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31.
Helena Grgi? Marcio Costa Robert M. Friendship Susy Carman éva Nagy Zvonimir Poljak 《PloS one》2015,10(6)
The objective of this study was to characterize H1N1 and H1N2 influenza A virus isolates detected during outbreaks of respiratory disease in pig herds in Ontario (Canada) in 2012. Six influenza viruses were included in analysis using full genome sequencing based on the 454 platform. In five H1N1 isolates, all eight segments were genetically related to 2009 pandemic virus (A(H1N1)pdm09). One H1N2 isolate had hemagglutinin (HA), polymerase A (PA) and non-structural (NS) genes closely related to A(H1N1)pdm09, and neuraminidase (NA), matrix (M), polymerase B1 (PB1), polymerase B2 (PB2), and nucleoprotein (NP) genes originating from a triple-reassortant H3N2 virus (tr H3N2). The HA gene of five Ontario H1 isolates exhibited high identity of 99% with the human A(H1N1)pdm09 [A/Mexico/InDRE4487/09] from Mexico, while one Ontario H1N1 isolate had only 96.9% identity with this Mexican virus. Each of the five Ontario H1N1 viruses had between one and four amino acid (aa) changes within five antigenic sites, while one Ontario H1N2 virus had two aa changes within two antigenic sites. Such aa changes in antigenic sites could have an effect on antibody recognition and ultimately have implications for immunization practices. According to aa sequence analysis of the M2 protein, Ontario H1N1 and H1N2 viruses can be expected to offer resistance to adamantane derivatives, but not to neuraminidase inhibitors. 相似文献
32.
Biotin carboxylase catalyzes the ATP-dependent carboxylation of biotin and is one component of the multienzyme complex acetyl-CoA carboxylase that catalyzes the first committed step in fatty acid synthesis in all organisms. In Escherichia coli, biotin carboxylase exists as a homodimer where each subunit contains a complete active site. In a previous study (Janiyani, K., Bordelon, T., Waldrop, G.L., Cronan Jr., J.E., 2001. J. Biol. Chem. 276, 29864-29870), hybrid dimers were constructed where one subunit was wild-type and the other contained an active site mutation that reduced activity at least 100-fold. The activity of the hybrid dimers was only slightly greater than the activity of the mutant homodimers and far less than the expected 50% activity for completely independent active sites. Thus, there is communication between the two subunits of biotin carboxylase. The dominant negative effect of the mutations on the wild-type active site was interpreted as alternating catalytic cycles of the active sites in the homodimer. In order to test the hypothesis of oscillating catalytic cycles, mathematical modeling and numerical simulations of the kinetics of wild-type, hybrid dimers, and mutant homodimers of biotin carboxylase were performed. Numerical simulations of biotin carboxylase kinetics were the most similar to the experimental data when an oscillating active site model was used. In contrast, alternative models where the active sites were independent did not agree with the experimental data. Thus, the numerical simulations of the proposed kinetic model support the hypothesis that the two active sites of biotin carboxylase alternate their catalytic cycles. 相似文献
33.
Morin C Sirois M Echave V Gomes MM Rousseau E 《Prostaglandins & other lipid mediators》2007,83(4):311-319
The present study investigated the ability of 5-oxo-EicosaTetraEnoic acid (5-oxo-ETE) for modulating airway smooth muscle (ASM) tone in human bronchi. 5-Oxo-ETE induced a concentration-dependent relaxing effect on human bronchi pre-contracted with methacholine (MCh) and arachidonic acid (AA). This relaxing response was highly sensitive to Iberiotoxin (IbTx), a large conducting Ca(2+)-activated K(+) channel (BK(Ca)) inhibitor. Furthermore, microelectrode measurements revealed that 5-oxo-ETE (0.1-10 microM) hyperpolarizes the membrane potential of human bronchial ASM cells. These hyperpolarizing effects were also inhibited in the presence of 10nM IbTx. Lastly, 5-oxo-ETE was shown to directly activate reconstituted BK(Ca) channels derived from human airway smooth muscles. In summary, the 5-oxo-ETE eicosanoid activates a specific K(+) conductance, involved in membrane hyperpolarization, which in turn reduces Ca(2+) entry and facilitates relaxation of smooth muscle cells. 相似文献
34.
International Journal of Primatology - Primates might be particularly vulnerable to experiencing adverse effects from climate change, given their level of exposure, sensitivity to climatic... 相似文献
35.
36.
