Integration of metabolite with transcript and enzyme activity profiling during diurnal cycles in Arabidopsis rosettes

ABSTRACT: BACKGROUND: Genome-wide transcript profiling and analyses of enzyme activities from central carbon and nitrogen metabolism has shown that transcript levels undergo marked and rapid changes during diurnal cycles and after transfer to darkness, whereas changes of enzyme activities are smaller and delayed. In the starchless pgm mutant, where sugars are depleted every night, there are accentuated diurnal changes of transcript levels. Enzyme activities do not show larger diurnal changes; instead they shift towards the levels found in wild-type after several days of darkness. These results indicate that enzyme activities change slowly, integrating the changes of transcript levels over several diurnal cycles. RESULTS: To generalize this conclusion, 137 metabolites were profiled using GC-MS and LC-MS. Amplitudes of the diurnal changes of metabolites in pgm were (with the exception of sugars) similar or smaller than in wild-type. The average levels shifted towards those found after several days of darkness in wild-type. Examples include increased levels of many amino acids due to protein degradation, decreased levels of many fatty acids, increased tocopherol and decreased myo-inositol. Many metabolite-transcript correlations were found and the proportion of transcripts correlated with sugars increased dramatically in the starchless pgm mutant. CONCLUSION: Rapid diurnal changes of transcripts are integrated over time to generate quasi-stable changes across large sectors of metabolism. The slow response of enzyme activities and metabolites implies that correlations between metabolites and transcripts are due to regulation of gene expression by metabolites, rather than metabolites being changed as a consequence of a change in gene expression

Global transcript levels respond to small changes of the carbon status during progressive exhaustion of carbohydrates in Arabidopsis rosettes

The balance between the supply and utilization of carbon (C) changes continually. It has been proposed that plants respond in an acclimatory manner, modifying C utilization to minimize harmful periods of C depletion. This hypothesis predicts that signaling events are initiated by small changes in C status. We analyzed the global transcriptional response to a gradual depletion of C during the night and an extension of the night, where C becomes severely limiting from 4 h onward. The response was interpreted using published datasets for sugar, light, and circadian responses. Hundreds of C-responsive genes respond during the night and others very early in the extended night. Pathway analysis reveals that biosynthesis and cellular growth genes are repressed during the night and genes involved in catabolism are induced during the first hours of the extended night. The C response is amplified by an antagonistic interaction with the clock. Light signaling is attenuated during the 24-h light/dark cycle. A model was developed that uses the response of 22K genes during a circadian cycle and their responses to C and light to predict global transcriptional responses during diurnal cycles of wild-type and starchless pgm mutant plants and an extended night in wild-type plants. By identifying sets of genes that respond at different speeds and times during C depletion, our extended dataset and model aid the analysis of candidates for C signaling. This is illustrated for AKIN10 and four bZIP transcription factors, and sets of genes involved in trehalose signaling, protein turnover, and starch breakdown

Overexpression of Plastid Transketolase in Tobacco Results in a Thiamine Auxotrophic Phenotype

To investigate the effect of increased plastid transketolase on photosynthetic capacity and growth, tobacco (Nicotiana tabacum) plants with increased levels of transketolase protein were produced. This was achieved using a cassette composed of a full-length Arabidopsis thaliana transketolase cDNA under the control of the cauliflower mosaic virus 35S promoter. The results revealed a major and unexpected effect of plastid transketolase overexpression as the transgenic tobacco plants exhibited a slow-growth phenotype and chlorotic phenotype. These phenotypes were complemented by germinating the seeds of transketolase-overexpressing lines in media containing either thiamine pyrophosphate or thiamine. Thiamine levels in the seeds and cotyledons were lower in transketolase-overexpressing lines than in wild-type plants. When transketolase-overexpressing plants were supplemented with thiamine or thiamine pyrophosphate throughout the life cycle, they grew normally and the seed produced from these plants generated plants that did not have a growth or chlorotic phenotype. Our results reveal the crucial importance of the level of transketolase activity to provide the precursor for synthesis of intermediates and to enable plants to produce thiamine and thiamine pyrophosphate for growth and development. The mechanism determining transketolase protein levels remains to be elucidated, but the data presented provide evidence that this may contribute to the complex regulatory mechanisms maintaining thiamine homeostasis in plants

