Metabolite Profiles reveal Inter-Specific Variation in Operation of the Calvin-Benson Cycle in both C4 and C3 plants.

Low atmospheric CO2 in recent geological time led to the evolution of carbon concentrating mechanisms (CCMs) like C4 photosynthesis in over 65 terrestrial plant lineages. We know little about the impact of low CO2 on the Calvin-Benson cycle (CBC) in C3 species that did not evolve CCMs, representing over 90% of terrestrial plant species. Metabolite profiling provides a top-down strategy to investigate the operational balance in a pathway. We profiled CBC-intermediates in a panel of C4 (Zeamays, Setaria viridis, Flaveria bidentis,F.trinervia) and C3 species (Oryza sativa, Triticiumaestivum, Arabidopsis thaliana, Nicotiana tabacum, Manihot esculenta). Principal component analysis revealed differences between C4 and C3 species that were driven by many metabolites, including lower ribulose 1,5-bisphosphate in C4 species. Strikingly, there was also considerable variation between C3 species. This was partly due to different chlorophyll and protein contents, but mainly to differences in relative levels of metabolites. Correlation analysis indicated one contributory factor was the balance between fructose-1,6-bisphosphatase, sedoheptulose-1,7-bisphosphatase, phosphoribulokinase and RuBisCO. Our results point to the CBC having experienced different evolutionary trajectories in C3 species since the ancestors of modern plant lineages diverged. They underline the need to understand CBC operation in a wide range of species.