Myriam Dauzat

Arabidopsis Plants Acclimate to Water Deficit at Low Cost through Changes of Carbon Usage: An Integrated Perspective Using Growth, Metabolite, Enzyme, and Gene Expression Analysis

Growth and carbon (C) fluxes are severely altered in plants exposed to soil water deficit. Correspondingly, it has been suggested that plants under water deficit suffer from C shortage. In this study, we test this hypothesis in Arabidopsis (Arabidopsis thaliana) by providing an overview of the responses of growth, C balance, metabolites, enzymes of the central metabolism, and a set of sugar-responsive genes to a sustained soil water deficit. The results show that under drought, rosette relative expansion rate is decreased more than photosynthesis, leading to a more positive C balance, while root growth is promoted. Several soluble metabolites accumulate in response to soil water deficit, with K+ and organic acids as the main contributors to osmotic adjustment. Osmotic adjustment costs only a small percentage of the daily photosynthetic C fixation. All C metabolites measured (not only starch and sugars but also organic acids and amino acids) show a diurnal turnover that often increased under water deficit, suggesting that these metabolites are readily available for being metabolized in situ or exported to roots. On the basis of 30 enzyme activities, no in-depth reprogramming of C metabolism was observed. Water deficit induces a shift of the expression level of a set of sugar-responsive genes that is indicative of increased, rather than decreased, C availability. These results converge to show that the differential impact of soil water deficit on photosynthesis and rosette expansion results in an increased availability of C for the roots, an increased turnover of C metabolites, and a low-cost C-based osmotic adjustment, and these responses are performed without major reformatting of the primary metabolism machinery.

Control of leaf expansion: a developmental switch from metabolics to hydraulics

Leaf expansion is the central process by which plants colonize space, allowing energy capture and carbon acquisition. Water and carbon emerge as main limiting factors of leaf expansion, but the literature remains controversial about their respective contributions. Here, we tested the hypothesis that the importance of hydraulics and metabolics is organized according to both dark/light fluctuations and leaf ontogeny. For this purpose, we established the developmental pattern of individual leaf expansion during days and nights in the model plant Arabidopsis (Arabidopsis thaliana). Under control conditions, decreases in leaf expansion were observed at night immediately after emergence, when starch reserves were lowest. These nocturnal decreases were strongly exaggerated in a set of starch mutants, consistent with an early carbon limitation. However, low-light treatment of wild-type plants had no influence on these early decreases, implying that expansion can be uncoupled from changes in carbon availability. From 4 d after leaf emergence onward, decreases of leaf expansion were observed in the daytime. Using mutants impaired in stomatal control of transpiration as well as plants grown under soil water deficit or high air humidity, we gathered evidence that these diurnal decreases were the signature of a hydraulic limitation that gradually set up as the leaf developed. Changes in leaf turgor were consistent with this pattern. It is concluded that during the course of leaf ontogeny, the predominant control of leaf expansion switches from metabolics to hydraulics. We suggest that the leaf is better armed to buffer variations in the former than in the latter.

Combined Genetic and Modeling Approaches Reveal That Epidermal Cell Area and Number in Leaves Are Controlled by Leaf and Plant Developmental Processes in Arabidopsis

Both leaf production and leaf expansion are tightly linked to cell expansion and cell division, but the functional relationships between all these variables are not clearly established. To get insight into these relationships, a quantitative genetic analysis was performed in 118 recombinant inbred lines derived from a cross between the Landsberg erecta and Antwerp accessions and was combined with a structural equation modeling approach. Main effects and epistatic interactions at the quantitative trait locus (QTL) level were detected for rosette area, rosette leaf number, leaf 6 area, epidermal cell area and number. A QTL at ERECTA marker (ER) controlled cell expansion and cell division, in interaction with two other QTLs at SNP295 and SNP21 markers. Moreover, both the screening for marker association involved in the variation of the relationships between leaf growth variables and the test of alternative functional models by structural equation modeling revealed that the allelic value at ER controlled epidermal cell area and epidermal cell number in a leaf. These effects are driven both by a whole plant mechanism associated with leaf production and by a single leaf mechanism associated with leaf expansion. The complex effects of the QTL at ER were validated in selected heterogeneous inbred families. The ERECTA gene, which is mutated in the Landsberg erecta parental line, was found to be a putative candidate responsible for these mapped effects by phenotyping mutants of this gene at the cellular level. Together, these results give insight into the complex determination of leaf epidermal cell number and area.

