Metabolism is essential for plant growth and has become a major target for crop improvement by enhancing nutrient use efficiency. Metabolic engineering is also the basis for producing high-value plant products such as pharmaceuticals, biofuels and industrial biochemicals. An inherent problem for such engineering endeavours is the tendency to view metabolism as a series of distinct metabolic pathways-glycolysis, the tricarboxylic acid cycle, the Calvin-Benson cycle and so on. While these canonical pathways may represent a dominant or frequently occurring flux mode, systematic analyses of metabolism via computational modelling have emphasized the inherent flexibility of the metabolic network to carry flux distributions that are distinct from the canonical pathways. Recent experimental estimates of metabolic network fluxes using 13C-labelling approaches have revealed numerous instances in which non-canonical pathways occur under different conditions and in different tissues. In this Review, we bring these non-canonical pathways to the fore, summarizing the evidence for their occurrence and the context in which they operate. We also emphasize the importance of non-canonical pathways for metabolic engineering. We argue that the introduction of a high-flux pathway to a desired metabolic product will, by necessity, require non-canonical supporting fluxes in central metabolism to provide the necessary carbon skeletons, energy and reducing power. We illustrate this using the overproduction of isoprenoids and fatty acids as case studies.