Indicators of restoration success in riparian tropical forests using multiple reference ecosystems
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Forest restoration by planting trees often accelerates succession, but the trajectories toward reference ecosystems have rarely been evaluated. Using a chronosequence (4–53 years) of 26 riparian forest undergoing restoration in the Brazilian Atlantic Forest, we modeled how the variables representing forest structure, tree species richness and composition, and the proportion of plant functional guilds change through time. We also estimated the time required for these variables to reach different types of reference ecosystems: old‐growth forest (OGF), degraded forest, and secondary forest. Among the attributes which follow a predictable trajectory over time are: the basal area, canopy cover, density and tree species richness, as well as proportions of shade tolerant and slow growing species or individuals. Most of the variation in density of pteridophythes, lianas, shrubs and phorophythes, proportion of animal‐dispersed individuals, rarefied richness and floristic similarity with reference ecosystems remain unexplained. Estimated time to reach the reference ecosystems is, in general, shorter for structural attributes than for species composition or proportion of functional guilds. The length of this time varies among the three types of reference ecosystems for most attributes. For instance, tree species richness and proportion of shade tolerant or slow growing individuals become similar to secondary forests in about 40 years, but is estimated to take 70 years or more to reach the OGF. Of all the variables considered, canopy cover, basal area, density, and richness of the understory—by their ecological relevance and predictability—are recommended as ecological indicators for monitoring tropical forest restoration success. 相似文献
37.
The enzyme biotin carboxylase (BC) uses adenosine triphosphate (ATP) to carboxylate biotin and is involved in fatty acid synthesis. Structural evidence suggests that the B domain of BC undergoes a large hinge motion of ~45° when binding and releasing substrates. Escherichia coli BC can function as a natural homodimer and as a mutant monomer. Using molecular dynamics simulations, we evaluate the free energy profile along a closure angle of the B domain of E. coli BC for three cases: a monomer without bound Mg(2)ATP, a monomer with bound Mg(2)ATP, and a homodimer with bound Mg(2)ATP in one subunit. The simulation results show that a closed state is the most probable for the monomer with or without bound Mg(2)ATP. For the dimer with Mg(2)ATP in one of its subunits, communication between the two subunits was observed. Specifically, in the dimer, the opening of the subunit without Mg(2)ATP caused the other subunit to open, and hysteresis was observed upon reclosing it. The most stable state of the dimer is one in which the B domain of both subunits is closed; however, the open state for the B domain without Mg(2)ATP is only approximately 2k(B)T higher in free energy than the closed state. A simple diffusion model indicates that the mean times for opening and closing of the B domain in the monomer with and without Mg(2)ATP are much smaller than the overall reaction time, which is on the order of seconds. 相似文献
38.
Michael Sandmann Kamil Sk?odowski Pawel Gajdanowicz Erwan Michard Marcio Rocha Judith L Gomez-Porras Wendy González Luiz Gustavo Guedes Corraa Santiago J Ramírez-Aguilar Tracey Ann Cuin Joost T van Dongen Jean-Baptiste Thibaud Ingo Dreyer 《Plant signaling & behavior》2011,6(4):558-562
Potassium (K+) is an important nutrient for plants. It serves as a cofactor of various enzymes and as the major inorganic solute maintaining plant cell turgor. In a recent study, an as yet unknown role of K+ in plant homeostasis was shown. It was demonstrated that K+ gradients in vascular tissues can serve as an energy source for phloem (re)loading processes and that the voltage-gated K+ channels of the AKT2-type play a unique role in this process. The AKT2 channel can be converted by phosphorylation of specific serine residues (S210 and S329) into a non-rectifying channel that allows a rapid efflux of K+ from the sieve element/companion cells (SE/CC) complex. The energy of this flux is used by other transporters for phloem (re)loading processes. Nonetheless, the results do indicate that post-translational modifications at S210 and S329 alone cannot explain AKT2 regulation. Here, we discuss the existence of multiple post-translational modification steps that work in concert to convert AKT2 from an inward-rectifying into a non-rectifying K+ channel.Key words: potassium, channel, potassium channel, AKT2, phloem (re)loading, post-translational modifications, potassium batteryPotassium (K+) is the most abundant mineral element in plants, and together with nitrogen and phosphorous, is