Plasticity and implications of water-use traits in contrasting tropical tree species under climate change.

Plants face a trade-off between hydraulic safety and growth, leading to a range of water-use strategies in different species. However, little is known about such strategies in tropical trees and whether different water-use traits can acclimate to warming. We studied five water-use traits in 20 tropical tree species grown at three different altitudes in Rwanda (RwandaTREE): stomatal conductance (gs), leaf minimum conductance (gmin), plant hydraulic conductance (Kplant), leaf osmotic potential (ψo) and net defoliation during drought. We also explored the links between these traits and growth and mortality data. Late successional (LS) species had low Kplant, gs and gmin and, thus, low water loss, while low ψo helped improve leaf water status during drought. Early successional (ES) species, on the contrary, used more water during both moist and dry conditions and exhibited pronounced drought defoliation. The ES strategy was associated with lower mortality and more pronounced growth enhancement at the warmer sites compared to LS species. While Kplant and gmin showed downward acclimation in warmer climates, ψo did not acclimate and gs measured at prevailing temperature did not change. Due to distinctly different water use strategies between successional groups, ES species may be better equipped for a warmer climate as long as defoliation can bridge drought periods.

Transcription factor ZmDof22 enhances drought tolerance by regulating stomatal movement and antioxidant enzymes activities in maize (Zea mays L.).

KEY MESSAGE: The Dof22 gene encoding a deoxyribonucleic acid binding with one finger in maize, which is associated with its drought tolerance. The identification of drought stress regulatory genes is essential for the genetic improvement of maize yield. Deoxyribonucleic acid binding with one finger (Dof), a plant-specific transcription factor family, is involved in signal transduction, morphogenesis, and environmental stress responses. In present study, by weighted correlation network analysis (WGCNA) and gene co-expression network analysis, 15 putative Dof genes were identified from maize that respond to drought and rewatering. A real-time fluorescence quantitative PCR showed that these 15 genes were strongly induced by drought and ABA treatment, and among them ZmDof22 was highly induced by drought and ABA treatment. Its expression level increased by nearly 200 times after drought stress and more than 50 times after ABA treatment. After the normal conditions were restored, the expression levels were nearly 100 times and 40 times of those before treatment, respectively. The Gal4-LexA/UAS system and transcriptional activation analysis indicate that ZmDof22 is a transcriptional activator regulating drought tolerance and recovery ability in maize. Further, overexpressed transgenic and mutant plants of ZmDof22 by CRISPR/Cas9, indicates that the ZmDof22, improves maize drought tolerance by promoting stomatal closure, reduces water loss, and enhances antioxidant enzyme activity by participating in the ABA pathways. Taken together, our findings laid a foundation for further functional studies of the ZmDof gene family and provided insights into the role of the ZmDof22 regulatory network in controlling drought tolerance and recovery ability of maize.

A network-based modeling framework reveals the core signal transduction network underlying high carbon dioxide-induced stomatal closure in guard cells.

Stomata are pores on plant aerial surfaces, each bordered by a pair of guard cells. They control gas exchange vital for plant survival. Understanding how guard cells respond to environmental signals such as atmospheric carbon dioxide (CO2) levels is not only insightful to fundamental biology but also relevant to real-world issues of crop productivity under global climate change. In the past decade, multiple important signaling elements for stomatal closure induced by elevated CO2 have been identified. Yet, there is no comprehensive understanding of high CO2-induced stomatal closure. In this work, we assemble a cellular signaling network underlying high CO2-induced stomatal closure by integrating evidence from a comprehensive literature analysis. We further construct a Boolean dynamic model of the network, which allows in silico simulation of the stomatal closure response to high CO2 in wild-type Arabidopsis thaliana plants and in cases of pharmacological or genetic manipulation of network nodes. Our model has a 91% accuracy in capturing known experimental observations. We perform network-based logical analysis and reveal a feedback core of the network, which dictates cellular decisions in closure response to high CO2. Based on these analyses, we predict and experimentally confirm that applying nitric oxide (NO) induces stomatal closure in ambient CO2 and causes hypersensitivity to elevated CO2. Moreover, we predict a negative regulatory relationship between NO and the protein phosphatase ABI2 and find experimentally that NO inhibits ABI2 phosphatase activity. The experimental validation of these model predictions demonstrates the effectiveness of network-based modeling and highlights the decision-making role of the feedback core of the network in signal transduction. We further explore the model's potential in predicting targets of signaling elements not yet connected to the CO2 network. Our combination of network science, in silico model simulation, and experimental assays demonstrates an effective interdisciplinary approach to understanding system-level biology.

