Regulation of photosynthetic electron flow on dark to light transition by ferredoxin:NADP(H) oxidoreductase interactions.

During photosynthesis, electron transport is necessary for carbon assimilation and must be regulated to minimize free radical damage. There is a longstanding controversy over the role of a critical enzyme in this process (ferredoxin:NADP(H) oxidoreductase, or FNR), and in particular its location within chloroplasts. Here we use immunogold labelling to prove that FNR previously assigned as soluble is in fact membrane associated. We combined this technique with a genetic approach in the model plant Arabidopsis to show that the distribution of this enzyme between different membrane regions depends on its interaction with specific tether proteins. We further demonstrate a correlation between the interaction of FNR with different proteins and the activity of alternative photosynthetic electron transport pathways. This supports a role for FNR location in regulating photosynthetic electron flow during the transition from dark to light.

Ferredoxin:NADP(H) Oxidoreductase Abundance and Location Influences Redox Poise and Stress Tolerance.

In linear photosynthetic electron transport, ferredoxin:NADP(H) oxidoreductase (FNR) transfers electrons from ferredoxin (Fd) to NADP+ Both NADPH and reduced Fd (Fdred) are required for reductive assimilation and light/dark activation/deactivation of enzymes. FNR is therefore a hub, connecting photosynthetic electron transport to chloroplast redox metabolism. A correlation between FNR content and tolerance to oxidative stress is well established, although the precise mechanism remains unclear. We investigated the impact of altered FNR content and localization on electron transport and superoxide radical evolution in isolated thylakoids, and probed resulting changes in redox homeostasis, expression of oxidative stress markers, and tolerance to high light in planta. Our data indicate that the ratio of Fdred to FNR is critical, with either too much or too little FNR potentially leading to increased superoxide production, and perception of oxidative stress at the level of gene transcription. In FNR overexpressing plants, which show more NADP(H) and glutathione pools, improved tolerance to high-light stress indicates that disturbance of chloroplast redox poise and increased free radical generation may help "prime" the plant and induce protective mechanisms. In fnr1 knock-outs, the NADP(H) and glutathione pools are more oxidized relative to the wild type, and the photoprotective effect is absent despite perception of oxidative stress at the level of gene transcription.

N-terminal structure of maize ferredoxin:NADP+ reductase determines recruitment into different thylakoid membrane complexes.

To adapt to different light intensities, photosynthetic organisms manipulate the flow of electrons through several alternative pathways at the thylakoid membrane. The enzyme ferredoxin:NADP(+) reductase (FNR) has the potential to regulate this electron partitioning because it is integral to most of these electron cascades and can associate with several different membrane complexes. However, the factors controlling relative localization of FNR to different membrane complexes have not yet been established. Maize (Zea mays) contains three chloroplast FNR proteins with totally different membrane association, and we found that these proteins have variable distribution between cells conducting predominantly cyclic electron transport (bundle sheath) and linear electron transport (mesophyll). Here, the crystal structures of all three enzymes were solved, revealing major structural differences at the N-terminal domain and dimer interface. Expression in Arabidopsis thaliana of maize FNRs as chimeras and truncated proteins showed the N-terminal determines recruitment of FNR to different membrane complexes. In addition, the different maize FNR proteins localized to different thylakoid membrane complexes on expression in Arabidopsis, and analysis of chlorophyll fluorescence and photosystem I absorbance demonstrates the impact of FNR location on photosynthetic electron flow.