limiting for plant production in many natural and agricultural habitats. Voltage-gated K+ channels are key players in the acquisition of K+ ions from the soil and in its redistribution within the plant.1 Structurally, these channels result from the assembly of four so-called α-subunits. The subunits are encoded by nine genes in Arabidopsis and both homo- and hetero-tetramers are expressed.2,3 The K+ channel α-subunits can be categorized into four different subfamilies, based on the voltage-gating characteristics of the exogenous K+ conductance when expressed in an appropriate heterologous expression system. Kin α-subunits form hyperpolarization-activated channels that mediate K+ uptake.4–7 Kout α-subunits form depolarization-activated channels that mediate K+ release from cells.8–10 Ksilent subunits appear unable to yield functional homomeric channels, but can combine with Kin subunits and fine-tune the K+-uptake properties of the resulting heteromeric channels.11–14 Finally, Kweak α-subunits form channels with complex voltage-gating; they allow both K+ uptake and release.15–19 In Arabidopsis, a single member is found in this subfamily, AKT2, and this channel can assemble in heteromeric channels with the Kin subunit KAT2.20To date, only scarce and speculative information has been obtained for the function of Kweak channels. When expressed in heterologous expression systems, two different subpopulations of AKT2 channels differing in their sensitivity to voltage were found.21 Channels of the first type showed gating properties and currents analogous to that of Kin channels, while the other sort enabled a non-rectified (leak-like) current; they were open over the entire physiological voltage range.A given channel can be converted from one type to the other by post-translational modifications.21 A voltage-dependent phosphorylation was found to be an essential step for this switch,22,23 although the kinase responsible for this conversion still needs to be uncovered.24 In biophysical studies, mutant versions of the Arabidopsis Kweak channel subunit AKT2 have been created that showed impaired gating mode settings.22,23 Recently, Gajdanowicz et al. generated transgenic Arabidopsis thaliana plants that express these mutant AKT2 channels in the background of the akt2-1 null-allele plant.25 The major conclusion from analyses of these mutants is that the status switching of AKT2 from an inward-rectifying to a non-rectifying channel is crucial for plants to overcome energy-limiting conditions. This function of AKT2 could be correlated to its expression in phloem tissues. Selective expression of AKT2 under the control of the phloem companion cell-specific AtSUC2 promoter rescued the akt2-1 line, but conversely, selective expression of AKT2 under the control of the guard cell-specific GC1 promoter,26 resulted in further impairment of plant growth (Fig. 1). By combining diverse experimental approaches with mathematical simulation methods, an existing model for phloem (re)loading18,27 was fundamentally improved. This allowed the uncovering of a novel and interesting role of K+ in phloem physiology: K+ gradients present between the sieve element/companion cell (SE/CC) complex and the apoplast can serve as an energy source in phloem (re)loading processes. This “potassium battery” can be tapped by means of AKT2 regulation. This clarifies the observation of Deeken et al.28 that in AKT2 loss-of-function mutant plants, assimilates leaking away from the sieve tube were not efficiently reloaded into the main phloem stream.Open in a separate windowFigure 1AKT2 expressed only in guard cells delays plant development. (A–C) Representative wild-type, akt2-1 and akt2-1+pGC1:AKT2 complementation plants grown for 7 weeks (A), 9 weeks (B) and 12 weeks (C) under 12-h day/12-h night conditions at normal light intensity (150 µmol m−2 s−1). (D) akt2-1+pGC1:AKT2 developed a similar number of leaves as the akt2-1 knock out plants, but bolting-time was delayed. (B and E) After 9 weeks, wild-type plants were at an advanced bolting stage, akt2-1 plants had started bolting, but only initial signs of bolting were visible in akt2-1+pGC1:AKT2 plants. (C and F) At 12 weeks, akt2-1 plants had caught up with the wild-type and akt2-1+pGC1:AKT2 was just starting to bolt, although rosette-leaves were showing clear signs of senescence. For the generation of akt2-1+pGC1:AKT2, the AKT2 cDNA was fused to the guard cell-specific GC1 promoter26 kindly provided by J.I. Schroeder, San Diego. The pGC1:AKT2 construct was cloned into pGreen0229-35S by replacing the 35S promoter and then transformed into the akt2-1 knockout plant. All seeds were cold-treated for 24 h at 4°C. Plants were grown on artificial substrate (type GS-90, Einheitserde). After 2 weeks, seedlings were transferred to single pots. Plants were grown in 60% relative humidity at 21°C during the day and 18°C at night. Phenotypical