C4 photosynthesis is one of the most impressive examples of convergent evolution with at least 66 independent origins. It employs a metabolic C4 cycle operating across two distinct cell types, mesophyll and bundle sheath (BS) cells, which builds up CO2 partial pressure in the BS, where Rubisco resides. While this CO2 concentrating mechanism allows Rubisco to operate close to maximum efficiency, it also doubles the ATP cost of CO2 fixation. There are three subtypes of C4 photosynthesis with differences in biochemistry and the efficiency of utilizing sunlight for carbon fixation. The subtype which predominantly relies on NADP-malic enzyme (ME) to release CO2 in the C4 cycle is the most efficient and evolutionary successful, featuring many economically critical species like maize, sorghum, and sugarcane.
In the C4 system, electron transport chains of mesophyll and BS chloroplasts are attuned to specific metabolic needs of each cell type. Cyclic electron flow (CEF) builds up proton motive force which can be used for generation of ATP or to initiate energy-dependent non-photochemical quenching (NPQ). There are two proposed CEF pathways mediated by the chloroplast NADH dehydrogenase-like (NDH) complex and by PROTON GRADIENT REGULATION5 (PGR5)/PGR5-LIKE 1 (PGRL1). We investigate specific roles of these pathways in supplying additional ATP and/or initiating NPQ in the two-cell C4 system using gene-edited model NADP-ME grass Setaria viridis. Our results suggest that cell-preferential operation of CEF pathways contributes to the superior efficiency of NADP-ME subtype of C4 photosynthesis.