Trehalose 6-phosphate (Tre6P) is an essential signal metabolite in plants, linking growth and development to carbon metabolism. The sucrose-Tre6P nexus model postulates that Tre6P acts as both a signal and negative feedback regulator of sucrose levels. To test this model, short-term metabolic responses to induced increases in Tre6P levels were investigated in Arabidopsis thaliana plants expressing the Escherichia coli Tre6P synthase (otsA) under the control of an ethanol-inducible promoter. Increased Tre6P led to a transient drop in sucrose content, post-translational activation of nitrate reductase and phosphoenolpyruvate carboxylase, and increased levels of organic and amino acids. Radio- ((14) CO2 ) and stable-isotope ((13) CO2 ) labelling experiments showed no change in the rates of photoassimilate export in plants with elevated Tre6P, but increased labelling of organic acids. It is concluded that high Tre6P lowers sucrose levels by stimulating nitrate assimilation and anaplerotic synthesis of organic acids, thereby diverting photoassimilates away from sucrose to generate carbon skeletons and fixed nitrogen for amino acid synthesis. These results are consistent with the sucrose-Tre6P nexus model and implicate Tre6P in coordinating carbon and nitrogen metabolism in plants. This article is protected by copyright. All rights reserved.