Nothing ventured, nothing gained. In oxygenic photosynthesis, CO₂ is converted into sugar using H₂O and light energy. Additionally, O₂ is released from oxygenic photosynthesis as a by-product. Thus, oxygenic photoautotrophs (i.e., cyanobacteria, algae, and plants) support life on Earth with only sun light energy. However, this process is not risk-free, and every photoautotroph is constantly exposed to potential photo-oxidative damage.

Oxygenic photosynthesis is achieved through the assimilation of CO₂ from the atmosphere in the Calvin-Benson cycle (dark reaction) and the utilization of light for the production of nicotinamide adenine dinucleotide phosphate (NADPH) and adenosine triphosphate (ATP) to meet the demand for CO₂ assimilation in the photosynthetic electron transport system (light reaction). Light energy from the sun is absorbed by pigments, such as chlorophyll (Chl), in light-harvesting complexes (LHC) around two photosystems (PSII and PSI) on the thylakoid membrane, and excites the reaction center Chls (P680 and P700 in PSII and PSI, respectively) to drive charge separation and photosynthetic linear electron flow from PSII to PSI via plastoquinone (PQ) pool, cytochrome (Cyt) b₆/f complex, and plastocyanin (PC) or Cyt c₆ (Barber and Andersson, 1994). On the electron donor side of PSII, photo-oxidized P680 (P680⁺) oxidizes H₂O with O₂ evolved with the help of the oxygen-evolving complex (Nathan and Wolfgang, 2015). On the electron acceptor side of PSI, NADP⁺ is reduced to NADPH with electrons from P700 via ferredoxin (Fd) and Fd-NADP⁺ reductase (FNR). Photo-oxidized P700 (P700⁺) is reduced with electrons from PSII via the Cyt b₆/f complex and PC (or Cyt c₆) in photosynthetic linear electron flow (Jensen et al., 2007). The proton gradient across the thylakoid membrane (ΔpH), which is the motive force of the chloroplast ATP synthase (ATPase), is established at both the steps of H₂O oxidation on the luminal side of PSII and electron transport in the Q-cycle of the Cytb₆f complex (Cramer et al., 2006). NADPH and ATP produced in the photosynthetic electron transport system are utilized for driving CO₂ assimilation in the Calvin-Benson cycle (Calvin and Benson, 1948) and are required to be in concordance. However, dynamic natural environmental variations can easily unbalance the production and utilization of NADPH and ATP, resulting in photo-oxidative damage to photosynthetic cells.

Absorption of light energy exceeding the demand for photosynthetic CO₂ assimilation can cause inactivation of these photosystems. This light-dependent inactivation of photosynthesis was first observed in the green alga Chlorella sp. more than 50 years ago and was termed photoinhibition by Kok (1956). Based on the characterization of photoinhibition using isolated thylakoid membranes, it has been separately recognized as deriving from the inactivation of PSII or PSI (Satoh, 1970). Photoinhibition of PSII leads to light-dependent degradation of the reaction center subunit (D1 protein), which is observed in vivo under various stress conditions, such as high light, high temperature, and drought. Photoinhibited PSII can rapidly recover by replacing the degraded D1 protein with a newly synthesized D1 protein in several hours. The processes that lead to the photo-oxidative damage of PSII are still under debate, although numerous studies have provided extensive and remarkable insights into the mechanisms of PSII photoinhibition (Krause et al., 1985; Aro et al., 1993; Sundby et al., 1993; Andersson and Barber, 1996; Neidhardt et al., 1998; Melis, 1999; Allakhverdiev and Murata, 2004; Hakala et al., 2005; Murata et al., 2007; Tyystjärvi, 2008; Fischer et al., 2013; Pospíšil, 2016; Jimbo et al., 2018).

