SLAH1, a homologue of the slow type anion channel SLAC1, modulates shoot Cl(-) accumulation and salt tolerance in Arabidopsis thaliana

Formally published in vol. 67, no. 15, 2016Salinity tolerance is correlated with shoot chloride (Cl⁻) exclusion in multiple crops, but the molecular mechanisms of long-distance Cl⁻ transport are poorly defined. Here, we characterize the in planta role of AtSLAH1 (a homologue of the slow type anion channel-associated 1 (SLAC1)). This protein, localized to the plasma membrane of root stelar cells, has its expression reduced by salt or ABA, which are key predictions for a protein involved with loading Cl– into the root xylem. Artificial microRNA knockdown mutants of AtSLAH1 had significantly reduced shoot Cl− accumulation when grown under low Cl⁻, whereas shoot Cl– increased and the shoot nitrate/chloride ratio decreased following AtSLAH1 constitutive or stelar-specific overexpression when grown in high Cl–. In both sets of overexpression lines a significant reduction in shoot biomass over the null segregants was observed under high Cl⁻ supply, but not low Cl⁻ supply. Further in planta data showed AtSLAH3 overexpression increased the shoot nitrate/chloride ratio, consistent with AtSLAH3 favouring nitrate transport. Heterologous expression of AtSLAH1 in Xenopus laevis oocytes led to no detectible transport, suggesting the need for post-translational modifications for AtSLAH1 to be active. Our in planta data are consistent with AtSLAH1 having a role in controlling root-to-shoot Cl⁻ transport.Jiaen Qiu, Sam W Henderson, Mark Tester, Stuart J Roy and Mathew Gilliha

Cofinement, entropy, and single-particle dynamics of equilibrium hard-sphere mixtures

We use discontinuous molecular dynamics and grand-canonical transition-matrix Monte Carlo simulations to explore how confinement between parallel hard walls modifies the relationships between packing fraction, self-diffusivity, partial molar excess entropy, and total excess entropy for binary hard-sphere mixtures. To accomplish this, we introduce an efficient algorithm to calculate partial molar excess entropies from the transition-matrix Monte Carlo simulation data. We find that the species-dependent self-diffusivities of confined fluids are very similar to those of the bulk mixture if compared at the same, appropriately defined, packing fraction up to intermediate values, but then deviate negatively from the bulk behavior at higher packing fractions. On the other hand, the relationships between self-diffusivity and partial molar excess entropy (or total excess entropy) observed in the bulk fluid are preserved under confinement even at relatively high packing fractions and for different mixture compositions. This suggests that the partial molar excess entropy, calculable from classical density functional theories of inhomogeneous fluids, can be used to predict some of the nontrivial dynamical behaviors of fluid mixtures in confined environments.Comment: submitted to JC

Effects of environmental variables on invasive amphibian activity: using model selection on quantiles for counts

Many different factors influence animal activity. Often, the value of an environmental variable may influence significantly the upper or lower tails of the activity distribution. For describing relationships with heterogeneous boundaries, quantile regressions predict a quantile of the conditional distribution of the dependent variable. A quantile count model extends linear quantile regression methods to discrete response variables, and is useful if activity is quantified by trapping, where there may be many tied (equal) values in the activity distribution, over a small range of discrete values. Additionally, different environmental variables in combination may have synergistic or antagonistic effects on activity, so examining their effects together, in a modeling framework, is a useful approach. Thus, model selection on quantile counts can be used to determine the relative importance of different variables in determining activity, across the entire distribution of capture results. We conducted model selection on quantile count models to describe the factors affecting activity (numbers of captures) of cane toads (Rhinella marina) in response to several environmental variables (humidity, temperature, rainfall, wind speed, and moon luminosity) over eleven months of trapping. Environmental effects on activity are understudied in this pest animal. In the dry season, model selection on quantile count models suggested that rainfall positively affected activity, especially near the lower tails of the activity distribution. In the wet season, wind speed limited activity near the maximum of the distribution, while minimum activity increased with minimum temperature. This statistical methodology allowed us to explore, in depth, how environmental factors influenced activity across the entire distribution, and is applicable to any survey or trapping regime, in which environmental variables affect activity

