A dual function of SnRK2 kinases in the regulation of SnRK1 and plant growth

[EN] Adverse environmental conditions trigger responses in plants that promote stress tolerance and survival at the expense of growth(1). However, little is known of how stress signalling pathways interact with each other and with growth regulatory components to balance growth and stress responses. Here, we show that plant growth is largely regulated by the interplay between the evolutionarily conserved energy-sensing SNF1-related protein kinase 1 (SnRK1) protein kinase and the abscisic acid (ABA) phytohormone pathway. While SnRK2 kinases are main drivers of ABA-triggered stress responses, we uncover an unexpected growth-promoting function of these kinases in the absence of ABA as repressors of SnRK1. Sequestration of SnRK1 by SnRK2-containing complexes inhibits SnRK1 signalling, thereby allowing target of rapamycin (TOR) activity and growth under optimal conditions. On the other hand, these complexes are essential for releasing and activating SnRK1 in response to ABA, leading to the inhibition of TOR and growth under stress. This dual regulation of SnRK1 by SnRK2 kinases couples growth control with environmental factors typical for the terrestrial habitat and is likely to have been critical for the water-to-land transition of plants.We thank J.-K. Zhu for the snrk2 mutants, M. Bennett for the SnRK2.2-GFP line, C. Koncz for the SnRK1-GFP line, X. Li for the SnRK2.3-FLAG OE line, J. Schroeder for the GFP-His-FLAG and SnRK2.6-His-FLAG OE lines, C. Mackintosh for the TPS5 antibody and the Nottingham Arabidopsis stock centre for T-DNA mutant seeds. The IGC Plant Facility (Vera Nunes) is thanked for excellent plant care. This work was supported by Fundacao para a Ciencia e a Tecnologia through the R&D Units UIDB/04551/2020 (GREEN-IT-Bioresources for Sustainability) and UID/MAR/04292/2019, FCT project nos. PTDC/BIA-PLA/7143/2014, LISBOA-01-0145-FEDER-028128 and PTDC/BIA-BID/32347/2017, and FCT fellowships/contract nos. SFRH/BD/122736/2016 (M.A.), SFRH/BPD/109336/2015 (A.C.), PD/BD/150239/2019 (D.R.B.), and IF/00804/2013 (E.B.G.). Work in P.L.R.'s laboratory was funded by MCIU grant no. BIO2017-82503-R. C.M. thanks the LabEx Paris Saclay Plant Sciences-SPS (ANR-10-LABX-040-SPS) for support. B.B.P. was funded by Programa VALi+d GVA APOSTD/2017/039. This project has received funding from the European Union Horizon 2020 research and innovation programme (grant agreement no. 867426-ABA-GrowthBalance-H2020-WF-2018-2020/H2020-WF-01-2018, awarded to B.B.P.). This work is dedicated to the memory of our beloved friend and colleague Americo Rodrigues.Belda-Palazón, B.; Adamo, M.; Valerio, C.; Ferreira, LJ.; Confraria, A.; Reis-Barata, D.; Rodrigues, A.... (2020). A dual function of SnRK2 kinases in the regulation of SnRK1 and plant growth. 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Natural Variation in Arabidopsis thaliana as a Tool for Highlighting Differential Drought Responses

To test whether natural variation in Arabidopsis could be used to dissect out the genetic basis of responses to drought stress, we characterised a number of accessions. Most of the accessions belong to a core collection that was shown to maximise the genetic diversity captured for a given number of individual accessions in Arabidopsis thaliana. We measured total leaf area (TLA), Electrolyte Leakage (EL), Relative Water Content (RWC), and Cut Rosette Water Loss (CRWL) in control and mild water deficit conditions. A Principal Component Analysis revealed which traits explain most of the variation and showed that some accessions behave differently compared to the others in drought conditions, these included Ita-0, Cvi-0 and Shahdara. This study relied on genetic variation found naturally within the species, in which populations are assumed to be adapted to their environment. Overall, Arabidopsis thaliana showed interesting phenotypic variations in response to mild water deficit that can be exploited to identify genes and alleles important for this complex trait

Mg-chelatase H subunit affects ABA signaling in stomatal guard cells, but is not an ABA receptor in Arabidopsis thaliana

Mg-chelatase H subunit (CHLH) is a multifunctional protein involved in chlorophyll synthesis, plastid-to-nucleus retrograde signaling, and ABA perception. However, whether CHLH acts as an actual ABA receptor remains controversial. Here we present evidence that CHLH affects ABA signaling in stomatal guard cells but is not itself an ABA receptor. We screened ethyl methanesulfonate-treated Arabidopsis thaliana plants with a focus on stomatal aperture-dependent water loss in detached leaves and isolated a rapid transpiration in detached leaves 1 (rtl1) mutant that we identified as a novel missense mutant of CHLH. The rtl1 and CHLH RNAi plants showed phenotypes in which stomatal movements were insensitive to ABA, while the rtl1 phenotype showed normal sensitivity to ABA with respect to seed germination and root growth. ABA-binding analyses using 3H-labeled ABA revealed that recombinant CHLH did not bind ABA, but recombinant pyrabactin resistance 1, a reliable ABA receptor used as a control, showed specific binding. Moreover, we found that the rtl1 mutant showed ABA-induced stomatal closure when a high concentration of extracellular Ca2+ was present and that a knockout mutant of Mg-chelatase I subunit (chli1) showed the same ABA-insensitive phenotype as rtl1. These results suggest that the Mg-chelatase complex as a whole affects the ABA-signaling pathway for stomatal movements

Light-Induced Stomatal Opening Is Affected by the Guard Cell Protein Kinase APK1b.

