Chloroplast NADH dehydrogenase-like complex-mediated cyclic electron flow is the main electron transport route in C4 bundle sheath cells.

The superior productivity of C4 plants is achieved via a metabolic C4 cycle which acts as a CO2 pump across mesophyll and bundle sheath (BS) cells and requires an additional input of energy in the form of ATP. The importance of chloroplast NADH dehydrogenase-like complex (NDH) operating cyclic electron flow (CEF) around Photosystem I (PSI) for C4 photosynthesis has been shown in reverse genetics studies but the contribution of CEF and NDH to cell-level electron fluxes remained unknown. We have created gene-edited Setaria viridis with null ndhO alleles lacking functional NDH and developed methods for quantification of electron flow through NDH in BS and mesophyll cells. We show that CEF accounts for 84% of electrons reducing PSI in BS cells and most of those electrons are delivered through NDH while the contribution of the complex to electron transport in mesophyll cells is minimal. A decreased leaf CO2 assimilation rate and growth of plants lacking NDH cannot be rescued by supplying additional CO2. Our results indicate that NDH-mediated CEF is the primary electron transport route in BS chloroplasts highlighting the essential role of NDH in generating ATP required for CO2 fixation by the C3 cycle in BS cells.

Faster induction of photosynthesis increases biomass and grain yield in glasshouse-grown transgenic Sorghum bicolor overexpressing Rieske FeS.

Sorghum is one of the most important crops providing food and feed in many of the world's harsher environments. Sorghum utilizes the C4 pathway of photosynthesis in which a biochemical carbon-concentrating mechanism results in high CO2 assimilation rates. Overexpressing the Rieske FeS subunit of the Cytochrome b6 f complex was previously shown to increase the rate of photosynthetic electron transport and stimulate CO2 assimilation in the model C4 plant Setaria viridis. To test whether productivity of C4 crops could be improved by Rieske overexpression, we created transgenic Sorghum bicolor Tx430 plants with increased Rieske content. The transgenic plants showed no marked changes in abundances of other photosynthetic proteins or chlorophyll content. The steady-state rates of electron transport and CO2 assimilation did not differ between the plants with increased Rieske abundance and control plants, suggesting that Cytochrome b6 f is not the only factor limiting electron transport in sorghum at high light and high CO2 . However, faster responses of non-photochemical quenching as well as an elevated quantum yield of Photosystem II and an increased CO2 assimilation rate were observed from the plants overexpressing Rieske during the photosynthetic induction, a process of activation of photosynthesis upon the dark-light transition. As a consequence, sorghum with increased Rieske content produced more biomass and grain when grown in glasshouse conditions. Our results indicate that increasing Rieske content has potential to boost productivity of sorghum and other C4 crops by improving the efficiency of light utilization and conversion to biomass through the faster induction of photosynthesis.

Rieske FeS overexpression in tobacco provides increased abundance and activity of cytochrome b6 f.

Photosynthesis is fundamental for plant growth and yield. The cytochrome b6 f complex catalyses a rate-limiting step in thylakoid electron transport and therefore represents an important point of regulation of photosynthesis. Here we show that overexpression of a single core subunit of cytochrome b6 f, the Rieske FeS protein, led to up to a 40% increase in the abundance of the complex in Nicotiana tabacum (tobacco) and was accompanied by an enhanced in vitro cytochrome f activity, indicating a full functionality of the complex. Analysis of transgenic plants overexpressing Rieske FeS by the light-induced fluorescence transients technique revealed a more oxidised primary quinone acceptor of photosystem II (QA ) and plastoquinone pool and faster electron transport from the plastoquinone pool to photosystem I upon changes in irradiance, compared to control plants. A faster establishment of qE , the energy-dependent component of nonphotochemical quenching, in transgenic plants suggests a more rapid buildup of the transmembrane proton gradient, also supporting the increased in vivo cytochrome b6 f activity. However, there was no consistent increase in steady-state rates of electron transport or CO2 assimilation in plants overexpressing Rieske FeS grown in either laboratory conditions or field trials, suggesting that the in vivo activity of the complex was only transiently increased upon changes in irradiance. Our results show that overexpression of Rieske FeS in tobacco enhances the abundance of functional cytochrome b6 f and may have the potential to increase plant productivity if combined with other traits.

Enhanced abundance and activity of the chloroplast ATP synthase in rice through the overexpression of the AtpD subunit.

ATP, produced by the light reactions of photosynthesis, acts as the universal cellular energy cofactor fuelling all life processes. Chloroplast ATP synthase produces ATP using the proton motive force created by solar energy-driven thylakoid electron transport reactions. Here we investigate how increasing abundance of ATP synthase affects leaf photosynthesis and growth of rice, Oryza sativa variety Kitaake. We show that overexpression of AtpD, the nuclear-encoded subunit of the chloroplast ATP synthase, stimulates both abundance of the complex, confirmed by immunodetection of thylakoid complexes separated by Blue Native-PAGE, and ATP synthase activity, detected as higher proton conductivity of the thylakoid membrane. Plants with increased AtpD content had higher CO2 assimilation rates when a stepwise increase in CO2 partial pressure was imposed on leaves at high irradiance. Fitting of the CO2 response curves of assimilation revealed that plants overexpressing AtpD had a higher electron transport rate (J) at high CO2, despite having wild-type-like abundance of the cytochrome b6f complex. A higher maximum carboxylation rate (Vcmax) and lower cyclic electron flow detected in transgenic plants both pointed to an increased ATP production compared with wild-type plants. Our results present evidence that the activity of ATP synthase modulates the rate of electron transport at high CO2 and high irradiance.