Sugar Signaling In Plants Research Articles

A reactive oxygen species Ca2+ signalling pathway identified from a chemical screen for modifiers of sugar-activated circadian gene expression.

Sugars are essential metabolites for energy and anabolism that can also act as signals to regulate plant physiology and development. Experimental tools to disrupt major sugar signalling pathways are limited. We performed a chemical screen for modifiers of activation of circadian gene expression by sugars to discover pharmacological tools to investigate and manipulate plant sugar signalling. Using a library of commercially available bioactive compounds, we identified 75 confident hits that modified the response of a circadian luciferase reporter to sucrose in dark-adapted Arabidopsis thaliana seedlings. We validated the transcriptional effect on a subset of the hits and measured their effects on a range of sugar-dependent phenotypes for 13 of these chemicals. Chemicals were identified that appear to influence known and unknown sugar signalling pathways. Pentamidine isethionate was identified as a modifier of a sugar-activated Ca2+ signal that acts as a calmodulin inhibitor downstream of superoxide in a metabolic signalling pathway affecting circadian rhythms, primary metabolism and plant growth. Our data provide a resource of new experimental tools to manipulate plant sugar signalling and identify novel components of these pathways.

Exploring the Contribution of Autophagy to the Excess-Sucrose Response in Arabidopsis thaliana.

Autophagy is an essential intracellular eukaryotic recycling mechanism, functioning in, among others, carbon starvation. Surprisingly, although autophagy-deficient plants (atg mutants) are hypersensitive to carbon starvation, metabolic analysis revealed that they accumulate sugars under such conditions. In plants, sugars serve as both an energy source and as signaling molecules, affecting many developmental processes, including root and shoot formation. We thus set out to understand the interplay between autophagy and sucrose excess, comparing wild-type and atg mutant seedlings. The presented work showed that autophagy contributes to primary root elongation arrest under conditions of exogenous sucrose and glucose excess but not during fructose or mannitol treatment. Minor or no alterations in starch and primary metabolites were observed between atg mutants and wild-type plants, indicating that the sucrose response relates to its signaling and not its metabolic role. Extensive proteomic analysis of roots performed to further understand the mechanism found an accumulation of proteins essential for ROS reduction and auxin maintenance, which are necessary for root elongation, in atg plants under sucrose excess. The analysis also suggested mitochondrial and peroxisomal involvement in the autophagy-mediated sucrose response. This research increases our knowledge of the complex interplay between autophagy and sugar signaling in plants.

Molecular cues of sugar signaling in plants.

Sugars, the chemically bound form of energy, are formed by the absorption of photosynthetically active radiation and fixation in plants. During evolution, plants availed the sugar molecules as a resource, balancing molecule, and signaling molecule. The multifaceted role of sugar molecules in response to environmental stimuli makes it the central coordinator required for growth, survival, and continuity. During the course of evolution, the molecular networks have become complex to adapt or acclimate to the changing environment. Sugar molecules are sensed both intra and extracellularly by their specific sensors. The signal is transmitted by a signaling loop that involves various downstream signaling molecules, transcriptional factors and, most pertinent, the sensors TOR and SnRK1. In this review, the focus has been retained on the significance of the sugar sensors during signaling and induced modules to regulate plant growth, development, biotic and abiotic stress. It is interesting to visualize the sugar molecule as a signaling unit and not only a nutrient. Complete information on the downstream components of sugar signaling will open the gates for improving the qualitative and quantitative elements of crop plants.

Mechanism of Sugar Signaling in Plants

Mechanism of Sugar Signaling in Plants

Overexpression of TaCML20, a calmodulin-like gene, enhances water soluble carbohydrate accumulation and yield in wheat.

Calcium (Ca2+ ) is a universal messenger that mediates intracellular responses to extracellular stimuli in living organisms. Calmodulin (CaM) and calmodulin-like (CML) proteins are the important Ca2+ sensors in plants that decode Ca2+ -signatures to execute downstream intracellular level responses. Several studies indicate the interlinking of Ca2+ and sugar signaling in plants; however, no genes have been functionally characterized to provide molecular evidence. Our study found that expression of TaCML20 was significantly correlated with water soluble carbohydrate (WSC) concentrations in recombinant inbred lines in wheat. TaCML20 has four EF-hand motifs that may facilitate the binding of Ca2+ . To explore the role of CML20, we generated TaCML20 overexpressing transgenic lines in wheat. These lines accumulated higher WSC concentrations in the shoots, and we also found a significantly increased transcript level of sucrose:sucrose-1-fructosyltransferase (1-SST) in the internodes compared with the control plants. In addition, TaCML20 overexpressing plants showed significantly increased tillers per plant and also increased about 19% of grain weight per plant compared with control plants. The results also suggested a role for TaCML20 in drought stress, as its transcripts significantly increased in the shoots of wild-type plants under water deficit. These results uncovered the role of CML20 in determining multiple traits in wheat.