H2S is involved in drought-mediated stomatal closure through PLDα1 in Arabidopsis.

MAIN CONCLUSION: PLDα1 promoted H2S production by positively regulating the expression of LCD. Stomatal closure promoted by PLDα1 required the accumulation of H2S under drought stress. Phospholipase Dα1 (PLDα1) acting as one of the signal enzymes can respond to drought stress. It is well known that hydrogen sulfide (H2S) plays an important role in plant responding to biotic or abiotic stress. In this study, the functions and relationship between PLDα1 and H2S in drought stress resistance in Arabidopsis were explored. Our results indicated that drought stress promotes PLDα1 and H2S production by inducing the expression of PLDα1 and LCD genes. PLDα1 and LCD enhanced plant tolerance to drought by regulating membrane lipid peroxidation, proline accumulation, H2O2 content and stomatal closure. Under drought stress, the H2O2 content of PLDα1-deficient mutant (pldα1), L-cysteine desulfhydrase (LCD)-deficient mutant (lcd) was higher than that of ecotype (WT), the stomatal aperture of pldα1 and lcd was larger than that of WT. The transcriptional and translational levels of LCD were lower in pldα1 than that in WT. Exogenous application of the H2S donor NaHS or GYY reduced the stomatal aperture of WT, pldα1, PLDα1-CO, and PLDα1-OE lines, while exogenous application of the H2S scavenger hypotaurine (HT) increased the stomatal aperture. qRT-PCR analysis of stomatal movement-related genes showed that the expression of CAX1, ABCG5, SCAB1, and SLAC1 genes in pldα1 and lcd were down-regulated, while ACA1 and OST1 gene expression was significantly up-regulated. Thus, PLDα1 and LCD are required for stomatal closure to improve drought stress tolerance.

Potassium deficiency enhances imbalances in rice water relations under water deficit by decreasing leaf hydraulic conductance.

Potassium (K+) is an essential macronutrient for appropriate plant development and physiology. However, little is known about the mechanisms involved in the regulation of leaf water relations by K under water deficit. A pot experiment with two K supplies of 0.45 and 0 g K2O per pot (3 kg soil per pot) and two watering conditions (well-watered and water-deficit) was conducted to explore the effects of K deficiency on canopy transpiration characteristics, leaf water status, photosynthesis, and hydraulic traits in two rice genotypes with contrasting resistance to drought. The results showed that K deficiency reduced canopy transpiration rate by decreasing stomatal conductance, which led to higher canopy temperatures, resulting in limited water deficit tolerance in rice. In addition, K deficiency led to further substantial reductions in leaf relative water content and water potential under water deficit, which increased the imbalance in leaf water relations under water deficit. Notably, K deficiency limited leaf gas exchange by reducing leaf hydraulic conductance, but decreased the intrinsic water use efficiency under water deficit, especially for the drought-resistant cultivar. Further analysis of the underlying process of leaf hydraulic resistance revealed that the key limiting factor of leaf hydraulic conductance under K deficiency was the outside-xylem hydraulic conductance rather than the xylem hydraulic conductance. Overall, our results provide a comprehensive perspective for assessing leaf water relations under K deficiency, water deficit, and their combined stresses, which will be useful for optimal rice fertilization strategies.