In this review, we concentrate on the photoinhibition of PSI, which depends on both O₂ and electrons produced by PSII (Satoh, 1970). Compared with studies on PSII photoinhibition, those on PSI photoinhibition are relatively rare, since it hardly occurs in vivo, even if oxygenic photoautotrophs are exposed to a stress treatment with excess light (e.g., continuous light illumination with high light) (Critchley, 1981; Powles and Björkman, 1982; Havaux and Eyletters, 1991). Previous studies, using isolated thylakoid membranes and chloroplasts, have suggested that PSI photoinhibition is derived from a dysfunction in the [4Fe–4S] clusters on the acceptor side of PSI (i.e., F_X, F_A, and F_B), caused by reactive oxygen species (ROS) (Satoh, 1970; Inoue et al., 1986). PSI photoinhibition was first observed in vivo in the intact leaves of the chilling-sensitive plant Cucumis sativus under chilling stress (Terashima et al., 1994). The characterization of PSI photoinhibition under such conditions corroborated previous findings from in vitro studies and worked toward establishing the present theory of the mechanisms of PSI photoinhibition (Sonoike, 2011).

On the electron acceptor side of PSI, excess photo-excitation energy can reduce O₂, generating ROS, including superoxide anion radical (O₂^-), hydrogen peroxide (H₂O₂), and hydroxyl radical (⋅OH) (Mehler, 1951; Asada, 2006; Rutherford et al., 2012). Owing to their high reactivity, ROS can immediately inactivate PSI (Sonoike, 1996; Sonoike et al., 1997). In comparison with PSII, the damaged PSI takes a long time (days or weeks) to completely recover (Kudoh and Sonoike, 2002; Zivcak et al., 2015b). Therefore, PSI photoinhibition is a lethal event for oxygenic photoautotrophs. That is why PSI photoinhibition hardly occurs in vivo except for under specific conditions, such as chilling. The mechanisms of prevention of PSI photoinhibition had remained unknown for long.

The generation of ROS in PSI should be strictly suppressed for the purpose of preventing PSI photoinhibition in vivo. Based on the simple concept of oxygenic photosynthesis, the electron acceptor side of PSI is expected to be over-reduced when the Calvin-Benson cycle cannot follow the production of NADPH in the photosynthetic electron transport system. Nevertheless, PSI is always kept in an oxidized state in response to situations where the Calvin-Benson cycle is suppressed, which has been observed using in vivo spectroscopic measurement techniques for P700⁺ (Foyer et al., 1990; Harbinson and Hedley, 1993; Klughammer and Schreiber, 1994; Golding and Johnson, 2003; Miyake et al., 2005). The universal physiological response of oxygenic photoautotrophs is termed “P700 oxidation” and refers to the increase in the ratio of P700⁺ to the total amount of photo-oxidizable P700. In comparison with P700, P700⁺ cannot drive its photo-oxidation/reduction cycle but directly dissipate the photo-excitation energy as heat (Nuijs et al., 1986; Trissl, 1997; Bukhov and Carpentier, 2003). Therefore, P700 oxidation is expected to be directly linked to the quenching of excess light energy in PSI.

Recently, the impact of P700 oxidation on the alleviation of PSI photoinhibition has been demonstrated by a method to easily and selectively induce PSI photoinhibition in intact plant leaves at room temperature (Sejima et al., 2014). In the method named “repetitive short-pulse (SP) illumination (rSP illumination),” SP light (e.g., 300-1000 ms, 2000-20,000 μmol photons m^-2 s^-1) is repetitively applied to plant leaves (e.g., every 10 s) under darkness. This experimental procedure is similar to continuously shooting the plants with a camera with a strobe light at night and can be defined as a severe form of artificial fluctuating light. This is different to illumination with continuous light; during rSP illumination, PSI is inactivated significantly faster than PSII in intact plant leaves, depending on the intensity and length of the SP light (Sejima et al., 2014; Zivcak et al., 2015b). Eliminating or limiting O₂ suppress the inactivation of PSI, indicating that PSI photoinhibition during rSP illumination is caused by ROS (Sejima et al., 2014). This observation corresponds to results of previous studies on chilling-induced PSI photoinhibition (Sonoike, 1996; Sonoike et al., 1997). Therefore, rSP illumination is a useful tool to induce PSI photoinhibition in vivo (Zivcak et al., 2015a,b; Kono and Terashima, 2016; Takagi et al., 2017b; Mikko and Steffen, 2018).

