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TROL-FNR complex is integrated into cellular redox networks and reveals alternative pathways of energy dissipation in photosynthesis

In photosynthesis, final electron transfer from ferredoxin (Fd) to NADP+ is accomplished by the flavoenzyme ferredoxin:NADP+ oxidoreductase (FNR). FNR is recruited to thylakoid membranes via integral membrane thylakoid rodanese-like protein (TROL). TROL is located in the vicinity of photosystem I and its deletion leads to changes in electron transfer efficiency and enhancement of non photochemical quenching. Depending in its location and isoforms, FNR can serve dual function in photosynthetic energy conversion: it catalyzes the production of NADPH and can function as diaphorase, consuming NADPH. Thus, FNR can balance redox status of chloroplast stroma depending on its association with thylakoid membrane and its organization into homo- or hetero-complexes. Here we address the activities of several antioxidant mechanisms (pathways) in TROL knock-out plants (trol) grown under different light regimes. In all tested light environments trol plants accumulated significantly more H2O2 than the WT. Regularly, this should be accounted to the interplay of enzymes such as superoxide dismutase (SOD) and catalase (CAT). In the dark- and growth light (GL)-acclimated trol plants SOD activity remained unchanged, whereas in high light (HL) conditions markedly decreased when compared to the WT. It remains unclear which processes contributed to H2O2 levels increase in all tested light environments, since the CAT activity was significantly lower only in GL acclimated trol plants. To test possible connection of FNR-related ROS antioxidant mechanisms and FNR-dependent NADPH production, we have monitored the activity of glutathione reductase (GR) and the accumulation of glutathione (GSH) in several cell compartments. After a short term light stress GSH levels in trol plants were not increased in any of investigated compartments, suggesting that plants without TROL are much less stressed. Ascorbic acid (AA) cycle works in synchrony with the GR system to prevent or minimize the damage caused by ROS. By monitoring the AA concentration we observed that AA content is reduced in the dark- and HL-acclimated trol plants. Reduction in the content of AA under these conditions is unexpected, but it may indicate that regeneration of AA from its oxidized form is impaired in conditions when unbound FNR is not able to maintain optimal rates of NADPH production, or when stroma is more oxidized. Further, cellular biochemistry can be extensively altered when redox pathways are not optimally poised. ROS exert the most damaging effect on lipids and proteins, resulting in lipid peroxidation (LP) and protein oxidation (PO). Surprisingly, LP was significantly lower in the dark- and HL-acclimated trol plants, while it was only slightly increased under GL conditions. We also addressed the extent of PO, measured by the content of carbonyls (CAR). In both GL and HL conditions CARs were not significantly increased in trol plants, and only slight increase was recorded in dark-acclimated plants. We concluded that trol plants have modified ROS scavenging strategies to efficiently avoid toxic lipid and protein modifications. We conclude that TROL-FNR complex is fully integrated into cellular redox networks and it appears that it can even serve as the source element in various redox pathway of ROS detoxification.

Photosystem I; TROL; ferredoxin:NADP+ oxidoreductase (FNR); ROS; electron partitioning

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