Effects of high-temperature stress on gene expression related to photosynthesis in two jujube (Ziziphus jujuba Mill.) varieties.

Elevated temperatures critically impact crop growth, development, and yield, with photosynthesis being the most temperature-sensitive physiological process in plants. This study focused on assessing the photosynthetic response and genetic adaptation of two different heat-resistant jujube varieties 'Junzao' (J) and 'Fucuimi' (F), to high-temperature stress (42°C Day/30°C Night). Comparative analyses of leaf photosynthetic indices, microstructural changes, and transcriptome sequencing were conducted. Results indicated superior high-temperature adaptability in F, evidenced by alterations in leaf stomatal behavior - particularly in J, where defense cells exhibited significant water loss, shrinkage, and reduced stomatal opening, alongside a marked increase in stomatal density. Through transcriptome sequencing 13,884 differentially expressed genes (DEGs) were identified, significantly enriched in pathways related to plant-pathogen interactions, amino acid biosynthesis, starch and sucrose metabolism, and carbohydrate metabolism. Key findings include the identification of photosynthetic pathway related DEGs and HSFA1s as central regulators of thermal morphogenesis and heat stress response. Revealing their upregulation in F and downregulation in J. The results indicate that these genes play a crucial role in improving heat tolerance in F. This study unveils critical photosynthetic genes involved in heat stress, providing a theoretical foundation for comprehending the molecular mechanisms underlying jujube heat tolerance.

Effects of different light conditions on morphological, anatomical, photosynthetic and biochemical parameters of Cypripedium macranthos Sw.

In this study, the morphological (plant height, leaf length and width, stem diameter and leaf number), anatomical (epidermal cell density and thickness, Stomatal length and width), photosynthetic (net photosynthetic rate, transpiration rate, stomatal conductance, intercellular CO2 concentration, relative humidity, leaf temperature and chlorophyll fluorescence parameters) and biochemical parameters (the content of soluble sugar, soluble protein, proline, malondialdehyde and electrical conductivity) of Cypripedium macranthos Sw. in Changbai Mountain were determined under different light conditions (L10, L30, L50, L100). The results showed that morphological values including plant height, leaf area, stem diameter and leaf number of C. macranthos were smaller under the condition of full light at L100. The epidermal cell density and epidermal thickness of C. macranthos were the highest under L30 and L50 treatments, respectively. It had the highest net photosynthetic rate (Pn) and chlorophyll content under L50 treatment. Meanwhile, correlation analysis indicated that photosynthetically active radiation (PAR) and water use efficiency (WUE) were the main factors influencing Pn. C. macranthos accumulated more soluble sugars and soluble proteins under L100 treatment, while the degree of membrane peroxidation was the highest and the plant was severely damaged. In summary, the adaptability of C. macranthos to light conditions is ranked as follows L50 > L30 > L10 > L100. Appropriate light conditions for C. macranthos are 30%-50% of full light, which should be taken into account in protection and cultivation.

Evaluation of potential increase in photosynthetic efficiency of cassava (Manihot esculenta Crantz) plants exposed to elevated carbon dioxide.

Cassava (Manihot esculenta Crantz), an important tropical crop, is affected by extreme climatic events, including rising CO2 levels. We evaluated the short-term effect of elevated CO2 concentration (ECO2 ) (600, 800 and 1000ppm) on the photosynthetic efficiency of 14 cassava genotypes. ECO2 significantly altered gaseous exchange parameters (net photosynthetic rate (P n ), stomatal conductance (g s ), intercellular CO2 (C i ) and transpiration (E )) in cassava leaves. There were significant but varying interactive effects between ECO2 and varieties on these physiological characteristics. ECO2 at 600 and 800ppm increased the P n rate in the range of 13-24% in comparison to 400ppm (ambient CO2 ), followed by acclimation at the highest concentration of 1000ppm. A similar trend was observed in g s and E . Conversely, C i increased significantly and linearly across increasing CO2 concentration. Along with C i , a steady increase in water use efficiency [WUEintrinsic (P n /g s ) and WUEinstantaneous (P n /E )] across various CO2 concentrations corresponded with the central role of restricted stomatal activity, a common response under ECO2 . Furthermore, P n had a significant quadratic relationship with the ECO2 (R 2 =0.489) and a significant and linear relationship with C i (R 2 =0.227). Relative humidity and vapour pressure deficit during the time of measurements remained at 70-85% and ~0.9-1.31kPa, respectively, at 26±2°C leaf temperature. Notably, not a single variety exhibited constant performance for any of the parameters across CO2 concentrations. Our results indicate that the potential photosynthesis can be increased up to 800ppm cassava varieties with high sink capacity can be cultivated under protected cultivation to attain higher productivity.