Sejima et al. (2014) have applied rSP illumination to sunflower leaves under constant actinic light at different intensities, producing different P700 oxidation levels (Figures 1A,B), resulting in a linear relationship of P700 oxidation with the alleviation of PSI photoinhibition (Figure 1C) (Sejima et al., 2014). The effects of P700 oxidation on the protection of PSI against photoinhibition is also evidenced by the kinetics of P700⁺ in response to SP light during rSP illumination. In the intact leaves of angiosperms, P700 is excited by SP light and is kept in a reduced state during exposure to SP light (Figure 2A), which suggests that electron transport in PSI is limited on the acceptor side, but not on the donor side, during exposure to SP light. Contrarily, in the presence of a continuously high intensity background light, P700 is kept in an oxidized state during exposure to SP light (Figure 2B), which is due to a change in the limitation step of the electron transport system in PSI from acceptor to donor sides by P700 oxidation system (described in the next chapter). Furthermore, the addition of a far-red light in the background during rSP illumination can also stimulate P700 oxidation in the SP light to suppress PSI photoinhibition (Kono et al., 2017), which might suggest that shaded plants in an understory can efficiently keep P700 oxidized during natural “sunflecks.”

The impact of P700 oxidation on the alleviation of PSI photoinhibition has been observed not only during rSP illumination but also continuous light. In cyanobacteria, the progenitor of oxygenic photosynthesis, P700, is kept in an oxidized state in response to a suppression of photosynthesis under CO₂ limitation, similarly to that in intact plant leaves (Badger and Schreiber, 1993). Shimakawa et al. (2016b) validated the common response of the redox state of PSI to CO₂ limitation in three different cyanobacteria species, Synechocystis sp. PCC 6803, Synechococcus elongatus PCC 7942, and Synechococcus sp. PCC 7002 (Figure 3). Among the mutants deficient in flavodiiron protein (FLV) in each species, only the mutant of Synechococcus sp. PCC 7002 cannot keep P700 oxidized under CO₂ limitation and suffers from PSI photoinhibition. These facts led us to suggest that the fate of PSI is determined by whether P700 can be kept in an oxidized state under excess light conditions. Furthermore, the inactivation of PSI in the mutant of Synechococcus sp. PCC 7002 has been observed even in the range of photon flux density between 200 and 300 μmol photons m^-2 s^-1 (Shimakawa et al., 2016b), which indicates that PSI has the potential to generate ROS and suffer from PSI photoinhibition even under constant light 10 times less intense than sunlight, if the light exceeds the demand of the electron sinks, such as the Calvin-Benson cycle. Overall, without P700 oxidation, oxygenic photoautotrophs would easily suffer from PSI photoinhibition under natural environmental variation. The diverse strategies to keep P700 oxidized in these cyanobacteria species are further discussed at the section “Flavodiiron Protein (FLV)” in the chapter “Regulatory Mechanisms to Keep P700 in an Oxidized State, P700 Oxidation System.”

Besides being a quencher of light energy to suppress the generation of ROS in PSI, P700⁺ is assumed to play other important roles. For example, the charge recombination of P700⁺ with the electron acceptors in PSI (e.g., F_X) can occur in the range of the μs- or ms-order half times (Semenov et al., 2000). That is, P700⁺ can act as an electron sink to oxidize [4Fe–4S] clusters on the acceptor side of PSI, which might suppress the generation of ⋅OH (Sonoike, 1996). Additionally, P700⁺ possibly thermally dissipate excess light energy not only in PSI but also around PSII via energy transfer mechanisms, including state transition and/or spillover (Ueno et al., 2018; Yokono and Akimoto, 2018), which ultimately, also has the potential to alleviate PSII photoinhibition.

P700 oxidation is strictly regulated by diverse molecular mechanisms (collectively termed P700 oxidation system) in oxygenic photoautotrophs (Figure 4). Importantly, the redox state of P700 depends on both the electron donor and acceptor sides of PSI. There are various regulatory mechanisms functioning on both sides of PSI. P700 oxidation is commonly observed in oxygenic photoautotrophs in response to excess light conditions, and the strategies to keep P700 in an oxidized state are diverse. Many diverse ways to oxidize P700 have been recognized already in the photosynthetic prokaryote cyanobacteria (Shimakawa et al., 2016b; Figure 3), which have supposedly developed and changed during the evolutionary history of oxygenic photoautotrophs.