analyses were done in the middle of the day. Data are shown as means ± SD of n ≥ 9 plants. Statistical analyses using Student''s t test: (D, WT/akt2-1: p < 2e-08; D, WT/pGC-AKT2: p < 2e-08; D, akt2-1/pGC-AKT2: p < 5e-03; E, WT/akt2-1: p < 4e-06; E, WT/pGC-AKT2: p < 1e-10; E, akt2-1/pGC-AKT2: p < 5e-04; F, WT/akt2-1: p = 0.51; F, WT/pGC-AKT2: p < 1e-10; F, akt2-1/pGC-AKT2: p < 1e-10).AKT2 expression is especially abundant in phloem tissues and the root stele, both of which are characterized by a poor availability of oxygen.29,30 This local internal hypoxia impairs respiratory activity of the vascular tissue and concomitantly, respiratory ATP production is reduced.31 As a consequence, phloem transport is very susceptible to decreasing oxygen supply to the plant.29,32 It is therefore comprehensible that the above mentioned support by the K+ driving force for sucrose retrieval is especially relevant in the phloem. Indeed Gajdanowicz et al.25 showed that transgenic plants lacking the AKT2 K+ channel were severely impaired in growth when exposed to mild hypoxia (10% v:v), whereas growth of wild-type plants was unaffected by this treatment. These observations illustrate the importance of biochemical flexibility in plant cells to cope with the energetic consequences of the steep oxygen concentration gradients that generally occur in plant stems and roots.In fact, the role of K+ gradients in driving sugar, amino acid and organic acid transport across plant cell membranes was first suggested several decades ago.33,34 Experimental evidence for this concept was provided by various tests in which pieces of plant tissue were incubated in solutions with different K+ concentrations and pH levels.33,34 Unfortunately, at that time the lack of genetic information to support this hypothesis (e.g., identifying transporter proteins that could provide a molecular mechanism to explain the working mechanism of substrate transport driven by a K+-motive force) resulted in this idea falling into oblivion. Indeed, the unequivocal experimental observation of this new role of K+ gradients in phloem reloading is extremely challenging. Under normal experimental conditions, K+ fluxes and sucrose fluxes are coupled during phloem loading in source tissues and unloading in sink tissues. Nonetheless, computational simulations predict that under certain conditions, a local K+/Suc antiport is also thermodynamically possible. In this antiport system, the energy from the K+ gradient is used to transport Suc into the phloem. This process is only transient; flooding the apoplast with K+ will decrease the K+ gradient. However, the gradient can be maintained for longer if surrounding cells take up the apoplastic K+ for their own use. A K+/Suc antiport will not occur in obvious sink or source tissues since the energy balances in such cells are fundamentally different. Consequently, in these tissues only the coupled symport of K+ and Suc can be observed. However, the computational predictions allowed the identification of the experimental conditions under which the effect of the K+/Suc antiport system is empirically observable at the whole plant level.An essential role in the regulation of AKT2 is played by (de)phosphorylation events of serine residues at positions S210 and S329. The replacement of both serines by asparagine (AKT2-S210N-S329N) resulted in a K+-selective leak that is locked in a continuously open mode when the channels are expressed in Xenopus oocytes. Under certain conditions, plants expressing the AKT2-S210N-S329N mutation showed growth benefits over wild-type plants; akt2-1+AKT2-S210N-S329N plants reach the generative state faster, possess an increased number of leaves and increased fresh weight (Fig. 2). Intuitively, one would expect a continuously open channel to cause severe problems for the plant, not a benefit as was observed here. We therefore have to postulate that phosphorylation at residues AKT2-S210 and AKT2-S329 is insufficient for converting AKT2 from an inward-rectifying into a non-rectifying channel; other, as yet unknown mechanisms, must contribute to the switch in the AKT2 gating mode. Such a concept would correspond to results that would otherwise be hard to explain. For instance, when both serine residues were replaced by glutamate, the mutant AKT2-S210E-S329E still showed wild-type characteristics.22 The S to E substitution is expected to mimic the phosphorylated state better than the S to N replacement. Furthermore, position AKT2-K197 has a fundamental influence on the AKT2 gating mode.23 AKT2 mutants with that particular lysine substituted with a serine are far less sensitive towards (de)phosphorylation; they display the characteristics of a pure inward-rectifying K+ channel,23 and transgenic Arabidopsis plants expressing AKT2 channels with this substitution showed the characteristics of akt2-1 knock-out plants.25 Initially, it was proposed that