L'usage de l'espace par les exploitations d'élevage de montagne et la gestion de la biodiversité

International audienceEn prenant appui sur divers résultats de recherches menées depuis 20 ans dans les Pyrénées centrales, les caractéristiques de l'utilisation de l'espace par les exploitations agricoles sont présentées. Les particularités spatiales des territoires d'exploitation influent sur les pratiques d'utilisation des prairies et sur la dynamique des couverts et des paysages. Un modèle de référence permet de diagnostiquer la maîtrise de l'exploitation de la végétation. L'impact des modes d'usage des prairies sur leur richesse en espèces est présenté sur quelques cas concrets. Une gestion mal maîtrisée conduit à une augmentation de la diversité intraparcellaire et à une diminution de la diversité interparcellaire, puis à une baisse rapide de ces 2 diversités. Raisonner l'organisation de l'usage des prairies à des niveaux d'organisation larges est une nécessité pour préserver la biodiversité en région de montagne

The potential of integrative phenomics to harness underutilized crops for improving stress resilience

The current agricultural and food system faces diverse and increasing challenges. These include feeding an ever-growing human population, expected to reach about 10 billion by 2050 combined with societal disruption, and the need to cope with the impact of climate change (FAO, 2022). Given that future environmental conditions will limit crop productivity (Zhao et al., 2017; Cooper et al., 2021) and the limited potential to continually increase the performance of staple crops by conventional breeding (Hickey et al., 2019), there is an urgent need to transform agricultural systems. Central to this transformation is the application of alternative, accelerated, and sustainable approaches for the improvement and development of underutilized crops (Hickey et al., 2019). Modern breeding strategies for major crops have widely integrated novel technologies, such as advanced phenotyping or genome-wide interactions, and even epigenomics within “beyond the gene” strategies (Crisp et al., 2022) to speed up crop/genotype selection (Hickey et al., 2019; Kumar et al., 2023). Deploying phenotyping at different scales has the potential to identify novel trait(s) components that can be targeted to accelerate crop improvement (Araus and Cairns, 2014; Großkinsky et al., 2015b; Zhao et al., 2019; Varshney et al., 2021). There is even greater potential for these technologies when used to improve underutilized crops and support the agricultural transformation, as underutilized crops typically lack a biased breeding/selection history, i.e., they often exhibit a high genetic diversity and potential, and are usually better adapted to challenging environments (Kumar et al., 2021; Kumar et al., 2023). To illustrate the application of an integrative phenomics approach we discuss how combining multi-omics and advanced phenotyping is being applied to the underutilized oilseed crop Camelina sativa (camelina, gold-of-pleasure, false flax) to facilitate the generation of climate-smart crops for future agricultural systems

Sugars and circadian regulation make major contributions to the global regulation of diurnal gene expression in Arabidopsis

The diurnal cycle strongly influences many plant metabolic and physiological processes. Arabidopsis thaliana rosettes were harvested six times during 12-h-light/12-h-dark treatments to investigate changes in gene expression using ATH1 arrays. Diagnostic gene sets were identified from published or in-house expression profiles of the response to light, sugar, nitrogen, and water deficit in seedlings and 4 h of darkness or illumination at ambient or compensation point [CO2]. Many sugar-responsive genes showed large diurnal expression changes, whose timing matched that of the diurnal changes of sugars. A set of circadian-regulated genes also showed large diurnal changes in expression. Comparison of published results from a free-running cycle with the diurnal changes in Columbia-0 (Col-0) and the starchless phosphoglucomutase (pgm) mutant indicated that sugars modify the expression of up to half of the clock-regulated genes. Principle component analysis identified genes that make large contributions to diurnal changes and confirmed that sugar and circadian regulation are the major inputs in Col-0 but that sugars dominate the response in pgm. Most of the changes in pgm are triggered by low sugar levels during the night rather than high levels in the light, highlighting the importance of responses to low sugar in diurnal gene regulation. We identified a set of candidate regulatory genes that show robust responses to alterations in sugar levels and change markedly during the diurnal cycle

