Search

Gas exchange measurements enable mechanistic insights into the processes that underpin carbon and water fluxes in plant leaves which in turn inform understanding of related processes at a range of scales from individual cells to entire ecosytems. Given the importance of photosynthesis for the global climate discussion it is important to (a) foster a basic understanding of the fundamental principles underpinning the experimental methods used by the broad community, and (b) ensure best practice and correct data interpretation within the research community. In this review, we outline the biochemical and biophysical parameters of photosynthesis that can be investigated with gas exchange measurements and we provide step-by-step guidance on how to reliably measure them. We advise on best practices for using gas exchange equipment and highlight potential pitfalls in experimental design and data interpretation. The Supporting Information contains exemplary data sets, experimental protocols and data-modelling routines. This review is a community effort to equip both the experimental researcher and the data modeller with a solid understanding of the theoretical basis of gas-exchange measurements, the rationale behind different experimental protocols and the approaches to data interpretation., (© 2024 The Authors. Plant, Cell & Environment published by John Wiley & Sons Ltd.)

Stomata of plant leaves open to enable CO 2 entry for photosynthesis and close to reduce water loss via transpiration. Compared with photosynthesis, stomata respond slowly to fluctuating light, reducing assimilation and water use efficiency. Efficiency gains are possible without a cost to photosynthesis if stomatal kinetics can be accelerated. Here we show that clustering of the GORK channel, which mediates K + efflux for stomatal closure in the model plant Arabidopsis, arises from binding between the channel voltage sensors, creating an extended 'sensory antenna' for channel gating. Mutants altered in clustering affect channel gating to facilitate K + flux, accelerate stomatal movements and reduce water use without a loss in biomass. Our findings identify the mechanism coupling channel clustering with gating, and they demonstrate the potential for engineering of ion channels native to the guard cell to enhance stomatal kinetics and improve water use efficiency without a cost in carbon fixation., (© 2022. The Author(s), under exclusive licence to Springer Nature Limited.)

Stomata serve dual and often conflicting roles, facilitating carbon dioxide influx into the plant leaf for photosynthesis and restricting water efflux via transpiration. Strategies for reducing transpiration without incurring a cost for photosynthesis must circumvent this inherent coupling of carbon dioxide and water vapor diffusion. We expressed the synthetic, light-gated K + channel BLINK1 in guard cells surrounding stomatal pores in Arabidopsis to enhance the solute fluxes that drive stomatal aperture. BLINK1 introduced a K + conductance and accelerated both stomatal opening under light exposure and closing after irradiation. Integrated over the growth period, BLINK1 drove a 2.2-fold increase in biomass in fluctuating light without cost in water use by the plant. Thus, we demonstrate the potential of enhancing stomatal kinetics to improve water use efficiency without penalty in carbon fixation., (Copyright © 2019 The Authors, some rights reserved; exclusive licensee American Association for the Advancement of Science. No claim to original U.S. Government Works.)

Stomatal movements depend on the transport and metabolism of osmotic solutes that drive reversible changes in guard cell volume and turgor. These processes are defined by a deep knowledge of the identities of the key transporters and of their biophysical and regulatory properties, and have been modeled successfully with quantitative kinetic detail at the cellular level. Transpiration of the leaf and canopy, by contrast, is described by quasilinear, empirical relations for the inputs of atmospheric humidity, CO 2 , and light, but without connection to guard cell mechanics. Until now, no framework has been available to bridge this gap and provide an understanding of their connections. Here, we introduce OnGuard2, a quantitative systems platform that utilizes the molecular mechanics of ion transport, metabolism, and signaling of the guard cell to define the water relations and transpiration of the leaf. We show that OnGuard2 faithfully reproduces the kinetics of stomatal conductance in Arabidopsis thaliana and its dependence on vapor pressure difference (VPD) and on water feed to the leaf. OnGuard2 also predicted with VPD unexpected alterations in K + channel activities and changes in stomatal conductance of the slac1 Cl - channel and ost2 H + -ATPase mutants, which we verified experimentally. OnGuard2 thus bridges the micro-macro divide, offering a powerful tool with which to explore the links between guard cell homeostasis, stomatal dynamics, and foliar transpiration., (© 2017 American Society of Plant Biologists. All rights reserved.)