PHENOPSIS DB: an information system for Arabidopsis thaliana phenotypic data in an environmental context.

BackgroundRenewed interest in plant × environment interactions has risen in the post-genomic era. In this context, high-throughput phenotyping platforms have been developed to create reproducible environmental scenarios in which the phenotypic responses of multiple genotypes can be analysed in a reproducible way. These platforms benefit hugely from the development of suitable databases for storage, sharing and analysis of the large amount of data collected. In the model plant Arabidopsis thaliana, most databases available to the scientific community contain data related to genetic and molecular biology and are characterised by an inadequacy in the description of plant developmental stages and experimental metadata such as environmental conditions. Our goal was to develop a comprehensive information system for sharing of the data collected in PHENOPSIS, an automated platform for Arabidopsis thaliana phenotyping, with the scientific community.DescriptionPHENOPSIS DB is a publicly available (URL: http://bioweb.supagro.inra.fr/phenopsis/) information system developed for storage, browsing and sharing of online data generated by the PHENOPSIS platform and offline data collected by experimenters and experimental metadata. It provides modules coupled to a Web interface for (i) the visualisation of environmental data of an experiment, (ii) the visualisation and statistical analysis of phenotypic data, and (iii) the analysis of Arabidopsis thaliana plant images.ConclusionsFirstly, data stored in the PHENOPSIS DB are of interest to the Arabidopsis thaliana community, particularly in allowing phenotypic meta-analyses directly linked to environmental conditions on which publications are still scarce. Secondly, data or image analysis modules can be downloaded from the Web interface for direct usage or as the basis for modifications according to new requirements. Finally, the structure of PHENOPSIS DB provides a useful template for the development of other similar databases related to genotype × environment interactions.

Keep on growing under drought: genetic and developmental bases of the response of rosette area using a recombinant inbred line population

Variation in leaf development caused by water deficit was analysed in 120 recombinant inbred lines derived from two Arabidopsis thaliana accessions, Ler and An-1. Main effect quantitative trait loci (QTLs) and QTLs in epistatic interactions were mapped for the responses of rosette area, leaf number and leaf 6 area to water deficit. An epistatic interaction between two QTLs affected the response of whole rosette area and individual leaf area but only with effects in well-watered condition. A second epistatic interaction between two QTLs controlled the response of rosette area and leaf number with specific effects in the water deficit condition. These effects were validated by generating and phenotyping new appropriate lines. Accordingly, a low reduction of rosette area was observed for lines with a specific allelic combination at the two interacting QTLs. This low reduction was accompanied by an increase in leaf number with a lengthening of the vegetative phase and a low reduction in individual leaf area with low reductions in epidermal cell area and number. Statistical analyses suggested that responses of epidermal cell area and number to water deficit in individual leaves were partly caused by delay in flowering time and reduction in leaf emergence rate, respectively.

New Insights into the Control of Endoreduplication: Endoreduplication Could be Driven by Organ Growth in Arabidopsis Leaves

Enormous progress has been achieved understanding the molecular mechanisms regulating endoreduplication. By contrast, how this process is coordinated with the cell cycle or cell expansion and contributes to overall growth in multicellular systems remains unclear. A holistic approach was used here to give insight into the functional links between endoreduplication, cell division, cell expansion, and whole growth in the Arabidopsis (Arabidopsis thaliana) leaf. Correlative analyses, quantitative genetics, and structural equation modeling were applied to a large data set issued from the multiscale phenotyping of 200 genotypes, including both genetically modified lines and recombinant inbred lines. All results support the conclusion that endoreduplication in leaf cells could be controlled by leaf growth itself. More generally, leaf growth could act as a “hub” that drives cell division, cell expansion, and endoreduplication in parallel. In many cases, this strategy allows compensations that stabilize leaf area even when one of the underlying cellular processes is limiting.

