Two cyclic electron flows around photosystem I differentially participate in C₄ photosynthesis

C-4 plants assimilate CO2 more efficiently than C-3 plants because of their C-4 cycle that concentrates CO2. However, the C-4 cycle requires additional ATP molecules, which may be supplied by cyclic electron flow (CEF) around photosystem I. One CEF route, which depends on a chloroplast NADH dehydrogenase-like (NDH) complex, is suggested to be crucial for C-4 plants despite the low activity in C-3 plants. The other route depends on proton gradient regulation 5 (PGR5) and PGR5-like photosynthetic phenotype 1 (PGRL1) and is considered a major CEF route to generate the proton gradient across the thylakoid membrane in C-3 plants. However, its contribution to C-4 photosynthesis is still unclear. In this study, we investigated the contribution of the two CEF routes to the NADP-malic enzyme subtype of C-4 photosynthesis in Flaveria bidentis. We observed that suppressing the NDH-dependent route drastically delayed growth and decreased the CO2 assimilation rate to approximately 30% of the wild-type rate. On the other hand, suppressing the PGR5/PGRL1-dependent route did not affect plant growth and resulted in a CO2 assimilation rate that was approximately 80% of the wild-type rate. Our data indicate that the NDH-dependent CEF substantially contributes to the NADP-malic enzyme subtype of C-4 photosynthesis and that the PGR5/PGRL1-dependent route cannot complement the NDH-dependent route in F. bidentis. These findings support the fact that during C-4 evolution, photosynthetic electron flow may have been optimized to provide the energy required for C-4 photosynthesis.