**2. Adaptation to changes in light conditions**

A distinctive feature of photosynthetic organisms is the presence of light-harvesting systems that funnel the absorbed light energy to the corresponding reaction centers and thereby considerably increase their absorption cross-section. Several regulatory mechanisms operate on these antenna systems for controlling the energy flux to the reaction centers. This is particularly important under changing environmental conditions when the photosynthetic apparatus needs to adapt quickly. Under limiting light, it optimizes its light absorption efficiency by adjusting the relative size of its antenna systems through the reversible allocation of a portion of LHCII between PSII and PSI, a process referred to as state transitions which occurs in algae, plants and cyanobacteria (for reviews see Refs. [17,18]). In contrast, when the absorbed light energy exceeds the capacity of the photosynthetic apparatus, it dissipates the excess excitation energy through nonphotochemical quenching (NPQ) as heat thereby avoiding photodamage (for reviews see Refs. [19,20]).

#### **2.1. State transitions**

Because the antenna systems of PSII and PSI have a different pigment composition, their relative light absorption properties change when the light quality varies. This is especially important for aquatic algae because the penetration of light in water changes depending on its wavelength; in particular, red light is more absorbed than blue light. Another example is provided by photosynthetic organisms growing under a canopy where far red light is enriched. These changes in light quality can result in an unequal excitation of PSII and PSI and thereby perturb the redox poise of the plastoquinone pool. Over excitation of PSII relative to PSI leads to increased reduction of the plastoquinone pool and favors thereby docking of plastoquinol to the Qo site of the Cyt*b*6*f* complex [21,22]. This process leads to activation of the chloroplast protein kinase Stt7/STN7 and to the phosphorylation of several proteins from LHCII [23,24]. Although the direct phosphorylation of LHCII by the St7/STN7 kinase has not yet been demonstrated, this kinase is the best candidate for the LHCII kinase because it is firmly associated with the Cyt*b*6*f* complex, and in its absence, state transitions no longer occur [25]. Furthermore, it is widely conserved in land plants, mosses and algae. As a result of this phosphorylation, a part of the LHCII antenna is detached from PSII and moves and binds to PSI thereby rebalancing the light excitation of PSII and PSI and enhancing photosynthetic yield. This process is reversible as overexcitation of PSI leads to the inactivation of the kinase and to dephosphorylation of LHCII by the PPH1/TAP38 protein phosphatase and its return to PSI [26,27]. Thus two different states can be defined, state 1 and state 2 corresponding to the association of the mobile LHCII antenna to PSII and PSI, respectively. However, a strict causal link between LHCII phosphorylation and its migration from PSII to PSI has been questioned recently by the finding that some phosphorylated LHCII remains associated with PSII supercomplexes and that LHCII serves as antenna for both photosystems under most natural light conditions [13,28,29]. In plants, the LHCII S trimers comprise Lhcb1 and Lhcb2, whereas the M trimers contain Lhcb1 and Lhcb3 [30]. Both the S and M trimers are most likely not involved in state transitions because the PSII-LHCII supercomplex is unchanged upon phosphorylation [29] and PSI does not bind Lhcb3 in state 2 [30]. Thus LHCII phosphorylation is not sufficient to dissociate all LHCII trimers from PSII. It has therefore been proposed that the peripherally bound L trimers associate with PSI in state 2 [30]. Moreover, although Lhcb1 and Lhcb2 display similar phosphorylation kinetics during a state 1 to state 2 transition, only phosphorylated Lhcb2 but not Lhcb1 is part of the PSI-LHCII supercomplex [31]. A PSI supercomplex has been isolated and characterized in *Chlamydomonas*. It consists of PSI, Cyt*b*6*f*, LHCII, FNR (ferredoxin–NADPH reductase), PGRL1, a protein involved in CEF [3] and additional factors (**Figure 1**). The correlation between the occurrence of state transitions and CEF raised the possibility that state transitions may act as a switch between LEF and CEF in *Chlamydomonas* [32]. This interpretation is however not compatible with recent studies which indicate that CEF is activated in the *stt7* mutant when the metabolic demand for ATP increases during the induction of the carbon concentrating mechanism when CO2 is limiting [33]. Also, analysis of the *stt7* and *ptox2* mutants, locked in state 1 and state 2, respectively, independent of the redox conditions led to similar conclusions [34]. The *ptox2* mutant is deficient in the plastid terminal oxidase which controls the redox state of the PQ pool in the dark [35]. Whereas the accumulation of reducing power and transition to state 2 correlated well with the en‐ hancement of CEF in the wild type, this was not the case for *ptox2*. In this mutant, CEF was not enhanced under aerobic conditions in the dark even though it is locked in state 2 with phosphorylated LHCII. Moreover, CEF enhancement and formation of the PSI-Cyt*b*6*f* supercomplex were still observed in the *stt7* mutant when the PQ pool was reduced. It can be concluded that both of these processes occur under reducing conditions with no correlation with state transitions and their associated LHCII reorganization and that it is the redox state of the photosynthetic electron transport rather than state transitions that control CEF [34].

subunits through an extensive signaling network in chloroplasts and between chloroplasts and