Structural assessment of the impact of environmental constraints on Arabidopsis thaliana leaf growth: a 3D approach

Light and soil water content affect leaf surface area expansion through modifications in epidermal cell numbers and area, while effects on leaf thickness and mesophyll cell volumes are far less documented. Here, three-dimensional imaging was applied in a study of Arabidopsis thaliana leaf growth to determine leaf thickness and the cellular organization of mesophyll tissues under moderate soil water deficit and two cumulative light conditions. In contrast to surface area, thickness was highly conserved in response to water deficit under both low and high cumulative light regimes. Unlike epidermal and palisade mesophyll tissues, no reductions in cell number were observed in the spongy mesophyll; cells had rather changed in volume and shape. Furthermore, leaf features of a selection of genotypes affected in leaf functioning were analysed. The low-starch mutant pgm had very thick leaves because of unusually large palisade mesophyll cells, together with high levels of photosynthesis and stomatal conductance. By means of an open stomata mutant and a 9-cis-epoxycarotenoid dioxygenase overexpressor, it was shown that stomatal conductance does not necessarily have a major impact on leaf dimensions and cellular organization, pointing to additional mechanisms for the control of CO(2) diffusion under high and low stomatal conductance, respectively.

Quantifying spatial heterogeneity of chlorophyll fluorescence during plant growth and in response to water stress

BackgroundEffects of abiotic and biotic stresses on plant photosynthetic performance lead to fitness and yield decrease. The maximum quantum efficiency of photosystem II (Fv/Fm) is a parameter of chlorophyll fluorescence (ChlF) classically used to track changes in photosynthetic performance. Despite recent technical and methodological advances in ChlF imaging, the spatio-temporal heterogeneity of Fv/Fm still awaits for standardized and accurate quantification.ResultsWe developed a method to quantify the dynamics of spatial heterogeneity of photosynthetic efficiency through the distribution-based analysis of Fv/Fm values. The method was applied to Arabidopsis thaliana grown under well-watered and severe water deficit (survival rate of 40%). First, whole-plant Fv/Fm shifted from unimodal to bimodal distributions during plant development despite a constant mean Fv/Fm under well-watered conditions. The establishment of a bimodal distribution of Fv/Fm reflects the occurrence of two types of leaf regions with contrasted photosynthetic efficiency. The distance between the two modes (called S) quantified the whole-plant photosynthetic heterogeneity. The weighted contribution of the most efficient/healthiest leaf regions to whole-plant performance (called Wmax) quantified the spatial efficiency of a photosynthetically heterogeneous plant. Plant survival to water deficit was associated to high S values, as well as with strong and fast recovery of Wmax following soil rewatering. Hence, during stress surviving plants had higher, but more efficient photosynthetic heterogeneity compared to perishing plants. Importantly, S allowed the discrimination between surviving and perishing plants four days earlier than the mean Fv/Fm. A sensitivity analysis from simulated dynamics of Fv/Fm showed that parameters indicative of plant tolerance and/or stress intensity caused identifiable changes in S and Wmax. Finally, an independent comparison of six Arabidopsis accessions grown under well-watered conditions indicated that S and Wmax are related to the genetic variability of growth.ConclusionsThe distribution-based analysis of ChlF provides an efficient tool for quantifying photosynthetic heterogeneity and performance. S and Wmax are good indicators to estimate plant survival under water stress. Our results suggest that the dynamics of photosynthetic heterogeneity are key components of plant growth and tolerance to stress.

Buffering growth variations against water deficits through timely carbon usage.

Water stresses reduce plant growth but there is no consensus on whether carbon metabolism has any role in this reduction. Sugar starvation resulting from stomatal closure is often proposed as a cause of growth impairment under long-term or severe water deficits. However, growth decreases faster than photosynthesis in response to drought, leading to increased carbohydrate stores under short-term or moderate water deficits. Here, we addressed the question of the role of carbon availability on growth under moderate water deficits using two different systems. Firstly, we monitored the day/night pattern of leaf growth in Arabidopsis plants. We show that a moderate soil water deficit promotes leaf growth at night in mutants severely disrupted in their nighttime carbohydrate availability. This suggests that soil water deficit promotes carbon satiation. Secondly, we monitored the sub-hourly growth variations of clementine fruits in response to daily, natural fluctuations in air water deficit, and at contrasting source–sink balances obtained by defoliation. We show that high carbohydrate levels prevent excessive, hydraulic shrinkage of the fruit during days with high evaporative demand, most probably through osmotic adjustment. Together, our results contribute to the view that growing organs under moderate soil or air water deficit are not carbon starved, but use soluble carbohydrate in excess to partly release a hydromechanical limitation of growth.

Multivariate genetic analysis of plant responses to water deficit and high temperature revealed contrasting adaptive strategies

Summary Plants need to control transpiration and photosynthesis tightly in order to grow under drought and high temperature. Contrasting genetic-by-environment interactions are exploited by Arabidopsis thaliana to improve stress tolerance.