Regulatory principles and experimental approaches to the circadian control of starch turnover

Peer reviewedPublisher PD

L'usage de l'espace par les exploitations d'élevage de montagne et la gestion de la biodiversité

En prenant appui sur divers résultats de recherches menées depuis 20 ans dans les Pyrénées centrales, les caractéristiques de l'utilisation de l'espace par les exploitations agricoles sont présentées. Les particularités spatiales des territoires d'exploitation influent sur les pratiques d'utilisation des prairies et sur la dynamique des couverts et des paysages. Un modèle de référence permet de diagnostiquer la maîtrise de l'exploitation de la végétation. L'impact des modes d'usage des prairies sur leur richesse en espèces est présenté sur quelques cas concrets. Une gestion mal maîtrisée conduit à une augmentation de la diversité intraparcellaire et à une diminution de la diversité interparcellaire, puis à une baisse rapide de ces 2 diversités. Raisonner l'organisation de l'usage des prairies à des niveaux d'organisation larges est une nécessité pour préserver la biodiversité en région de montagn

Stress-Induced GSK3 Regulates the Redox Stress Response by Phosphorylating Glucose-6-Phosphate Dehydrogenase in Arabidopsis.

Diverse stresses such as high salt conditions cause an increase in reactive oxygen species (ROS), necessitating a redox stress response. However, little is known about the signaling pathways that regulate the antioxidant system to counteract oxidative stress. Here, we show that a Glycogen Synthase Kinase3 from Arabidopsis thaliana (ASKα) regulates stress tolerance by activating Glc-6-phosphate dehydrogenase (G6PD), which is essential for maintaining the cellular redox balance. Loss of stress-activated ASKα leads to reduced G6PD activity, elevated levels of ROS, and enhanced sensitivity to salt stress. Conversely, plants overexpressing ASKα have increased G6PD activity and low levels of ROS in response to stress and are more tolerant to salt stress. ASKα stimulates the activity of a specific cytosolic G6PD isoform by phosphorylating the evolutionarily conserved Thr-467, which is implicated in cosubstrate binding. Our results reveal a novel mechanism of G6PD adaptive regulation that is critical for the cellular stress response

PageMan: An interactive ontology tool to generate, display, and annotate overview graphs for profiling experiments

BACKGROUND: Microarray technology has become a widely accepted and standardized tool in biology. The first microarray data analysis programs were developed to support pair-wise comparison. However, as microarray experiments have become more routine, large scale experiments have become more common, which investigate multiple time points or sets of mutants or transgenics. To extract biological information from such high-throughput expression data, it is necessary to develop efficient analytical platforms, which combine manually curated gene ontologies with efficient visualization and navigation tools. Currently, most tools focus on a few limited biological aspects, rather than offering a holistic, integrated analysis. RESULTS: Here we introduce PageMan, a multiplatform, user-friendly, and stand-alone software tool that annotates, investigates, and condenses high-throughput microarray data in the context of functional ontologies. It includes a GUI tool to transform different ontologies into a suitable format, enabling the user to compare and choose between different ontologies. It is equipped with several statistical modules for data analysis, including over-representation analysis and Wilcoxon statistical testing. Results are exported in a graphical format for direct use, or for further editing in graphics programs. PageMan provides a fast overview of single treatments, allows genome-level responses to be compared across several microarray experiments covering, for example, stress responses at multiple time points. This aids in searching for trait-specific changes in pathways using mutants or transgenics, analyzing development time-courses, and comparison between species. In a case study, we analyze the results of publicly available microarrays of multiple cold stress experiments using PageMan, and compare the results to a previously published meta-analysis. PageMan offers a complete user's guide, a web-based over-representation analysis as well as a tutorial, and is freely available at . CONCLUSION: PageMan allows multiple microarray experiments to be efficiently condensed into a single page graphical display. The flexible interface allows data to be quickly and easily visualized, facilitating comparisons within experiments and to published experiments, thus enabling researchers to gain a rapid overview of the biological responses in the experiments