Filip Rolland

A central integrator of transcription networks in plant stress and energy signalling

Photosynthetic plants are the principal solar energy converter sustaining life on Earth. Despite its fundamental importance, little is known about how plants sense and adapt to darkness in the daily light–dark cycle, or how they adapt to unpredictable environmental stresses that compromise photosynthesis and respiration and deplete energy supplies. Current models emphasize diverse stress perception and signalling mechanisms. Using a combination of cellular and systems screens, we show here that the evolutionarily conserved Arabidopsis thaliana protein kinases, KIN10 and KIN11 (also known as AKIN10/At3g01090 and AKIN11/At3g29160, respectively), control convergent reprogramming of transcription in response to seemingly unrelated darkness, sugar and stress conditions. Sensing and signalling deprivation of sugar and energy, KIN10 targets a remarkably broad array of genes that orchestrate transcription networks, promote catabolism and suppress anabolism. Specific bZIP transcription factors partially mediate primary KIN10 signalling. Transgenic KIN10 overexpression confers enhanced starvation tolerance and lifespan extension, and alters architecture and developmental transitions. Significantly, double kin10 kin11 deficiency abrogates the transcriptional switch in darkness and stress signalling, and impairs starch mobilization at night and growth. These studies uncover surprisingly pivotal roles of KIN10/11 in linking stress, sugar and developmental signals to globally regulate plant metabolism, energy balance, growth and survival. In contrast to the prevailing view that sucrose activates plant SnRK1s (Snf1-related protein kinases), our functional analyses of Arabidopsis KIN10/11 provide compelling evidence that SnRK1s are inactivated by sugars and share central roles with the orthologous yeast Snf1 and mammalian AMPK in energy signalling.

Sugar sensing and signaling in plants

In addition to their essential roles as substrates in carbon and energy metabolism and in polymer biosynthesis, sugars have important hormone-like functions as primary messengers in signal transduction. The pivotal role of sugars as signaling molecules is well illustrated by the variety of sugar

Sugar signals and molecular networks controlling plant growth

In recent years, several regulatory systems that link carbon nutrient status to plant growth and development have emerged. In this paper, we discuss the growth promoting functions of the hexokinase (HXK) glucose sensor, the trehalose 6-phosphate (T6P) signal and the Target of Rapamycin (TOR) kinase pathway, and the growth inhibitory function of the SNF1-related Protein Kinase1 (SnRK1) and the C/S1 bZIP transcription factor network. It is crucial that these systems interact closely in regulating growth and in several cases crosstalk has been demonstrated. Importantly, these nutrient controlled systems must interact with other growth regulatory pathways.

Glucose-sensing mechanisms in eukaryotic cells.

Glucose not only serves as a nutrient but also exerts many hormone-like regulatory effects in a wide variety of eukaryotic cell types. Recently, interest in identifying general mechanisms and principles used to sense the presence of glucose has significantly increased and promising advances have been made: in yeast, the first proteins with an apparently specific function in glucose detection have been discovered; in plant cells, there is increasing evidence for a diverse array of glucose-induced signalling mechanisms; and in mammals, glucose-sensing phenomena have turned out to be much more widespread than just in the well-known example of pancreatic beta cells.

Sugar signalling and antioxidant network connections in plant cells

Sugars play important roles as both nutrients and regulatory molecules throughout plant life. Sugar metabolism and signalling function in an intricate network with numerous hormones and reactive oxygen species (ROS) production, signalling and scavenging systems. Although hexokinase is well known to fulfil a crucial role in glucose sensing processes, a scenario is emerging in which the catalytic activity of mitochondria‐associated hexokinase regulates glucose‐6‐phosphate and ROS levels, stimulating antioxidant defence mechanisms and the synthesis of phenolic compounds. As a new concept, it can be hypothesized that the synergistic interaction of sugars (or sugar‐like compounds) and phenolic compounds forms part of an integrated redox system, quenching ROS and contributing to stress tolerance, especially in tissues or organelles with high soluble sugar concentrations.

Glucose-induced cAMP signalling in yeast requires both a G-protein coupled receptor system for extracellular glucose detection and a separable hexose kinase-dependent sensing process.