Recent studies have not only supported the hypothetical model of the mechanisms of PSI photoinhibition, established on the basis of the experimental findings of a study based on chilling stress (Sonoike, 2011), but have also provided novel insights into mechanisms for the generation of ROS in PSI. In this review, we revisited the hypothetical model of the mechanisms of PSI photoinhibition and the dynamics of ROS in PSI in oxygenic photoautotrophs. Originally, in chloroplasts the production of the ROS O₂^- on the electron acceptor side of PSI has been defined as the Mehler reaction (Mehler, 1951). There are four electron acceptors for photo-excited P700 in PSI: chlorophyll A₀ (primary acceptor), phylloquinone A₁, and [4Fe–4S] clusters F_X and F_A/F_B. Both phylloquinones exist asymmetrically in the heterodimeric reaction centers of each of PsaA and PsaB (i.e., A_0A, A_0B, A_1A, and A_1B) (Joliot and Joliot, 1999). Among these acceptors, the Mehler reaction has been proposed to mainly occur at A_1B (Kruk et al., 2003) or F_X (Takahashi and Asada, 1988), which is supported by the lower midpoint redox potential (versus NHE) for A_1B (-820 mV) and F_X (-730 mV) (Brettel and Leibl, 2001; Kozuleva and Ivanov, 2010) than estimated (O₂/O₂^-) in the lipid bilayer (from -500 to -600 mV versus NHE) (Wardman, 1990).

The increase in the photo-oxidative damage in PSI during rSP illumination in the range of light intensity for SP light, from 2000 to 20,000 μmol photons m^-2 s^-1 (Sejima et al., 2014), corresponds to the non-light saturation manner of the production of O₂^- at phylloquinones in PSI (Kozuleva et al., 2014), which implies that O₂^- produced by A_1B is likely to cause PSI photoinhibition. The production of O₂^- can occur on both the stromal and luminal sides of PSI (Takahashi and Asada, 1988; Mubarakshina et al., 2006). O₂^- produced during this process should immediately be disproportionated into H₂O₂ by the oxidation of ascorbate and superoxide dismutase in chloroplasts (Scarpa et al., 1983; Miyake and Asada, 1992). Further, H₂O₂ can react with the reduced [4Fe–4S] centers to produce ⋅OH where the acceptor side of PSI is reduced (Youngman and Elstner, 1981; Sonoike et al., 1997). The addition of methyl viologen, which strongly oxidizes the [4Fe–4S] centers and reduces O₂ to O₂^- on the stromal side of PSI, clearly alleviates PSI photoinhibition, which suggests that the photo-oxidative damage in PSI is caused by ⋅OH produced, depending on the reduced [4Fe–4S] centers (Sonoike, 1996) and/or ROS generated inside PSI (Takagi et al., 2016b).

Although all oxygenic photoautotrophs develop a variety of scavenging enzymes for ROS, including superoxide dismutase, ascorbate peroxidase, and catalase (Asada, 2006), at least in a physiological sense, these scavenging enzymes possibly have no impact on the alleviation of PSI photoinhibition. Originally, Terashima et al. (1998) has found the slight accumulation of H₂O₂ with the lower activity of thylakoid-bound ascorbate peroxidase in the C₃ plant Cucumis sativus in the transition to chilling stress where PSI photoinhibition occurs in this plant. Whereas this finding clearly suggests that the ascorbate peroxidase modulates the H₂O₂ concentration in the plant leaves, it is still unclear if the accumulated H₂O₂ causes PSI photoinhibition. Overall, the effects of P700 oxidation can be lost in the isolated thylakoid membrane. Indeed, it has been proposed that the addition of superoxide dismutase and catalase do not alleviate PSI photoinhibition during a high light-stress treatment in an isolated PSI submembrane (Subramanyam et al., 2005). In the cyanobacterial FLV mutant that cannot keep P700 oxidized under CO₂ limitation, PSI photoinhibition is rapidly induced even under constant light with approximately 200 μmol photons m^-2 s^-1 (Shimakawa et al., 2016b). In chloroplasts of plant leaves, PSI photoinhibition is observed during rSP illumination even in the presence of sufficient activities of superoxide dismutase and ascorbate peroxidase (Takagi et al., 2016b). These data clearly suggest that the scavenging enzymes of ROS cannot prevent PSI photoinhibition. That is, once produced, O₂^- is supposed to immediately attack PSI and/or trigger the production of ⋅OH before it is scavenged, which may be supported by the significantly shorter lives of O₂^- (2-4 μs) and ⋅OH (<1 μs) than that of H₂O₂ (1 ms; Van Breusegem et al., 2001). Contrarily, the production and diffusion of H₂O₂ can be easily detected in photosynthetic cells (Michelet et al., 2013; Roach et al., 2015), which is reasonable considering that H₂O₂ functions as a signaling molecule in oxygenic photoautotrophs (Van Breusegem et al., 2001; Gläßer et al., 2014; Dietz et al., 2016). Overall, we propose that the production of ROS leading to PSI photoinhibition can be completely distinguished from those related to dynamic metabolic and signaling mechanisms. It is possible that the different production site of O₂^- causes the different effects of ROS on photosynthetic cells (Takagi et al., 2016b). Unfortunately, identification of the primary site in PSI attacked by ROS is still controversial (Tjus et al., 1999; Subramanyam et al., 2005; Takagi et al., 2016b), and more research is needed to identify the proper dynamics of ROS around PSI in vivo. The qualitative and quantitative relationships between the production of ROS and PSI photoinhibition should be addressed in future.

