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

The case for evidence-based policy to support stress-resilient cropping systems

Research and the dissemination of evidence-based guidelines for best practice in crop production are fundamental for the protection of our crop yields against biotic and abiotic threats, and for meeting ambitious food production targets by 2050. The advances in knowledge required for sustaining crop productivity targets will be gained through three research tracks: (1) basic strategic research in the field, for example, crop breeding, agronomy, and advanced phenotyping; (2) translational research involving the application of advances in fundamental science; and (3) pure fundamental research to fuel future translational research. We propose that policy and funding structures need to be improved to facilitate and encourage more interactions between scientists involved in all three research tracks, and also between researchers and farmers, to improve the effectiveness of delivering improvements in crop stress resilience. History illustrates that it is challenging for public researchers to “stretch across” all of these research tracks, with effective farm-level solutions being more likely when end-users and industry are directly engaged in the research pipeline. As research proceeds from fundamental through to applied research, the demand for experimental rigor and a wider understanding of appropriate methods and outcomes is paramount, that is, demonstrating value in yield at the field level requires the input of experienced practitioners from each research track. The development of evidence-based policies to support all funding structures and the engagement of producers with both the development of research, and with the findings of such research, will form an important capability in meeting food security targets. This commentary, concentrating on the development of policies to support research and its dissemination, is based on discussions held at the Stress Resilience Symposium organized by the Global Plant Council and Society of Experimental Biology in October 2015.Matthew Gilliham, Scott Chapman, Lisa Martin, Sarah Jose and Ruth Basto

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

Heterodimerization of Arabidopsis calcium/proton exchangers contributes to regulation of guard cell dynamics and plant defense responses

Arabidopsis thaliana cation exchangers (CAX1 and CAX3) are closely related tonoplast-localized calcium/proton (Ca²⁺/H⁺) antiporters that contribute to cellular Ca²⁺ homeostasis. CAX1 and CAX3 were previously shown to interact in yeast; however, the function of this complex in plants has remained elusive. Here, we demonstrate that expression of CAX1 and CAX3 occurs in guard cells. Additionally, CAX1 and CAX3 are co-expressed in mesophyll tissue in response to wounding or flg22 treatment, due to the induction of CAX3 expression. Having shown that the transporters can be co-expressed in the same cells, we demonstrate that CAX1 and CAX3 can form homomeric and heteromeric complexes in plants. Consistent with the formation of a functional CAX1-CAX3 complex, CAX1 and CAX3 integrated into the yeast genome suppressed a Ca²⁺-hypersensitive phenotype of mutants defective in vacuolar Ca²⁺ transport, and demonstrated enzyme kinetics different from those of either CAX protein expressed by itself. We demonstrate that the interactions between CAX proteins contribute to the functioning of stomata, because stomata were more closed in cax1-1, cax3-1, and cax1-1/cax3-1 loss-of-function mutants due to an inability to buffer Ca²⁺ effectively. We hypothesize that the formation of CAX1-CAX3 complexes may occur in the mesophyll to affect intracellular Ca²⁺ signaling during defense responses.Bradleigh Hocking, Simon J. Conn, Murli Manohar, Bo Xu, Asmini Athman, Matthew A. Stancombe, Alex R. Webb, Kendal D. Hirschi and Matthew Gilliha

Plants fighting back: to transport or not to transport, this is a structural question

Abstract not availableMaria Hrmova and Matthew Gilliha

Effectiveness and cost-effectiveness of implementing HIV testing in primary care in East London: protocol for an interrupted time series analysis

Introduction HIV remains underdiagnosed. Guidelines recommend routine HIV testing in primary care, but evidence on implementing testing is lacking. In a previous study, the Rapid HIV Assessment 2 (RHIVA2) cluster randomised controlled trial, we showed that providing training and rapid point-of-care HIV testing at general practice registration (RHIVA2 intervention) in Hackney led to cost-effective, increased and earlier diagnosis of HIV. However, interventions effective in a trial context may be less so when implemented in routine practice. We describe the protocol for an MRC phase IV implementation programme, evaluating the impact of rolling out the RHIVA2 intervention in a post-trial setting. We will use a longitudinal study to examine if the post-trial implementation in Hackney practices is effective and cost-effective, and a cross-sectional study to compare Hackney with two adjacent boroughs providing usual primary care (Newham) and an enhanced service promoting HIV testing in primary care (Tower Hamlets). Methods and analysis Service evaluation using interrupted time series and cost-effectiveness analyses. We will include all general practices in three contiguous high HIV prevalence East London boroughs. All adults aged 16 and above registered with the practices will be included. The interventions to be examined are: a post-trial RHIVA2 implementation programme (including practice-based education and training, external quality assurance, incentive payments for rapid HIV testing and incorporation of rapid HIV testing in the sexual health Local Enhanced Service) in Hackney; the general practice sexual health Network Improved Service in Tower Hamlets and usual care in Newham. Coprimary outcomes are rates of HIV testing and new HIV diagnoses. Ethics and dissemination The chair of the Camden and Islington NHS Research Ethics Committee, London, has endorsed this programme as an evaluation of routine care. Study results will be published in peer-reviewed journals and reported to commissioners