Stomata are microscopic pores formed by specialized cells in the leaf epidermis and permit gaseous exchange between the interior of the leaf and the atmosphere. Stomata in most plants are separated by at least one epidermal pavement cell and, individually, overlay a single substomatal cavity within the leaf. This spacing is thought to enhance stomatal function. Yet, there are several genera naturally exhibiting stomata in clusters and therefore deviating from the one-cell spacing rule with multiple stomata overlaying a single substomatal cavity. We made use of two Begonia species to investigate whether clustering of stomata alters guard cell dynamics and gas exchange under different light and dark treatments. Begonia plebeja, which forms stomatal clusters, exhibited enhanced kinetics of stomatal conductance and CO2 assimilation upon light stimuli that in turn were translated into greater water use efficiency. Our findings emphasize the importance of spacing in stomatal clusters for gaseous exchange and plant performance under environmentally limited conditions., (© The Author 2017. Published by Oxford University Press on behalf of the Society for Experimental Biology.)

Stomata enable gaseous exchange between the interior of the leaf and the atmosphere through the stomatal pore. Control of the pore aperture depends on osmotic solute accumulation by, and its loss from the guard cells surrounding the pore. Stomata in most plants are separated by at least one epidermal cell, and this spacing is thought to enhance stomatal function, although there are several genera that exhibit stomata in clusters. We made use of Arabidopsis (Arabidopsis thaliana) stomatal patterning mutants to explore the impact of clustering on guard cell dynamics, gas exchange, and ion transport of guard cells. These studies showed that stomatal clustering in the Arabidopsis too many mouths (tmm1) mutant suppressed stomatal movements and affected CO2 assimilation and transpiration differentially between dark and light conditions and were associated with alterations in K(+) channel gating. These changes were consistent with the impaired dynamics of tmm1 stomata and were accompanied by a reduced accumulation of K(+) ions in the guard cells. Our findings underline the significance of spacing for stomatal dynamics. While stomatal spacing may be important as a reservoir for K(+) and other ions to facilitate stomatal movements, the effects on channel gating, and by inference on K(+) accumulation, cannot be explained on the basis of a reduced number of epidermal cells facilitating ion supply to the guard cells., (© 2016 American Society of Plant Biologists. All rights reserved.)

Phototropins (phots) regulate a range of adaptive processes in plants that serve to optimize photosynthetic efficiency and promote growth. Light sensing by Arabidopsis thaliana phots is predominantly mediated by the Light, Oxygen and Voltage sensing 2 (LOV2) flavin-binding motif located within the N-terminus of the photoreceptor. Here we characterize the photochemical and biochemical properties of phot from the marine picoalga Ostreococcus tauri phototropin (Otphot) and examine its ability to replace phot-mediated function in Arabidopsis. Photochemical properties of Otphot rely on both LOV1 and LOV2. Yet, biochemical analysis indicates that light-dependent receptor autophosphorylation is primarily dependent on LOV2. As found for Arabidopsis phots, Otphot associates with the plasma membrane and partially internalizes, albeit to a limited extent, in response to blue-light irradiation. Otphot is able to elicit a number of phot-regulated processes in Arabidopsis, including petiole positioning, leaf expansion, stomatal opening and chloroplast accumulation movement. However, Otphot is unable to restore phototropism and chloroplast avoidance movement. Consistent with its lack of phototropic function in Arabidopsis, Otphot does not associate with or trigger dephosphorylation of the phototropic signalling component Non-Phototropic Hypocotyl 3 (NPH3). Taken together, these findings indicate that the mechanism of action of plant and evolutionarily distant algal phots is less well conserved than previously thought., (© 2015 The Authors. New Phytologist © 2015 New Phytologist Trust.)