the positive charge is important for sensitizing AKT2 to phosphorylation. However, the charge-conserving mutant AKT2-K197R is similar to the charge inverting mutant AKT2-K197D,23 a purely inward-rectifying channel (Fig. 3). We therefore need to take into account that in plants, K197 may also be a target of post-translational modification.35 At present, we can explain the beneficial effect of the AKT2-S210N-S329N mutant on plant growth only by a multiple step regulation of AKT2 (Fig. 4). The double-N mutation would then bypass the phosphorylation step, but AKT2-S210N-S329N could still be deregulated into an inward-rectifying channel. Thus, AKT2 can be considered as a highly specialized Kin channel that can be converted into a leak-like channel by a cascade of post-translational modification steps.Open in a separate windowFigure 2Plants expressing the AKT2-S210N-S329N mutant reach the generative state faster than wild-type plants. The mutant channel AKT2-S210N-S329N was expressed under the control of the native AKT2 promoter in the akt2-1 knock-out background. (A) Photos of representative Arabidopsis thaliana plants grown 7 weeks under short day conditions (12-h day/12-h night, light intensity = 150 µE m−2s−1). Seven weeks after sowing, plants expressing only AKT2-S210N-S329N mutant channels (n = 22) differed significantly (Student''s t test, p < 4e-05) from wild-type plants (n = 20) in the height of the main inflorescent stalk (B) and fresh weight (C). At later time points, these differences decrease.25Open in a separate windowFigure 3The mutant AKT2-K197R channel is inward-rectifying. Steady-state current-voltage characteristics measured at the end of activation voltage steps. Currents were normalized to the current values measured at −145 mV in 10 mM K+ and are shown as means ± SD (n = 6).Open in a separate windowFigure 4Minimal model for AKT2 gating-mode regulation. To switch AKT2 from an inward-rectifying into a non-rectifying channel, at least two post-translational steps are postulated. (1) Phosphorylation at residues AKT2-S210 and AKT2-S329 (transitions [1]→[2] and [3]→[4]) and (2) a yet unknown modification that most likely involves the residue AKT2-K197 (transitions [1]→[3] and [2]→[4]). Only after both modifications will AKT2 allow the efflux of K+ (state [4]). 相似文献
39.
Chi BH Yiannoutsos CT Westfall AO Newman JE Zhou J Cesar C Brinkhof MW Mwango A Balestre E Carriquiry G Sirisanthana T Mukumbi H Martin JN Grimsrud A Bacon M Thiebaut R;International Epidemiologic Databases to Evaluate AIDS Collaboration 《PLoS medicine》2011,8(10):e1001111
Background
Although patient attrition is recognized as a threat to the long-term success of antiretroviral therapy programs worldwide, there is no universal definition for classifying patients as lost to follow-up (LTFU). We analyzed data from health facilities across Africa, Asia, and Latin America to empirically determine a standard LTFU definition.Methods and Findings
At a set “status classification” date, patients were categorized as either “active” or “LTFU” according to different intervals from time of last clinic encounter. For each threshold, we looked forward 365 d to assess the performance and accuracy of this initial classification. The best-performing definition for LTFU had the lowest proportion of patients misclassified as active or LTFU. Observational data from 111 health facilities—representing 180,718 patients from 19 countries—were included in this study. In the primary analysis, for which data from all facilities were pooled, an interval of 180 d (95% confidence interval [CI]: 173–181 d) since last patient encounter resulted in the fewest misclassifications (7.7%, 95% CI: 7.6%–7.8%). A secondary analysis that gave equal weight to cohorts and to regions generated a similar result (175 d); however, an alternate approach that used inverse weighting for cohorts based on variance and equal weighting for regions produced a slightly lower summary measure (150 d). When examined at the facility level, the best-performing definition varied from 58 to 383 d (mean = 150 d), but when a standard definition of 180 d was applied to each facility, only slight increases in misclassification (mean = 1.2%, 95% CI: 1.0%–1.5%) were observed. Using this definition, the proportion of patients classified as LTFU by facility ranged from 3.1% to 45.1% (mean = 19.9%, 95% CI: 19.1%–21.7%).Conclusions
Based on this evaluation, we recommend the adoption of ≥180 d since the last clinic visit as a standard LTFU definition. Such standardization is an important step to understanding the reasons that underlie patient attrition and establishing more reliable and comparable program evaluation worldwide. Please see later in the article for the Editors'' Summary 相似文献40.
Analysis of the NAC transcription factor gene family in citrus reveals a novel member involved in multiple abiotic stress responses 总被引:1,自引:0,他引:1