Free oxygen radicals regulate plasma membrane Ca2+- and K+-permeable channels in plant root cells

Free oxygen radicals are an irrefutable component of life, underlying important biochemical and physiological phenomena in animals. Here it is shown that free oxygen radicals activate plasma membrane Ca²⁺- and K⁺-permeable conductances in Arabidopsis root cell protoplasts, mediating Ca²⁺ influx and K⁺ efflux, respectively. Free oxygen radicals generate increases in cytosolic Ca²⁺ mediated by a novel population of nonselective cation channels that differ in selectivity and pharmacology from those involved in toxic Na⁺ influx. Analysis of the free oxygen radical-activated K⁺ conductance showed its similarity to the Arabidopsis root K⁺ outward rectifier. Significantly larger channel activation was found in cells responsible for perceiving environmental signals and undergoing elongation. Quenching root free oxygen radicals inhibited root elongation, confirming the role of radical-activated Ca²⁺ influx in cell growth. Net free oxygen radical-stimulated Ca²⁺ influx and K⁺ efflux were observed in root cells of monocots, dicots, C3 and C4 plants, suggesting conserved mechanisms and functions. In conclusion, two functions for free oxygen radical cation channel activation are proposed: initialization/amplification of stress signals and control of cell elongation in root growth.Vadim Demidchik, Sergey N. Shabala, Katherine B. Coutts, Mark A. Tester and Julia M. Davie

Implementing multifactorial psychotherapy research in online virtual environments (IMPROVE-2): study protocol for a phase III trial of the MOST randomized component selection method for internet cognitive-behavioural therapy for depression.

BACKGROUND: Depression is a global health challenge. Although there are effective psychological and pharmaceutical interventions, our best treatments achieve remission rates less than 1/3 and limited sustained recovery. Underpinning this efficacy gap is limited understanding of how complex psychological interventions for depression work. Recent reviews have argued that the active ingredients of therapy need to be identified so that therapy can be made briefer, more potent, and to improve scalability. This in turn requires the use of rigorous study designs that test the presence or absence of individual therapeutic elements, rather than standard comparative randomised controlled trials. One such approach is the Multiphase Optimization Strategy, which uses efficient experimentation such as factorial designs to identify active factors in complex interventions. This approach has been successfully applied to behavioural health but not yet to mental health interventions. METHODS/DESIGN: A Phase III randomised, single-blind balanced fractional factorial trial, based in England and conducted on the internet, randomized at the level of the patient, will investigate the active ingredients of internet cognitive-behavioural therapy (CBT) for depression. Adults with depression (operationalized as PHQ-9 score ≥ 10), recruited directly from the internet and from an UK National Health Service Improving Access to Psychological Therapies service, will be randomized across seven experimental factors, each reflecting the presence versus absence of specific treatment components (activity scheduling, functional analysis, thought challenging, relaxation, concreteness training, absorption, self-compassion training) using a 32-condition balanced fractional factorial design (2IV(7-2)). The primary outcome is symptoms of depression (PHQ-9) at 12 weeks. Secondary outcomes include symptoms of anxiety and process measures related to hypothesized mechanisms. DISCUSSION: Better understanding of the active ingredients of efficacious therapies, such as CBT, is necessary in order to improve and further disseminate these interventions. This study is the first application of a component selection experiment to psychological interventions in depression and will enable us to determine the main effect of each treatment component and its relative efficacy, and cast light on underlying mechanisms, so that we can systematically enhance internet CBT. TRIAL REGISTRATION: Current Controlled Trials ISRCTN24117387 . Registered 26 August 2014.Funding for this trial is provided by grants from the Cornwall NHS Foundation Trust and South West Peninsula Academic Health Research Network to EW. LC is supported by United States National Institutes of Health grants P50DA039838, P01CA180945, R01DK097364, and R01AA022931