Besides O₂^- and ⋅OH, singlet O₂ (¹O₂) has been recently suggested to be generated in PSI and cause PSI photoinhibition. In the core and LHC complexes associated with PSI, triplet Chl can produce ¹O₂ to cause PSI photoinhibition, unless carotenoids such as β-carotene quench the triplet Chl (Subramanyam et al., 2005; Cazzaniga et al., 2012, 2016), which is considered a potential mechanism for alleviating PSI photoinhibition, in addition to P700 oxidation system. Long-lived triplet P700 suggests that ¹O₂ is unlikely to be generated from triplet P700 (Setif et al., 1981; Rutherford et al., 2012). Nevertheless, the generation of ¹O₂ originating from triplet P700 has recently been suggested during rSP illumination (Takagi et al., 2016b, 2017b). Ultimately, it is difficult to exclude the possibility that ¹O₂ has an impact on the photo-oxidative damage in PSI under severe excess light and in specific mutants.

Oxygenic photoautotrophs can safely undergo photosynthesis owing to P700 oxidation system. Despite the current poor understanding of the mechanisms of PSI photoinhibition, the effects of P700 oxidation on the alleviation of PSI photoinhibition discussed herein are likely to be true based on a number of experimental results (e.g., Figures 1–3). On the contrary, a recent study has reported an inconsistency between PSI photoinhibition and P700 oxidation in two different shade-established tropical tree species (Huang et al., 2015). Indeed, the degrees of PSI photoinhibition are diverse among a variety of oxygenic photoautotrophs, regardless of P700 oxidation levels (Takagi et al., 2017b), which probably reflects the different levels of robustness of PSI against ROS in each species. Most importantly, P700 oxidation is not directly linked to photosynthesis. Nevertheless, it is impossible for oxygenic photoautotrophs to live without P700 oxidation system because PSI photoinhibition is the lethal event for them (Shimakawa et al., 2016b). These facts reflect that all oxygenic photoautotrophs are confronted with the potential risk of photo-oxidative damage inevitably accompanied with exposure to light and O₂. Diverse molecular mechanisms, i.e., P700 oxidation system, support P700 oxidation (Figure 4). On the contrary, almost all the agents of P700 oxidation system are still not characterized at the molecular level. There would be various ways to keep P700 oxidized. Intriguingly, the cyanobacterium Leptolyngbya sp., the species thriving in the harsh conditions of the desert, has been suggested to induce P700 oxidation by constricting the thylakoid lumen to limit diffusion of PC (Bar-Eyal et al., 2015). Unimaginable diversity of the strategies for P700 oxidation are possibly still unexplored in a variety of oxygenic photoautotrophs.

CM conceived the project and GS wrote the manuscript.

The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest. The reviewer YS declared a past co-authorship with the authors GS and CM to the handling Editor.