In Saccharomyces cerevisiae, glucose activation of cAMP synthesis requires both the presence of the G‐protein‐coupled receptor (GPCR) system, Gpr1‐Gpa2, and uptake and phosphorylation of the sugar. In a hxt‐null strain that lacks all physiologically important glucose carriers, glucose transport as well as glucose‐induced cAMP signalling can be restored by constitutive expression of the galactose permease. Hence, the glucose transporters do not seem to have a regulatory function but are only required for glucose uptake. We established a system in which the GPCR‐dependent glucose‐sensing process is separated from the glucose phosphorylation process. It is based on the specific transport and hydrolysis of maltose providing intracellular glucose in the absence of glucose transport. Preaddition of a low concentration (0.7 mM) of maltose to derepressed hxt‐null cells and subsequent addition of glucose restored the glucose‐induced cAMP signalling, although there was no glucose uptake. Addition of a low concentration of maltose itself does not increase the cAMP level but enhances Glu6P and apparently fulfils the intracellular glucose phosphorylation requirement for activation of the cAMP pathway by extracellular glucose. This system enabled us to analyse the affinity and specificity of the GPCR system for fermentable sugars. Gpr1 displayed a very low affinity for glucose (apparent Ka = 75 mM) and responded specifically to extracellular α and βd‐glucose and sucrose, but not to fructose, mannose or any glucose analogues tested. The presence of the constitutively active Gpa2val132 allele in a wild‐type strain bypassed the requirement for Gpr1 and increased the low cAMP signal induced by fructose and by low glucose up to the same intensity as the high glucose signal. Therefore, the low cAMP increases observed with fructose and low glucose in wild‐type cells result only from the low sensitivity of the Gpr1‐Gpa2 system and not from the intracellular sugar kinase‐dependent process. In conclusion, we have shown that the two essential requirements for glucose‐induced activation of cAMP synthesis can be fulfilled separately: an extracellular glucose detection process dependent on Gpr1 and an intracellular sugar‐sensing process requiring the hexose kinases.

Sugar Sensing and Signaling

Abstract Plants, restricted by their environment, need to integrate a wide variety of stimuli with their metabolic activity, growth and development. Sugars, generated by photosynthetic carbon fixation, are central in coordinating metabolic fluxes in response to the changing environment and in providing cells and tissues with the necessary energy for continued growth and survival. A complex network of metabolic and hormone signaling pathways are intimately linked to diverse sugar responses. A combination of genetic, cellular and systems analyses have uncovered nuclear HXK1 (hexokinase1) as a pivotal and conserved glucose sensor, directly mediating transcription regulation, while the KIN10/11 energy sensor protein kinases function as master regulators of transcription networks under sugar and energy deprivation conditions. The involvement of disaccharide signals in the regulation of specific cellular processes and the potential role of cell surface receptors in mediating sugar signals add to the complexity. This chapter gives an overview of our current insight in the sugar sensing and signaling network and describes some of the molecular mechanisms involved.

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.

The AMPK/SNF1/SnRK1 fuel gauge and energy regulator: structure, function and regulation.

All life forms on earth require a continuous input and monitoring of carbon and energy supplies. The AMP‐activated kinase (AMPK)/sucrose nonfermenting1 (SNF1)/Snf1‐related kinase1 (SnRK1) protein kinases are evolutionarily conserved metabolic sensors found in all eukaryotic organisms from simple unicellular fungi (yeast SNF1) to animals (AMPK) and plants (SnRK1). Activated by starvation and energy‐depleting stress conditions, they enable energy homeostasis and survival by up‐regulating energy‐conserving and energy‐producing catabolic processes, and by limiting energy‐consuming anabolic metabolism. In addition, they control normal growth and development as well as metabolic homeostasis at the organismal level. As such, the AMPK/SNF1/SnRK1 kinases act in concert with other central signaling components to control carbohydrate uptake and metabolism, fatty acid and lipid biosynthesis and the storage of carbon energy reserves. Moreover, they have a tremendous impact on developmental processes that are triggered by environmental changes such as nutrient depletion or stress. Although intensive research by many groups has partly unveiled the factors that regulate AMPK/SNF1/SnRK1 kinase activity as well as the pathways and substrates they control, several fundamental issues still await to be clarified. In this review, we will highlight these issues and focus on the structure, function and regulation of the AMPK/SNF1/SnRK1 kinases.

Exploring the neutral invertase–oxidative stress defence connection in Arabidopsis thaliana

Over the past decades, considerable advances have been made in understanding the crucial role and the regulation of sucrose metabolism in plants. Among the various sucrose-catabolizing enzymes, alkaline/neutral invertases (A/N-Invs) have long remained poorly studied. However, recent findings have demonstrated the presence of A/N-Invs in various organelles in addition to the cytosol, and their importance for plant development and stress tolerance. A cytosolic (At-A/N-InvG, At1g35580) and a mitochondrial (At-A/N-InvA, At1g56560) member of the A/N-Invs have been analysed in more detail in Arabidopsis and it was found that At-A/N-InvA knockout plants show an even more severe growth phenotype than At-A/N-InvG knockout plants. The absence of either A/N-Inv was associated with higher oxidative stress defence gene expression, while transient overexpression of At-A/N-InvA and At-A/N-InvG in leaf mesophyll protoplasts down-regulated the oxidative stress-responsive ascorbate peroxidase 2 (APX2) promoter. Moreover, up-regulation of the APX2 promoter by hydrogen peroxide or abscisic acid could be blocked by adding metabolizable sugars or ascorbate. A hypothetical model is proposed in which both mitochondrial and cytosolic A/N-Invs can generate glucose as a substrate for mitochondria-associated hexokinase, contributing to mitochondrial reactive oxygen species homeostasis.