Accurate photosynthetic parameter estimation at low stomatal conductance: effects of cuticular conductance and instrumental noise.

Accurate estimation of photosynthetic parameters is essential for understanding plant physiological limitations and responses to environmental factors from the leaf to the global scale. Gas exchange is a useful tool to measure responses of net CO2 assimilation (A) to internal CO2 concentration (Ci), a necessary step in estimating photosynthetic parameters including the maximum rate of carboxylation (Vcmax) and the electron transport rate (Jmax). However, species and environmental conditions of low stomatal conductance (gsw) reduce the signal-to-noise ratio of gas exchange, challenging estimations of Ci. Previous works showed that not considering cuticular conductance to water (gcw) can lead to significant errors in estimating Ci, because it has a different effect on total conductance to CO2 (gtc) than does gsw. Here we present a systematic assessment of the need for incorporating gcw into Ci estimates. In this study we modeled the effect of gcw and of instrumental noise and quantified these effects on photosynthetic parameters in the cases of four species with varying gsw and gcw, measured using steady-state and constant ramping techniques, like the rapid A/Ci response method. We show that not accounting for gcw quantitatively influences Ci and the resulting Vcmax and Jmax, particularly when gcw exceeds 7% of the total conductance to water. The influence of gcw was not limited to low gsw species, highlighting the importance of species-specific knowledge before assessing A/Ci curves. Furthermore, at low gsw instrumental noise can affect Ci estimation, but the effect of instrumental noise can be minimized using constant-ramping rather than steady-state techniques. By incorporating these considerations, more precise measurements and interpretations of photosynthetic parameters can be obtained in a broader range of species and environmental conditions.

PIF transcriptional regulators are required for rhythmic stomatal movements.

Stomata govern the gaseous exchange between the leaf and the external atmosphere, and their function is essential for photosynthesis and the global carbon and oxygen cycles. Rhythmic stomata movements in daily dark/light cycles prevent water loss at night and allow CO2 uptake during the day. How the actors involved are transcriptionally regulated and how this might contribute to rhythmicity is largely unknown. Here, we show that morning stomata opening depends on the previous night period. The transcription factors PHYTOCHROME-INTERACTING FACTORS (PIFs) accumulate at the end of the night and directly induce the guard cell-specific K+ channel KAT1. Remarkably, PIFs and KAT1 are required for blue light-induced stomata opening. Together, our data establish a molecular framework for daily rhythmic stomatal movements under well-watered conditions, whereby PIFs are required for accumulation of KAT1 at night, which upon activation by blue light in the morning leads to the K+ intake driving stomata opening.

Simultaneous and Independent Abaxial and Adaxial Gas Exchange Measurements.

Stomata can be distributed exclusively on the abaxial or adaxial leaf surface, but they are most commonly found on both leaf surfaces. Variations in stomatal arrangement, patterning, and the impact on photosynthesis can be measured using an infrared gas exchange system. However, when using standard gas exchange techniques, both surfaces are measured together and averaged to provide leaf-level values. Employing an innovative gas exchange apparatus with two infrared gas analyzers, separate gaseous flux from both leaf surfaces can be quantified simultaneously and independently. Here, we provide examples of typical measurements that can be performed using a "split chamber" gas exchange system.

