian semi‐arid zone, is considered an area of extreme importance for conservation with its vegetation varying with the altitude and slope (windward vs. leeward). On the windward (wet) slope, rainforest dominates, whereas the leeward (dry) slope is dominated by seasonal forests and thorny woodland. The aim of this study was to contribute to the knowledge of the patterns of richness and diversity of the family Leguminosae on a local scale (Baturité Mountain Range) as well as a regional scale (northeastern Brazil). The two slopes present quite distinct floras. The dry slope presents higher richness and diversity indices for Leguminosae than the wet slope. The highest diversity of Leguminosae in the dry areas did not corroborate the ideas of other studies carried out in neotropical forests (total flora) that the higher species richness was predicted for wet areas. The present study indicates that the historical and evolutionary processes influence the diversity patterns on a local scale (Baturité Mountain Range), as well as on a regional scale (Brazilian semi‐arid). Our results reinforce the uniqueness of each portion of this area and its importance for conservation.     

Effective inhibition of C3a-mediated pro-inflammatory response by a human C3a-specific protein binder

Complement component 3a (C3a) plays a crucial role in the immune response and host defense, but it is also involved in pro-inflammatory responses, causing many inflammatory disorders. Blockade of C3a has been regarded as a potent therapeutic strategy for inflammatory diseases. Here, we present the development of a human C3a (hC3a)-specific protein binder, which effectively inhibits pro-inflammatory responses. The protein binder, which is composed of leucine-rich repeat modules, was selected against hC3a through phage display, and its binding affinity was matured up to 600 pM by further expanding the binding interface in a module-by-module manner. The developed protein binder was shown to have more than 10-fold higher specificity to hC3a compared with human C5a, exhibiting a remarkable suppression effect on pro-inflammatory response in monocyte, by blocking the interaction between hC3a and its receptor. The hC3a-specific protein binder is likely to have a therapeutic potential for C3a-mediated inflammatory diseases.     

Multiple paternity and mating group size in the European earwig,Forficula auricularia

1. The patterns of multiple paternity among the progeny of females are key properties of genetic mating systems. Female multiple mating should evolve due to direct or indirect benefits, but it may also partly be driven by the encounter rate with different potential mates. 2. In this study this hypothesis was experimentally tested in the European earwig (Forficula auricularia L.) by establishing experimental mating groups that differed in the number of males and females (i.e. density). The number of sires and mean sibling relatedness in each clutch were estimated using microsatellite‐based paternity analysis. 3. As predicted, the mean number of sires per clutch was significantly increased, and sibling relatedness decreased, in the higher density treatment where more potential male mates were available. This change was less than proportional to the number of males in the mating groups, indicating that mechanisms limiting multiple paternity in large mating groups were involved. There were no significant relationships between female reproductive success or male siring success with morphology (body size, weight, and forceps size). 4. The present results show that multiple paternity in F. auricularia clutches is partly determined by the availability of male mates and suggest that this effect is modulated by mechanisms in males and/or females that limit multiple paternity.     

Proportion of exotics and relatedness of host species mediate the positive effect of plant richness on the species richness of fruit flies

1. All else being equal, the greater the local species richness of plants, the greater the number of associated herbivore species. Because most herbivore insects feed on a subset of closely related plant species, plant phylogenetic diversity is expected to play a key role in determining the number of herbivore species. What is not well known, however, is how an increase in the species richness of exotic plants affects the species richness of herbivores. 2. In this study, we used plant–fruit fly interactions to investigate the influence of the proportion and species richness of exotic host plants on the species richness of herbivorous insects. We also tested whether the phylogenetic diversity of host plants increases when the number of exotic plant species increases. 3. We found that the species richness of fruit flies is more accurately predicted by the richness of native host plants than by total plant species richness (including both native and exotic species). The proportion of exotic host species and the phylogenetic diversity of host plants had negative and positive effects, respectively, on the species richness of fruit flies. 4. Our findings suggest that a positive effect of plant richness on herbivore richness occurs only when an increase in plant diversity involves plant species with which native herbivores share some evolutionary history.