Identification of a QTL on chromosome 7AS for sodium exclusion in bread wheat

Tese de doutoramento em Engenharia Física (Instrumentação), apresentada à Faculdade de Ciências e Tecnologia da Universidade de CoimbraO número de ciclotrões com capacidade para acelerar protões até cerca de 20 MeV tem vindo a aumentar em todo o mundo. Apesar de o objectivo principal das instalaçõoes que contêm estas máquinas ser a produção de radionuclídeos para tomografia por emissão de positrões (PET, do inglês positron emission tomography), algumas dessas instalações estão equipadas com várias linhas de feixe que podem ser adaptadas para investigação científica. Por exemplo, radiobiologia, radiofisiolgia e outros estudos de dosimetria podem ser realizados utilizando uma destas linhas de feixe devidamente adaptada para o efeito. Neste trabalho, uma das linhas de feixe do ciclotrão PET da Universidade de Coimbra foi complementada e instrumentada por forma a possibilitar a irradiação de um arranjo experimental com um feixe de protões de elevada qualidade. Esta nova linha de feixe foi construída de raíz e sem causar qualquer interferência com as demais linhas do ciclotrão, dedicadas à produção de radionuclídeos. São apresentados resultados tanto experimentais como de simulação, estes últimos obtidos através dos pacotes de simulação SRIM/TRIM e Geant4, tendo como objectivo a medição do pico de Bragg depositado pelo feixe de protões do ciclotrão PET, com uma energia nominal de 18 MeV. Utilizando um bloco de plástico cintilador e uma câmara fotográfica com ligação à internet da marca D-link foi possível medir tanto o alcance como a largura do feixe de protões com uma resolução espacial inferior a 0,1 mm. Os alcances do feixe medidos após este passar pelo meio de um tubo de alumínio com vácuo com 40 cm de comprimento e um segundo tubo com 2,4 m de comprimento permitiram confirmar que a energia efectiva do feixe é de 18 MeV. Mediu-se também o pico de Bragg utilizando um alvo constituído por várias folhas de alumínio intercaladas com lâminas de polietileno. O sinal de corrente foi recolhido das várias folhas de alumínio através de amplificadores de transimpedância fabricados no âmbito desta tese. Verificou-se que o pico de Bragg assim medido é consistente com simulações realizadas utilizando o pacote SRIM/TRIM. Após a instalação da linha de feixe no perímetro exterior do ciclotrão, esta foi caracterizada, calibrada e validada. Para tal, o sinal induzido pela passagem do feixe por uma folha de alumínio com 20 μm de espessura é lido através do amplificador de transimpedância mencionado. Este sinal amplificado providencia informação de dose em tempo real através de um programa desenvolvido em C/C++. Para além da dose, as principais variáveis de monitorização que este programa providencia incluem a corrente do feixe, a carga integrada em conjunto com a taxa de dose. Deste modo a dose e a corrente integrada (carga total) entregue até um dado instante na montagem experimental pode ser controlada por meio de um obturador controlado por computador. Feixes de protões com correntes tão baixas como 10 pA podem deste modo ser aferidas. A folha de alumínio foi escolhida por ser resistente à radiação, possuir baixa densidade e baixa probabilidade de radioactividade induzida pelo feixe e, finalmente, por representar um custo negligenciável. Junta-se a estas vantagens o facto de o método potenciar o cálculo da dose entregue a um alvo durante uma irradiação, com uma perda mínima da energia do feixe de protões, e com dispersão igualmente mínima. Resultados experimentais e simulações com o Geant4 são apresentados que revelam a aplicação, pela primeira vez, de um feixe de 18 MeV proveniente de um ciclotrão para irradiação de uma região seleccionada de um alvo. Fazendo uso do sistema de dosimetria apresentado no parágrafo anterior foi possível irradiar de modo homogéneo uma região circular com 18 mm de diâmetro. Torna-se assim possível irradiar culturas celulares localizadas em placas multi-poços com um diâmetro por poço de 16 mm, como é usual em experiências de radiobiologia. Verificou-se que o controlo do campo magnético aplicado dentro do ciclotrão é crucial para se obter uma irradiação uniforme em todo o campo do alvo. Para tal, efetua-se antes de cada irradiação e com o obturador fechado, um varrimento à corrente que gera o campo magnético dentro do ciclotrão, medindo-se um perfil quase gaussiano e tomando-se o seu valor central para se obter uma irradiação homogénea. As taxas de dose no alvo (entre 500 mGy/s e 5 mGy/s) são obtidas através de um disco em rotação posicionado na trajectória do feixe. O disco, com 150 mm de raio e uma fenda de 0,5 mm na sua extermidade, permite reduzir a taxa de dose por um factor de 