Using Infrared Thermography for High-Throughput Plant Phenotyping.

Infrared thermography offers a rapid, noninvasive method for measuring plant temperature, which provides a proxy for stomatal conductance and plant water status and can therefore be used as an index for plant stress. Thermal imaging can provide an efficient method for high-throughput screening of large numbers of plants. This chapter provides guidelines for using thermal imaging equipment and illustrative methodologies, coupled with essential considerations, to access plant physiological processes.

RotatedStomataNet: a deep rotated object detection network for directional stomata phenotype analysis.

KEY MESSAGE: Innovatively, we consider stomatal detection as rotated object detection and provide an end-to-end, batch, rotated, real-time stomatal density and aperture size intelligent detection and identification system, RotatedeStomataNet. Stomata acts as a pathway for air and water vapor in the course of respiration, transpiration, and other gas metabolism, so the stomata phenotype is important for plant growth and development. Intelligent detection of high-throughput stoma is a key issue. Nevertheless, currently available methods usually suffer from detection errors or cumbersome operations when facing densely and unevenly arranged stomata. The proposed RotatedStomataNet innovatively regards stomata detection as rotated object detection, enabling an end-to-end, real-time, and intelligent phenotype analysis of stomata and apertures. The system is constructed based on the Arabidopsis and maize stomatal data sets acquired destructively, and the maize stomatal data set acquired in a non-destructive way, enabling the one-stop automatic collection of phenotypic, such as the location, density, length, and width of stomata and apertures without step-by-step operations. The accuracy of this system to acquire stomata and apertures has been well demonstrated in monocotyledon and dicotyledon, such as Arabidopsis, soybean, wheat, and maize. The experimental results that the prediction results of the method are consistent with those of manual labeling. The test sets, the system code, and their usage are also given ( https://github.com/AITAhenu/RotatedStomataNet ).

Plant responses to co-occurring heat and water deficit stress: A comparative study of tolerance mechanisms in old and modern wheat genotypes.

Global climate change increases the likelihood of co-occurrence of hot and dry spells with increased intensity, frequency, and duration. Studying the impact of the two stresses provide a better understanding of tolerance mechanisms in wheat, and our study was focused on revealing plant stress responses to different severities of combined stress at two phenophases in old and modern wheat genotypes. During the stem elongation and anthesis stages, plants were exposed to four treatments: control, deficit irrigation, combined heat, and deficit irrigation at 31 °C (HD31) and 37 °C (HD37). The modern genotypes were less affected by deficit irrigation at stem elongation as they maintained higher photosynthesis, stomatal conductance, and leaf cooling than old genotypes. When the HD37 stress was imposed during anthesis, the modern genotypes exhibited superior performance compared to the old, which was due to their higher photosynthetic rates resulting from improved biochemical regulation and a higher chlorophyll content. The plant responses varied during two phenophases under the combined stress exposure. Genotypes subjected to HD37 stress during stem elongation, photosynthesis was mainly controlled by stomatal regulation, whereas at anthesis it was predominated by biochemical regulation. These findings contribute to a deeper comprehension of plant tolerance mechanisms in response to different intensities of co-occurring hot and dry weather conditions.

Aluminium tolerance and stomata operation: Towards optimising crop performance in acid soil.

Stomatal operation is crucial for optimising plant water and gas exchange and represents a major trait conferring abiotic stress tolerance in plants. About 56% of agricultural land around the globe is classified as acidic, and Al toxicity is a major limiting factor affecting plant performance in such soils. While most of the research work in the field discusses the impact of major abiotic stresses such as drought or salinity on stomatal operation, the impact of toxic metals and, specifically aluminium (Al) on stomatal operation receives much less attention. We aim to fill this knowledge gap by summarizing the current knowledge of the adverse effects of acid soils on plant stomatal development and operation. We summarised the knowledge of stomatal responses to both long-term and transient Al exposure, explored molecular mechanisms underlying plant adaptations to Al toxicity, and elucidated regulatory networks that alleviate Al toxicity. It is shown that Al-induced stomatal closure involves regulations of core stomatal signalling components, such as ROS, NO, and CO2 and key elements of ABA signalling. We also discuss possible targets and pathway to modify stomatal operation in plants grown in acid soils thus reducing the impact of Al toxicity on plant growth and yield.

