Lessons from relatives: C4 photosynthesis enhances CO2 assimilation during the low-light phase of fluctuations

Despite the global importance of species with C4 photosynthesis, there is a lack of consensus regarding C4 performance under fluctuating light. Contrasting hypotheses and experimental evidence suggest that C4 photosynthesis is either less or more efficient in fixing carbon under fluctuating light than the ancestral C3 form. Two main issues have been identified that may underly the lack of consensus: neglect of evolutionary distance between selected C3 and C4 species and use of contrasting fluctuating light treatments. To circumvent these issues, we measured photosynthetic responses to fluctuating light across 3 independent phylogenetically controlled comparisons between C3 and C4 species from Alloteropsis, Flaveria, and Cleome genera under 21% and 2% O2. Leaves were subjected to repetitive stepwise changes in light intensity (800 and 100 µmol m-2 s-1 photon flux density) with 3 contrasting durations: 6, 30, and 300 s. These experiments reconciled the opposing results found across previous studies and showed that (i) stimulation of CO2 assimilation in C4 species during the low-light phase was both stronger and more sustained than in C3 species; (ii) CO2 assimilation patterns during the high-light phase could be attributable to species or C4 subtype differences rather than photosynthetic pathway; and (iii) the duration of each light step in the fluctuation regime can strongly influence experimental outcomes.