5×10−4. Finalmente, vários filmes do tipo Gafchromic EBT2 foram expostos a diferente valores de dose por forma a validar toda a instalação para irradiação de um alvo com feixes de protões. Para tal validação fez-se uso do sistema de dosimetria em filme 2D do Serviço de Radioterapia do Centro Hospitalar Universitário de Coimbra. A dose absoluta nos filmes irradiados com protões foi verificada neste sistema e apresentou uma precisão melhor que 2%.The number of cyclotrons capable of accelerating protons to about 20MeV is increasing throughout the world. Originally aiming at the production of positron emission tomography (PET) radionuclides, some of these facilities are equipped with several beamlines suitable for scientific research. Radiobiology, radiophysiology, and other dosimetric studies can be performed using these beamlines. In this work, a PET cyclotron was fitted with a long beam transport line to irradiate with a good quality proton beam experimental setups. The beamline was configured as a natural extension of one of the cyclotron beam ports, while keeping available the other beam ports for PET radionuclides production. Experimental results are reported, together with SRIM/TRIM and Geant4 simulations, which aim at measuring the Bragg peak of the 18 -MeV proton beam from the PET cyclotron. By using a piece of plastic scintillator and a D-link Ethernet-based camera, the proton beam range and width were measured with a spatial resolution of 0.1mm. The ranges of the proton beam in the plastic scintillator were used to assess its energy after trespassing one or two Havar R windows and either a 40-cm-long or a 2.4-m-long aluminum pipe. The initial energy of the proton beam from the PET cyclotron was found to be 18 MeV. Additionally, the Bragg peak of the protons from the PET cyclotron was assessed using a stacked target consisting of several aluminum foils interleaved with polyethylene sheets, readout by in-house made transimpedance electronics. The measured Bragg peak is consistent with simulations performed using the SRIM/TRIM simulation toolkit. An out-of-yoke irradiation setup using the accelerated proton beam coming from the PET cyclotron was developed, characterized, calibrated, and validated. A 20-μm-thick aluminum transmission foil is readout by in-house made transimpedance electronics, providing online dose information via a C/C++ program. xvii Overview The main monitoring variables include beam current, integrated charge together with dose rate. The beam monitor is able to readout and deliver these variables in real-time. Hence the dose and integrated current (total charge) delivered upto a given instant to an experimental setup may be controlled via a computercontrolled shutter that was installed in the beam path. Proton beam currents down to 10 pA can be assessed using the thin aluminum foil. The aluminum was chosen for this task because it is radiation hard, it has low density and low radiation activity, and finally because it is easily available at negligible cost. In addition, this method allows for calculating the dose delivered to a target during an irradiation with high efficiency, and with minimal proton energy loss and scattering. Experimental results and Geant4 simulations are reported, which aim at using for the first time the 18 -MeV proton beam from a PET cyclotron to irradiate a selected region of a target using the developed dosimetry system. By using this system, a homogeneous beam spot on target with a diameter of 18mm can be controlled. This allows controlled irradiation of cell cultures located in typical biological multi-well dishes with diameters of 16mm each. It was found that the control of the magnetic field applied inside the cyclotron plays a major role for achieving said homogeneity. For that, scanning the magnet current and measuring the corresponding dose rate reveals a quasi-Gaussian shaped curve that must be known before any irradiation procedure (the final shutter is closed during such measurements). The optimum magnet current is taken from the center of the Gaussian-shaped curve, hence producing a homogenous dose on target. The measured dose rate on target ranges from 500mGy/s down to 5mGy/s. This is achieved with a 150mm radius rotating disk with a slit of 0.5mm width, that decreases target dose rates by a factor of 5 × 10−4. Several Gafchromic R EBT2 films were exposed to different values of dose for validating the developed irradiation setup using the 2D film dosimetry system of the Department of Radiotherapy of Coimbra University Hospital Center. The absolute dose in the irradiated films were assessed with a precision better than 2%. It is planned, in the near future, to irradiate small animals, cell cultures, or other materials or samples