Chemical intervention in plant sugar signalling increases yield and resilience.

The pressing global issue of food insecurity due to population growth, diminishing land and variable climate can only be addressed in agriculture by improving both maximum crop yield potential and resilience. Genetic modification is one potential solution, but has yet to achieve worldwide acceptance, particularly for crops such as wheat. Trehalose-6-phosphate (T6P), a central sugar signal in plants, regulates sucrose use and allocation, underpinning crop growth and development. Here we show that application of a chemical intervention strategy directly modulates T6P levels in planta. Plant-permeable analogues of T6P were designed and constructed based on a 'signalling-precursor' concept for permeability, ready uptake and sunlight-triggered release of T6P in planta. We show that chemical intervention in a potent sugar signal increases grain yield, whereas application to vegetative tissue improves recovery and resurrection from drought. This technology offers a means to combine increases in yield with crop stress resilience. Given the generality of the T6P pathway in plants and other small-molecule signals in biology, these studies suggest that suitable synthetic exogenous small-molecule signal precursors can be used to directly enhance plant performance and perhaps other organism function.

Kinome profiling of sugar signaling in plants using multiple platforms

Although the primary sequence of kinases shows substantial divergence between unrelated eukaryotes, variation in the motifs that are actually phosphorylated by eukaryotic kinases is much smaller. Hence arrays developed for kinome profiling of mammalian cells are useful for kinome profiling of plant tissues as well, facilitating the study of plant signal transduction. We recently employed the Pepscan kinomics chip to reveal the small GTPases in plant sucrose signaling. Here we show that employing a different peptide library (the Pepscan kinase chip) largely similar results are obtained, confirming these earlier data, but such a different library also contributes new insights into the molecular details mediating plant cell responses to a sugar stimulus. Thus when studying plant signal transduction employing peptide arrays, using multiple platforms both increases the confidence of results and provides additional information.

Application of Rice Nuclear Proteome Analysis to the Identification of Evolutionarily Conserved and Glucose-Responsive Nuclear Proteins

In the current study, we apply nuclear proteome analysis to the search for novel nuclear proteins that play evolutionarily conserved roles in the sugar response in plants. We initially identified 563 different proteins by nanoLC/ESI/MS/MS analysis of extracts from rice nuclei that were purified by Percoll density gradient centrifugation, whereas 307 different proteins were identified with nucleic acid-associated proteins that were enriched by DNA affinity chromatography. Since some proteins were identified in both samples, the identified proteins summed up to 657 different proteins. Among them, we found not only transcription and splicing factors, but also a mediator of sugar signaling in plants, hexokinase. For identification of evolutionarily conserved and sugar-responsive nuclear factors, we performed in silico screening that included the identification of Arabidopsis homologues of our panel of rice proteins and investigation of the sugar-responsiveness of the corresponding genes. Since two WD40-like proteins and an armadillo/pumilio-like protein were selected as candidates for such nuclear factors, we confirmed the nuclear localization of these proteins and sugar-induced expression of their corresponding genes in both Arabidopsis and rice. We thus improve our knowledge of plant nuclear proteins and present a novel approach to the identification of factors with particular characteristics.

D-Glucose sensing by a plasma membrane regulator of G signaling protein, AtRGS1

Plants use sugars as signaling molecules and possess mechanisms to detect and respond to changes in sugar availability, ranging from the level of secondary signaling molecules to altered gene transcription. G-protein-coupled pathways are involved in sugar signaling in plants. The Arabidopsis thaliana regulator of G-protein signaling protein 1 ( AtRGS1) combines a receptor-like seven transmembrane domain with an RGS domain, interacts with the Arabidopsis Gα subunit ( AtGPA1) in a d-glucose-regulated manner, and stimulates AtGPA1 GTPase activity. We determined that AtRGS1 interacts with additional components, genetically defined here, to serve as a plasma membrane sensor for d-glucose. This interaction between AtRGS1 and AtGPA1 involves, in part, the seven-transmembrane domain of AtRGS1. Structured summary MINT- 6743118: RGS1 (uniprotkb: Q8H1F2) and GPA1 (uniprotkb: P18064) physically interact (MI: 0218) by bimolecular fluorescence complementation (MI: 0809)

Antisense oligodeoxynucleotide inhibition as a potent strategy in plant biology: identification of SUSIBA2 as a transcriptional activator in plant sugar signalling

Sugar signalling cascades are important components of regulatory networks in cells. Compared with the situation in bacteria, yeast and animals, participants of the sugar signalling pathways in plants are poorly understood. Several genes involved in starch synthesis are known to be sugar inducible, although the signal transduction pathways remain undisclosed. We reported recently the isolation of SUSIBA2, a transcription factor involved in sugar-mediated regulation of starch synthesis. Here, we used antisense oligodeoxynucleotide (ODN) inhibition, a powerful approach in medical sciences, to block the effects of SUSIBA2 in sugar-treated barley leaves. The uptake and intracellular trafficking of an 18-mer susiba2 antisense ODN in leaves were followed by confocal microscopy. Administration of the antisense ODN to the leaves impeded susiba2 expression by RNase H activation. This dramatically diminished the ectopic expression of the iso1 and sbeIIb genes and resulted in altered starch synthesis. This study illustrates the successful exploitation of the antisense ODN technology in plant biology, e.g. as a rapid antecedent to time-consuming transgenic studies, and identifies SUSIBA2 as a transcriptional activator in plant sugar signalling. Based on our findings, we propose a model for sugar-signalling control of starch synthesis.