Linking metabolism to membrane signaling: the GABA–malate connection

γ-Aminobutyric acid (GABA) concentration increases rapidly in tissues when plants encounter abiotic or biotic stress, and GABA manipulation affects growth. This, coupled to GABA's well-described role as a neurotransmitter in mammals, led to over a decade of speculation that GABA is a signal in plants. The discovery of GABA-regulated anion channels in plants provides compelling mechanistic proof that GABA is a legitimate plant-signaling molecule. Here we examine research avenues unlocked by this finding and propose that these plant 'GABA receptors' possess novel properties ideally suited to translating changes in metabolic status into physiological responses. Specifically, we suggest they have a role in signaling altered cycling of tricarboxylic acid (TCA) intermediates during stress via eliciting changes in electrical potential differences across membranes.Matthew Gilliham and Stephen D. Tyerma

Plant High-Affinity Potassium (HKT) transporters involved in salinity tolerance: structural insights to probe differences in ion selectivity

High-affinity Potassium Transporters (HKTs) belong to an important class of integral membrane proteins (IMPs) that facilitate cation transport across the plasma membranes of plant cells. Some members of the HKT protein family have been shown to be critical for salinity tolerance in commercially important crop species, particularly in grains, through exclusion of Na+ ions from sensitive shoot tissues in plants. However, given the number of different HKT proteins expressed in plants, it is likely that different members of this protein family perform in a range of functions. Plant breeders and biotechnologists have attempted to manipulate HKT gene expression through genetic engineering and more conventional plant breeding methods to improve the salinity tolerance of commercially important crop plants. Successful manipulation of a biological trait is more likely to be effective after a thorough understanding of how the trait, genes and proteins are interconnected at the whole plant level. This article examines the current structural and functional knowledge relating to plant HKTs and how their structural features may explain their transport selectivity. We also highlight specific areas where new knowledge of plant HKT transporters is needed. Our goal is to present how knowledge of the structure of HKT proteins is helpful in understanding their function and how this understanding can be an invaluable experimental tool. As such, we assert that accurate structural information of plant IMPs will greatly inform functional studies and will lead to a deeper understanding of plant nutrition, signalling and stress tolerance, all of which represent factors that can be manipulated to improve agricultural productivity.Shane Waters, Matthew Gilliham and Maria Hrmov

Chloride on the move

Chloride (Cl-) is an essential plant nutrient but under saline conditions it can accumulate to toxic levels in leaves; limiting this accumulation improves the salt tolerance of some crops. The rate-limiting step for this process - the transfer of Cl- from root symplast to xylem apoplast, which can antagonize delivery of the macronutrient nitrate (NO3-) to shoots - is regulated by abscisic acid (ABA) and is multigenic. Until recently the molecular mechanisms underpinning this salt-tolerance trait were poorly defined. We discuss here how recent advances highlight the role of newly identified transport proteins, some that directly transfer Cl- into the xylem, and others that act on endomembranes in 'gatekeeper' cell types in the root stele to control root-to-shoot delivery of Cl-.Bo Li, Mark Tester and Matthew Gilliha

Plant transporters involved in combating boron toxicity: beyond 3D structures

Version of Record published: 11 August 2020Membrane transporters control the movement and distribution of solutes, including the disposal or compartmentation of toxic substances that accumulate in plants under adverse environmental conditions. In this minireview, in the light of the approaching 100th anniversary of unveiling the significance of boron to plants (K. Warington, 1923; Ann. Bot.37, 629) we discuss the current state of the knowledge on boron transport systems that plants utilise to combat boron toxicity. These transport proteins include: (i) nodulin-26-like intrinsic protein-types of aquaporins, and (ii) anionic efflux (borate) solute carriers. We describe the recent progress made on the structure–function relationships of these transport proteins and point out that this progress is integral to quantitative considerations of the transporter's roles in tissue boron homeostasis. Newly acquired knowledge at the molecular level has informed on the transport mechanics and conformational states of boron transport systems that can explain their impact on cell biology and whole plant physiology. We expect that this information will form the basis for engineering transporters with optimised features to alleviate boron toxicity tolerance in plants exposed to suboptimal soil conditions for sustained food production.Maria Hrmova, Matthew Gilliham and Stephen D. Tyerma