Acoustic Investigation of Jet Mixing Noise in Dual Stream Nozzles

In an earlier study, a prediction model for jet noise in dual stream jets was proposed that is founded on velocity scaling laws in single stream jets and similarity features of the mean velocity and turbulent kinetic energy in dual stream flows. The model forms a composite spectrum from four component single-stream jets each believed to represent noise-generation from a distinct region in the actual flow. While the methodology worked effectively at conditions considered earlier, recent examination of acoustic data at some unconventional conditions indicate that further improvements are necessary in order to expand the range of applicability of the model. The present work demonstrates how these predictions compare with experimental data gathered by NASA and industry for the purpose of examining the aerodynamic and acoustic performance of such nozzles for a wide range of core and fan stream conditions. Of particular interest are jets with inverted velocity and temperature profiles and the appearance of a second spectral peak at small aft angles to the jet under such conditions. It is shown that a four-component spectrum succeeds in modeling the second peak when the aft angle refraction effects are properly incorporated into the model. A tradeoff of noise emission takes place between two turbulent regions identified as transition and fully mixed regions as the fan stream velocity exceeds that of the core stream. The effect of nozzle discharge coefficients will also be discussed

Novel Characteristics of Valveless Pumping

This study investigates the occurrence of valveless pumping in a fluidfilled system consisting of two open tanks connected by an elastic tube. We show that directional flow can be achieved by introducing a periodic pinching applied at an asymmetrical location along the tube, and that the flow direction depends on the pumping frequency. We propose a relation between wave propagation velocity, tube length, and resonance frequencies associated with shifts in the pumping direction using numerical simulations. The eigenfrequencies of the system are estimated from the linearized system, and we show that these eigenfrequencies constitute the resonance frequencies and the horizontal slope frequencies of the system; 'horizontal slope frequency' being a new concept. A simple model is suggested, explaining the effect of the gravity driven part of the oscillation observed in response to the tank and tube diameter changes. Results are partly compared with experimental findings.Art. no. 22450

AtNPF2.5 modulates chloride (Cl -bar) efflux from roots of Arabidopsis thaliana

The accumulation of high concentrations of chloride (Cl(-)) in leaves can adversely affect plant growth. When comparing different varieties of the same Cl(-) sensitive plant species those that exclude relatively more Cl(-) from their shoots tend to perform better under saline conditions; however, the molecular mechanisms involved in maintaining low shoot Cl(-) remain largely undefined. Recently, it was shown that the NRT1/PTR Family 2.4 protein (NPF2.4) loads Cl(-) into the root xylem, which affects the accumulation of Cl(-) in Arabidopsis shoots. Here we characterize NPF2.5, which is the closest homolog to NPF2.4 sharing 83.2% identity at the amino acid level. NPF2.5 is predominantly expressed in root cortical cells and its transcription is induced by salt. Functional characterisation of NPF2.5 via its heterologous expression in yeast (Saccharomyces cerevisiae) and Xenopus laevis oocytes indicated that NPF2.5 is likely to encode a Cl(-) permeable transporter. Arabidopsis npf2.5 T-DNA knockout mutant plants exhibited a significantly lower Cl(-) efflux from roots, and a greater Cl(-) accumulation in shoots compared to salt-treated Col-0 wild-type plants. At the same time, [Formula: see text] content in the shoot remained unaffected. Accumulation of Cl(-) in the shoot increased following (1) amiRNA-induced knockdown of NPF2.5 transcript abundance in the root, and (2) constitutive over-expression of NPF2.5. We suggest that both these findings are consistent with a role for NPF2.5 in modulating Cl(-) transport. Based on these results, we propose that NPF2.5 functions as a pathway for Cl(-) efflux from the root, contributing to exclusion of Cl(-) from the shoot of Arabidopsis.Bo Li, Jiaen Qiu, Maheswari Jayakannan, Bo Xu, Yuan Li, Gwenda M. Mayo, Mark Tester, Matthew Gilliham and Stuart J. Ro