Extracellular invertase is the key enzyme of an apoplasmic phloem unloading pathway and catalyses the hydrolytic cleavage of the transport sugar sucrose released into the apoplast. This mechanism contributes to long-distance assimilate transport, provides the substrate to sustain heterotrophic growth and generates metabolic signals known to effect various processes of primary metabolism and defence responses. The essential function of extracellular invertase for supplying carbohydrates to sink organs was demonstrated by the finding that antisense repression of an anther-specific isoenzyme provides an efficient method for metabolic engineering of male sterility. The regulation of extracellular invertase by all classes of phytohormones indicates an essential link between the molecular mechanism of phytohormone action and primary metabolism. The up-regulation of extracellular invertase appears to be a common response to various biotic and abiotic stress-related stimuli such as pathogen infection and salt stress, in addition to specific stress-related reactions. Based on the observed co-ordinated regulation of source/sink relations and defence responses by sugars and stress-related stimuli, the identified activation of distinct subsets of MAP kinases provides a mechanism for signal integration and distribution within such complex networks. Sucrose derivatives not synthesized by higher plants, such as turanose, were shown to elicit responses distinctly different from metabolizable sugars and are rather perceived as stress-related stimuli.

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.In recent years, sugars have been recognized as important signal molecules that affect a variety of physiological responses and in particular regulate genes involved in photosynthesis, sink metabolism, and defense response (Koch, 1996;Smeekens, 1998;Roitsch, 1999;Sheen et al., 1999). Whereas the effect of sugars on gene regulation is well established, the nature of the sugar signal, and the molecular mechanisms involved in sugar perception and intracellular signal transmission, are largely unknown. Suc is the major form of translocated carbon in higher plants and was shown to regulate a number of carbohydrate-responsive genes. Whereas in many cases the effects of Suc could be mimicked by hexoses, such as Glc and Fru, a few studies demonstrated the existence of Suc-specific regulatory pathways (Chiou and Bush, 1998;Rook et al., 1998). In principle, a sugar signal could be generated by extracellular recognition via a soluble or membranebound receptor molecule or by intracellular sensing at different stages of sugar metabolism. For hexoses, a dual role of hexokinase in sugar sensing and glycolysis has been proposed (Jang and Sheen, 1997; Jang et al., 1997) that is a matter of a controversial debate (Halford et al., 1999). Additional membranebased sensing systems have been implied both for hexoses and Suc. Primary lines of evidence are ...

Adaptation to elevated temperatures is of major importance for the survival of plants. The role of kinases in heat stress response was studied in tomato by in gel and in solution kinase assays using myelin basic protein as substrate. The application of heat stress in a naturally occurring temperature range resulted in a fast and transient activation of a 50 kDa mitogen-activated protein (MAP) kinase both in a photoautotrophic cell suspension culture and in leaves of mature plants. The heat activation of the MAP kinase was shown to be calcium-dependent. The speci¢c phosphorylation of tomato heat stress transcription factor HsfA3 by a partially puri¢ed preparation of the heat-activated MAP kinase supports a physiological role of the identi¢ed kinase activity in transducing the heat stress signal.

This paper originates from a presentation at the International Conference on Assimilate Transport and Partitioning, Newcastle, NSW, August 1999 Carbohydrates are synthesised in photosynthetically active source tissues and exported, in most species in the form of sucrose, to photosynthetically less active or inactive sink tissues. Sucrose hydrolysis at the site of utilisation contributes to phloem unloading. This phenomenon links sink metabolism with phloem transport to, and partitioning between, sinks. Invertases catalyse the irreversible hydrolysis of sucrose and thus are expected to contribute to carbohydrate partitioning. Different invertase isoenzymes may be distinguished based on their intracellular location, their isoelectric points and pH optima. Extracellular, cell-wall-bound invertase is uniquely positioned to supply carbohydrates to sink tissues via an apoplasmic pathway, and links the transport sugar sucrose to hexose transporters. A number of studies demonstrate an essential function of this invertase isoenzyme for phloem unloading, carbohydrate partitioning and growth of sink tissues. Extracellular invertases were shown to be specifically expressed under conditions that require a high carbohydrate supply to sink tissues. Further, their expression is upregulated by a number of stimuli that affect source–sink relations. Substrate and reaction products of invertases are not only nutri-ents, but also signal molecules. Like hormones and in combination with hormones and other stimuli, they can regu-late many aspects of plant development from gene expression to long-distance nutrient allocation. Based on studies in Chenopodium rubrum, tomato (Lycopersicon esculentum) and tobacco (Nicotiana tabacum), the regulation of extracellular invertase and its function in assimilate partitioning, defence reactions and sugar signal transduction pathways are discussed.

Activation of mitogen-activated protein (MAP) kinases is a common reaction of plant cells in defense-related signal transduction pathways. To gain insight into the mechanisms that determine specificity in response to a particular stimulus, a biochemical approach has been employed. Photoautotrophic suspension culture cells of tomato (Lycopersicon peruvianum) were used as experimental system to characterize MAP kinase activation by different stress-related stimuli. An elicitor preparation of the tomato-specific pathogen Fusarium oxysporum lycopersici was shown to result in the simultaneous induction of four kinase activities that could be separated by ion-exchange chromatography. The simultaneous activation of multiple MAP kinases was further substantiated by distinct pharmacological and immunological properties: a differential sensitivity toward various protein kinase inhibitors and a differential cross-reaction with isoform-specific MAP kinase antibodies. In contrast to the two fungal elicitors chitosan and the F. oxysporum lycopersici preparation, the plant-derived stimuli polygalacturonic acid and salicylic acid were shown to activate distinctly different subsets of MAP kinases. Application of a voltage pulse was introduced as a transient stress-related stimulus that does not persist in the culture. Voltage application activates a distinct set of MAP kinases, resembling those activated by salicylic acid treatment, and generates a refractory state for the salicylic acid response. The inhibitory effect of nifedipine indicates that current application may directly affect voltage-gated calcium channels, thus, providing a tool to study various calcium-dependent pathways.During their whole life, plants need to cope with a variety of attacking pathogens. They developed appropriate defense responses that protect them against impairment by most of them. These defense responses against several different pathogens have been extensively studied, however, the components that determine the specificity of the defense gene activation remain to be elucidated. After a plant cell is challenged by an elicitor, one or more signal transduction pathways are invoked by a ligand-receptor interaction (Nü rnberger, 1999) that lead to the activation of a set of defense-related genes appropriate for the defense against the attacking pathogen (Jabs et al., 1997).The involvement of mitogen-activated protein (MAP) kinases in biotic and abiotic stress-mediated defense gene activation has been extensively studied and MAP kinases that respond to elicitors (Zhang et al., 1998), wounding (Stratmann and Ryan, 1997), cold and drought stress (Jonak et al., 1996), salinity (Munnik et al., 1999), and endogenous signals (Zhang and Klessig, 1997) have been described. The activities were assigned to MAP kinase genes by the observation of coordinated transcriptional up-regulation and the use of specific antibodies. In-depth analysis of MAP kinase activation in tobacco revealed that the MAP kinase salicylate-induced protein kinase is activated by salicyli...