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

Abstract

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.

Figure 1

Differential effect of metabolizable sugars, Suc isomers, and an elicitor preparation of F. oxysporum lycopersici (E-FOL) on mRNA regulation and photosynthetic parameters. A, Regulation of mRNAs for extracellular invertase Lin6 and RbcS. Thirty micrograms of total RNA was separated on formaldehyde agarose gels, blotted onto nitrocellulose, and probed with random primer labeled cDNA fragments. Equal loading of RNA was confirmed by ethidium bromide staining of the rRNA (data not shown). The data presented are representative of five independent sets of experiments. B, Regulation of the rate of oxygen evolution (●) and effective photochemical yield Y (○). The data represent the mean values of five independent experiments.

Figure 2

Time course of changes in external sugar concentrations. Suspension cultures cells were treated with 50 mm of the indicated sugars and the remaining concentration in the culture supernatant was determined at the given time points. A, Suspension cultures were treated with Glc (○), palatinose (▴), or turanose (▵). B, Suspension cultures were treated with Suc and the concentration of Suc (●), Glc (○), and Fru (□) in the same supernatant were determined. The data represent the mean values of six independent measurements.

Figure 3

Differential effect of metabolizable sugars, Suc isomers, and an elicitor preparation of F. oxysporum lycopersici (E-FOL) on activation of MAPK. Cells were harvested exactly after 5 min after the addition of stimuli. A, Study of activation of MAPK by different sugars. Mannitol was taken as osmotic control for the different concentrations of sugars used. B, Activation of MAPK by different amount of E-FOL. The data presented in A and B are representative of five independent sets of experiments.

Figure 4

Enzymatic synthesis of 1′-deoxy-1′-fluoro-Suc with a recombinant Suc synthase (SuSy) from potato and alkaline phosphatase (AP).

Figure 5

A, Regulation of mRNAs of extracellular invertase Lin6 and RbcS by fluoro-Suc and Suc. Suspension culture cells were treated with 20 mm fluoro-Suc and 50 mm Suc for 6 h. Thirty micrograms of total RNA was separated on formaldehyde agarose gels, blotted onto nitrocellulose, and probed with random primer-labeled cDNA fragments of extracellular invertase Lin6 or RbcS. Equal loading of RNA was confirmed by ethidium bromide staining of the rRNA (data not shown). B, Activation of MAPK by fluoro-Suc and Suc. The cell cultures were incubated with 20 mm fluoro-Suc and 50 mm Suc and harvested exactly after 5 min for the MAPK assay. The data presented in A and B are representative of three independent sets of experiments.