Guard cells allow land plants to survive under restricted or fluctuating water availability. They control the exchange of gases between the external environment and the interior of the plant by regulating the aperture of stomatal pores in response to environmental stimuli such as light intensity, and are important regulators of plant productivity. Their turgor driven movements are under the control of a signalling network that is not yet fully characterised. A reporter gene fusion confirmed that the Arabidopsis APK1b protein kinase gene is predominantly expressed in guard cells. Infrared gas analysis and stomatal aperture measurements indicated that plants lacking APK1b are impaired in their ability to open their stomata on exposure to light, but retain the ability to adjust their stomatal apertures in response to darkness, abscisic acid or lack of carbon dioxide. Stomatal opening was not specifically impaired in response to either red or blue light as both of these stimuli caused some increase in stomatal conductance. Consistent with the reduction in maximum stomatal conductance, the relative water content of plants lacking APK1b was significantly increased under both well-watered and drought conditions. We conclude that APK1b is required for full stomatal opening in the light but is not required for stomatal closure

ESKIMO1 Disruption in Arabidopsis Alters Vascular Tissue and Impairs Water Transport

Water economy in agricultural practices is an issue that is being addressed through studies aimed at understanding both plant water-use efficiency (WUE), i.e. biomass produced per water consumed, and responses to water shortage. In the model species Arabidopsis thaliana, the ESKIMO1 (ESK1) gene has been described as involved in freezing, cold and salt tolerance as well as in water economy: esk1 mutants have very low evapo-transpiration rates and high water-use efficiency. In order to establish ESK1 function, detailed characterization of esk1 mutants has been carried out. The stress hormone ABA (abscisic acid) was present at high levels in esk1 compared to wild type, nevertheless, the weak water loss of esk1 was independent of stomata closure through ABA biosynthesis, as combining mutant in this pathway with esk1 led to additive phenotypes. Measurement of root hydraulic conductivity suggests that the esk1 vegetative apparatus suffers water deficit due to a defect in water transport. ESK1 promoter-driven reporter gene expression was observed in xylem and fibers, the vascular tissue responsible for the transport of water and mineral nutrients from the soil to the shoots, via the roots. Moreover, in cross sections of hypocotyls, roots and stems, esk1 xylem vessels were collapsed. Finally, using Fourier-Transform Infrared (FTIR) spectroscopy, severe chemical modifications of xylem cell wall composition were highlighted in the esk1 mutants. Taken together our findings show that ESK1 is necessary for the production of functional xylem vessels, through its implication in the laying down of secondary cell wall components

The Arabidopsis ABA-Activated Kinase OST1 Phosphorylates the bZIP Transcription Factor ABF3 and Creates a 14-3-3 Binding Site Involved in Its Turnover

indicates that members of the Snf1-Related Kinases 2 family (SnRK2) are essential in mediating various stress-adaptive responses. Recent reports have indeed shown that one particular member, OPEN STOMATA (OST)1, whose kinase activity is stimulated by the stress hormone abscisic acid (ABA), is a direct target of negative regulation by the core ABA co-receptor complex composed of PYR/PYL/RCAR and clade A Protein Phosphatase 2C (PP2C) proteins. and that phospho-T451 is important for stabilization of ABF3. on T451 to create a 14-3-3 binding motif. In a wider physiological context, we propose that the long term responses to ABA that require sustained gene expression is, in part, mediated by the stabilization of ABFs driven by ABA-activated SnRK2s

Signal transduction-related responses to phytohormones and environmental challenges in sugarcane

BACKGROUND: Sugarcane is an increasingly economically and environmentally important C4 grass, used for the production of sugar and bioethanol, a low-carbon emission fuel. Sugarcane originated from crosses of Saccharum species and is noted for its unique capacity to accumulate high amounts of sucrose in its stems. Environmental stresses limit enormously sugarcane productivity worldwide. To investigate transcriptome changes in response to environmental inputs that alter yield we used cDNA microarrays to profile expression of 1,545 genes in plants submitted to drought, phosphate starvation, herbivory and N(2)-fixing endophytic bacteria. We also investigated the response to phytohormones (abscisic acid and methyl jasmonate). The arrayed elements correspond mostly to genes involved in signal transduction, hormone biosynthesis, transcription factors, novel genes and genes corresponding to unknown proteins. RESULTS: Adopting an outliers searching method 179 genes with strikingly different expression levels were identified as differentially expressed in at least one of the treatments analysed. Self Organizing Maps were used to cluster the expression profiles of 695 genes that showed a highly correlated expression pattern among replicates. The expression data for 22 genes was evaluated for 36 experimental data points by quantitative RT-PCR indicating a validation rate of 80.5% using three biological experimental replicates. The SUCAST Database was created that provides public access to the data described in this work, linked to tissue expression profiling and the SUCAST gene category and sequence analysis. The SUCAST database also includes a categorization of the sugarcane kinome based on a phylogenetic grouping that included 182 undefined kinases. CONCLUSION: An extensive study on the sugarcane transcriptome was performed. Sugarcane genes responsive to phytohormones and to challenges sugarcane commonly deals with in the field were identified. Additionally, the protein kinases were annotated based on a phylogenetic approach. The experimental design and statistical analysis applied proved robust to unravel genes associated with a diverse array of conditions attributing novel functions to previously unknown or undefined genes. The data consolidated in the SUCAST database resource can guide further studies and be useful for the development of improved sugarcane varieties