Nocturnal stomatal behaviour and its impact on water use strategies of desert herbs in the Gurbantunggut Desert, Northwest China.

Nocturnal stomatal behaviour has the potential to exert a profound influence on plant-water relations, especially water use efficiency. However, we know very less about plant functional type differences in nocturnal stomatal conductance and their roles in plant stress adaptation, especially drought adaptation. To address this critical knowledge gap, we assessed diel leaf gas exchanges in eight ephemeral and perennial herbs growing on the southern edge of the Gurbantunggut Desert, Northwest China. For both ephemeral and perennial herbs, the nocturnal stomatal conductance (gs) exceeded 30 % of daytime gs, except for an ephemeral herb (Malcolmia scorpioides). The nocturnal gs in the studied perennial herbs were significantly higher than it in the ephemeral herbs. The results suggest that circadian-driven stomatal priming plays a pivotal role in accelerating the attainment of steady-state gs during the morning for perennial herbs, thereby enhancing their capacity for carbon capture. Moreover, the nocturnal stomatal behaviour of the ephemeral herbs favored water retention in the morning, consequently enhanced intrinsic water use efficiency and long-term water use efficiency. In summary, plant functional type differences in the magnitude of nocturnal stomatal opening were related to differences in water acquisition and utilization and highlighted diverse water use strategies in the desert plants.

Effects of increasing atmospheric CO2 on leaf water δ18O values are small and are attenuated in grasses and amplified in dicotyledonous herbs and legumes when transferred to cellulose δ18O values.

The oxygen isotope composition of cellulose (δ18O values) has been suggested to contain information on stomatal conductance (gs) responses to rising pCO2. The extent by which pCO2 affects leaf water and cellulose δ18O values (δ18OLW and δ18OC) and the isotope processes that determine pCO2 effects on δ18OLW and δ18OC are, however, unknown. We tested the effects of pCO2 on gs, δ18OLW and δ18OC in a glasshouse experiment, where six plant species were grown under pCO2 ranging from 200 to 500 ppm. Increasing pCO2 caused a decline in gs and an increase in δ18OLW, as expected. Importantly, the effects of pCO2 on gs and δ18OLW were small and pCO2 effects on δ18OLW were not directly transferred to δ18OC but were attenuated in grasses and amplified in dicotyledonous herbs and legumes. This is likely because of functional group-specific pCO2 effects on the model parameter pxpex. Our study highlights important uncertainties when using δ18OC as a proxy for gs. Specifically, pCO2-triggered gs effects on δ18OLW and δ18OC are possibly too small to be detected in natural settings and a pCO2 effect on pxpex may render the commonly assumed negative linkage between δ18OC and gs to be incorrect, potentially confounding δ18OC based gs reconstructions.

Coordinated hydraulic traits influence the two phases of time to hydraulic failure in five temperate tree species differing in stomatal stringency.