Sugar sensing and signalling networks in plants

Plant sugar signalling operates in a complex network with plant-specific hormone signalling pathways. Hexokinase was identified as an evolutionarily conserved glucose sensor that integrates light, hormone and nutrient signalling to control plant growth and development.

Transcription factors for sugar-inducible genes

Expression of a variety of sink-related genes for the synthesis of storage compounds are up-regulated in response to increased levels of sugars, while expression of many genes involved in photosynthesis and the breakdown of storage compounds is repressed by sugars. Despite of extensive studies on sugar signaling in plants, only few transcription factors involved in the expression of sugar-inducible genes have been identified so far. A transgenic Arabidopsis thaliana line carrying luciferase gene under the control of a short sugar-inducible promoter derived from a sweet potato sporamin gene was used to screen for loss-of-function type as well as gain-of-function type mutants. These genetic screens resulted in the identification of novel transcription factors that are involved in the expression of at least a subset of sugar-inducible genes in Arabidopsis.

Biochemical and Functional Characterization of PKS11, a Novel Arabidopsis Protein Kinase

The Arabidopsis SOS2 (Salt Overly Sensitive 2)-like protein kinases (PKS) are novel protein kinases that contain an SNF1-like catalytic domain with a putative activation loop and a regulatory domain with an FISL motif that binds calcium sensors. Very little biochemical and functional information is currently available on this family of kinases. Here we report on the expression of the PKS11 gene, activation and characterization of the gene product, and transgenic evaluation of its function in plants. PKS11 transcript was preferentially expressed in roots of Arabidopsis plants. Recombinant glutathione S-transferase fusion protein of PKS11 was inactive in substrate phosphorylation. However, the kinase can be highly activated by a threonine 161 to aspartate substitution (designated PKS11T161D) in the putative activation loop. Interestingly, PKS11 can also be activated by substitution of either a serine or tyrosine with aspartate within the activation loop. Deletion of the FISL motif also resulted in a slight activation of PKS11. PKS11T161D displayed an uncommon preference for Mn(2+) over Mg(2+) for substrate phosphorylation and autophosphorylation. The optimal pH and temperature values of PKS11T161D were determined to be 7.5 and 30 degrees C, respectively. The activated kinase showed substrate specificity, high affinity, and catalytic efficiency for a peptide substrate p3 and for ATP. AMP or ADP at concentrations from 10 microm to 1 mm did not activate PKS11T161D. Transgenic Arabidopsis plants expressing PKS11T161D were more resistant to high concentrations of glucose, suggesting the involvement of this protein kinase in sugar signaling in plants. These results provide insights into the function as well as regulation and biochemical properties of the PKS protein kinase.

Metabolizable and non-metabolizable sugars activate different signal transduction pathways in tomato.

To gain insight into the regulatory mechanisms of sugar signaling in plants, the effect of derivatives of the transport sugar sucrose (Suc), the Suc isomers palatinose and turanose, and the Suc analog fluoro-Suc were tested. Photo-autotrophic suspension culture cells of tomato (Lycopersicon peruvianum) were used to study their effect on the regulation of marker genes of source and sink metabolism, photosynthesis, and the activation of mitogen-activated protein kinases (MAPKs). Suc and glucose (Glc) resulted in reverse regulation of source and sink metabolism. Whereas the mRNA level of extracellular invertase (Lin6) was induced, the transcript level of small subunit of ribulose bisphosphate carboxylase (RbcS) was repressed. In contrast, turanose, palatinose, and fluoro-Suc only rapidly induced Lin6 mRNA level, whereas the transcript level of RbcS was not affected. The differential effect of the metabolizable and non-metabolizable sugars on RbcS mRNA regulation was reflected by the fact that only Suc and Glc inhibited photosynthesis and chlorophyll fluorescence. The activation of different signal transduction pathways by sugars was further supported by the analysis of the activation of MAPKs. MAPK activity was found to be strongly activated by turanose, palatinose, and fluoro-Suc, but not by Suc and Glc. To analyze the role of sugars in relation to pathogen perception, an elicitor preparation of Fusarium oxysporum lycopersici was used. The strong activation of MAPKs and the fast and transient induction of Lin6 expresssion by the fungal elicitor resembles the effect of turanose, palatinose, and fluoro-Suc and indicates that non-metabolizable sugars are sensed as stress-related stimuli.