Hydrogen sulfide (H2S) is the third biological gasotransmitter, and in animals, it affects many physiological processes by modulating ion channels. H2S has been reported to protect plants from oxidative stress in diverse physiological responses. H2S closes stomata, but the underlying mechanism remains elusive. Here, we report the selective inactivation of current carried by inward-rectifying K(+) channels of tobacco (Nicotiana tabacum) guard cells and show its close parallel with stomatal closure evoked by submicromolar concentrations of H2S. Experiments to scavenge H2S suggested an effect that is separable from that of abscisic acid, which is associated with water stress. Thus, H2S seems to define a unique and unresolved signaling pathway that selectively targets inward-rectifying K(+) channels., (© 2015 American Society of Plant Biologists. All Rights Reserved.)

GORK is the only outward-rectifying Kv-like K(+) channel expressed in guard cells. Its activity is tightly regulated to facilitate K(+) efflux for stomatal closure and is elevated in ABA in parallel with suppression of the activity of the inward-rectifying K(+) channel KAT1. Whereas the population of KAT1 is subject to regulated traffic to and from the plasma membrane, nothing is known about GORK, its distribution and traffic in vivo. We have used transformations with fluorescently-tagged GORK to explore its characteristics in tobacco epidermis and Arabidopsis guard cells. These studies showed that GORK assembles in puncta that reversibly dissociated as a function of the external K(+) concentration. Puncta dissociation parallelled the gating dependence of GORK, the speed of response consistent with the rapidity of channel gating response to changes in the external ionic conditions. Dissociation was also suppressed by the K(+) channel blocker Ba(2+) . By contrast, confocal and protein biochemical analysis failed to uncover substantial exo- and endocytotic traffic of the channel. Gating of GORK is displaced to more positive voltages with external K(+) , a characteristic that ensures the channel facilitates only K(+) efflux regardless of the external cation concentration. GORK conductance is also enhanced by external K(+) above 1 mm. We suggest that GORK clustering in puncta is related to its gating and conductance, and reflects associated conformational changes and (de)stabilisation of the channel protein, possibly as a platform for transmission and coordination of channel gating in response to external K(+) ., (© 2014 The Authors The Plant Journal © 2014 John Wiley & Sons Ltd.)

Much of the 70% of global water usage associated with agriculture passes through stomatal pores of plant leaves. The guard cells, which regulate these pores, thus have a profound influence on photosynthetic carbon assimilation and water use efficiency of plants. We recently demonstrated how quantitative mathematical modeling of guard cells with the OnGuard modeling software yields detail sufficient to guide phenotypic and mutational analysis. This advance represents an all-important step toward applications in directing "reverse-engineering" of guard cell function for improved water use efficiency and carbon assimilation. OnGuard is nonetheless challenging for those unfamiliar with a modeler's way of thinking. In practice, each model construct represents a hypothesis under test, to be discarded, validated or refined by comparisons between model predictions and experimental results. The few guidelines set out here summarize the standard and logical starting points for users of the OnGuard software.

Stomata account for much of the 70% of global water usage associated with agriculture and have a profound impact on the water and carbon cycles of the world. Stomata have long been modeled mathematically, but until now, no systems analysis of a plant cell has yielded detail sufficient to guide phenotypic and mutational analysis. Here, we demonstrate the predictive power of a systems dynamic model in Arabidopsis (Arabidopsis thaliana) to explain the paradoxical suppression of channels that facilitate K(+) uptake, slowing stomatal opening, by mutation of the SLAC1 anion channel, which mediates solute loss for closure. The model showed how anion accumulation in the mutant suppressed the H(+) load on the cytosol and promoted Ca(2+) influx to elevate cytosolic pH (pH(i)) and free cytosolic Ca(2+) concentration ([Ca(2+)](i)), in turn regulating the K(+) channels. We have confirmed these predictions, measuring pH(i) and [Ca(2+)](i) in vivo, and report that experimental manipulation of pH(i) and [Ca(2+)](i) is sufficient to recover K(+) channel activities and accelerate stomatal opening in the slac1 mutant. Thus, we uncover a previously unrecognized signaling network that ameliorates the effects of the slac1 mutant on transpiration by regulating the K(+) channels. Additionally, these findings underscore the importance of H(+)-coupled anion transport for pH(i) homeostasis.