Worldwide, forests are increasingly exposed to extreme droughts causing tree mortality. Because of the complex nature of the mechanisms involved, various traits have been linked to tree drought responses with contrasting results. This may be due to species-specific strategies in regulating water potential, a process that unfolds in two distinct phases: a first phase until stomatal closure, and a second phase until reaching lethal xylem hydraulic thresholds. We conducted dry-down experiments with five broadleaved temperate tree species differing in their degree of isohydry to estimate the time to stomatal closure (tsc) and subsequent time to critical hydraulic failure (tcrit). We measured various traits linked to tree drought responses, such as the water potentials at turgor loss point (Ptlp), stomatal closure (Pgs90), and 12%, 50% and 88% loss of xylem hydraulic conductance (P12, P50, P88), hydraulic capacitance (C), minimum leaf conductance (gmin), hydroscape area (HSA) and hydraulic safety margins (HSM). We found that Pgs90 followed previously recorded patterns of isohydry and was associated with HSA. Species ranked from more to less isohydric in the sequence Acer pseudoplatanus < Betula pendula < Tilia cordata < Sorbus aucuparia < Fagus sylvatica. Their degree of isohydry was associated with leaf safety (Ptlp and gmin), drought avoidance (C) and tsc, but decoupled from xylem safety (HSM and P88) and tcrit. Regardless of their stomatal stringency, species with wider HSM and lower P88 reached critical hydraulic failure later. We conclude that the duration of the first phase is determined by stomatal regulation, while the duration of the second phase is associated with xylem safety. Isohydry is thus linked to water use rather than to drought survival strategies, confirming the proposed use of HSA as a complement to HSM for describing plant drought responses before and after stomatal closure.

Leaf morpho-anatomical adjustments in a Quercus pubescens forest after 10 years of partial rain exclusion in the field.

In the Mediterranean region, a reduction of annual precipitation and a longer and drier summer season are expected with climate change by the end of the century, eventually endangering forest survival. To cope with such rapid changes, trees may modulate their morpho-anatomical and physiological traits. In the present study, we focused on the variation in leaf gas exchange and different leaf morpho-anatomical functional traits of Quercus pubescens Willd. in summer using a long-term drought experiment in natura consisting of a dynamic rainfall exclusion system where trees have been submitted to amplified drought (AD) (~-30% of annual precipitation) since April 2012 and compared them with trees under natural drought (ND) in a Mediterranean forest. During the study, we analyzed net CO2 assimilation (An), stomatal conductance (gs), transpiration (E), water-use efficiency (WUE), stomatal size and density, density of glandular trichomes and non-glandular trichomes, thickness of the different leaf tissues, specific leaf area and leaf surface. Under AD, tree functioning was slightly impacted, since only An exhibited a 49% drop, while gs, E and WUE remained stable. The decrease in An under AD was regulated by concomitant lower stomatal density and reduced leaf thickness. Trees under AD also featured leaves with a higher non-glandular trichome density and a lower glandular trichome density compared with ND, which simultaneously limits transpiration and production costs. This study points out that Q. pubescens exhibits adjustments of leaf morpho-anatomical traits which can help trees to acclimate to AD scenarios as those expected in the future in the Mediterranean region.

Guard cell and subsidiary cell sizes are key determinants for stomatal kinetics and drought adaptation in cereal crops.

Climate change-induced drought is a major threat to agriculture. C4 crops have a higher water use efficiency (WUE) and better adaptability to drought than C3 crops due to their smaller stomatal morphology and faster response. However, our understanding of stomatal behaviours in both C3 and C4 Poaceae crops is limited by knowledge gaps in physical traits of guard cell (GC) and subsidiary cell (SC). We employed infrared gas exchange analysis and a stomatal assay to explore the relationship between GC/SC sizes and stomatal kinetics across diverse drought conditions in two C3 (wheat and barley) and three C4 (maize, sorghum and foxtail millet) upland Poaceae crops. Through statistical analyses, we proposed a GCSC-τ model to demonstrate how morphological differences affect stomatal kinetics in C4 Poaceae crops. Our findings reveal that morphological variations specifically correlate with stomatal kinetics in C4 Poaceae crops, but not in C3 ones. Subsequent modelling and experimental validation provide further evidence that GC/SC sizes significantly impact stomatal kinetics, which affects stomatal responses to different drought conditions and thereby WUE in C4 Poaceae crops. These findings emphasize the crucial advantage of GC/SC morphological characteristics and stomatal kinetics for the drought adaptability of C4 Poaceae crops, highlighting their potential as future climate-resilient crops.