A distinctive feature of photosynthetic organisms is the presence of light-harvesting systems that funnel the absorbed light energy to the corresponding reaction centers and thereby considerably increase their absorption cross-section. Several regulatory mechanisms operate on these antenna systems for controlling the energy flux to the reaction centers. This is particularly important under changing environmental conditions when the photosynthetic apparatus needs to adapt quickly. Under limiting light, it optimizes its light absorption efficiency by adjusting the relative size of its antenna systems through the reversible allocation of a portion of LHCII between PSII and PSI, a process referred to as state transitions which occurs in algae, plants and cyanobacteria (for reviews see Refs. [17,18]). In contrast, when the absorbed light energy exceeds the capacity of the photosynthetic apparatus, it dissipates the excess excitation energy through nonphotochemical quenching (NPQ) as heat thereby

Because the antenna systems of PSII and PSI have a different pigment composition, their relative light absorption properties change when the light quality varies. This is especially important for aquatic algae because the penetration of light in water changes depending on its wavelength; in particular, red light is more absorbed than blue light. Another example is provided by photosynthetic organisms growing under a canopy where far red light is enriched. These changes in light quality can result in an unequal excitation of PSII and PSI and thereby perturb the redox poise of the plastoquinone pool. Over excitation of PSII relative to PSI leads to increased reduction of the plastoquinone pool and favors thereby docking of plastoquinol to the Qo site of the Cyt*b*6*f* complex [21,22]. This process leads to activation of the chloroplast protein kinase Stt7/STN7 and to the phosphorylation of several proteins from LHCII [23,24]. Although the direct phosphorylation of LHCII by the St7/STN7 kinase has not yet been demonstrated, this kinase is the best candidate for the LHCII kinase because it is firmly associated with the Cyt*b*6*f* complex, and in its absence, state transitions no longer occur [25]. Furthermore, it is widely conserved in land plants, mosses and algae. As a result of this phosphorylation, a part of the LHCII antenna is detached from PSII and moves and binds to PSI thereby rebalancing the light excitation of PSII and PSI and enhancing photosynthetic yield. This process is reversible as overexcitation of PSI leads to the inactivation of the kinase and to dephosphorylation of LHCII by the PPH1/TAP38 protein phosphatase and its return to PSI [26,27]. Thus two different states can be defined, state 1 and state 2 corresponding to the association of the mobile LHCII antenna to PSII and PSI, respectively. However, a strict causal link between LHCII phosphorylation and its migration from PSII to PSI has been questioned recently by the finding that some phosphorylated LHCII remains associated with PSII supercomplexes and that LHCII serves as antenna for both photosystems under most natural

nucleus which modulates nuclear and plastid gene expression.

**2. Adaptation to changes in light conditions**

avoiding photodamage (for reviews see Refs. [19,20]).

**2.1. State transitions**

26 Applied Photosynthesis - New Progress

State transitions do not occur under high light because the LHCII kinase is inactivated [36]. The current view is that inactivation of the kinase is mediated by the ferredoxin–thioredoxin system and that a disulfide bond in the kinase rather than in the substrate may be the target site of thioredoxin [37,38]. In this respect, the N-terminal region of the kinase contains indeed two Cys residues, which are conserved in all species examined [23,24]. Both of these Cys are essential for the kinase activity because changes of either Cys through site-directed mutagen‐ esis abolish the kinase activity [25]. It is noticeable that these Cys are located on the lumen side of the thylakoid membrane whereas the kinase catalytic domain is on the stromal side where the substrate sites of the LHCII proteins are located [23,25] (**Figure 2**). Although the conserved Cys residues in the lumen are on the opposite side of the stromal thioredoxin according to this model, one possibility is that thiol-reducing equivalents are transferred across the thylakoid membrane through the CcdA and Hcf164 proteins, which operate in this way during heme and Cyt*b*6*f* assembly [39,40]. Alternatively, transfer of thiol-reducing equivalents across the thylakoid membrane could also be mediated by Lto1 (lumen thiol oxidoreductase 1) which catalyzes the formation of disulfide bonds in the thylakoid lumen and is required for PSII assembly [41,42] (**Figure 2**). Its sulfhydryl oxidizing activity is linked to the reduction of phylloquinone, a redox component of PSI. It is not clear whether phylloquinone is involved in other electron transfer processes besides those in PSI. Although the two lumenal Cys are prime candidates for the redox control of the activity of the Stt7/STN7 kinase, high light treatment did not change the redox state of these Cys [43]. Another possibility is that high light affects the folding of the kinase in the thylakoid membrane, in particular through reactive oxygen species (ROS) generated by the high light treatment.

**Figure 2.** Regulation of the Stt7/STN7 kinase.

The Stt7/STN7 kinase is associated with the Cyt*b*6*f* complex. This kinase contains a transmem‐ brane domain connecting its N-terminus on the lumen side with two conserved Cys residues to the catalytic domain on the stromal side of the thylakoid membrane. The major substrates of this kinase are the LHCII proteins of the PSII antenna. The LHCII kinase is known to be inactivated by high light through the Fd/Trx system. This system could modulate the redox state of the two lumenal Cys through two proteins, CcdA and Hcf164, known to mediate the transfer of thiol-reducing equivalents across the thylakoid membrane. Another possibility is that this process is catalyzed by Lto1, the lumenal thiol oxidoreductase 1.

It is known that the activation of the kinase is intimately connected to the docking of plasto‐ quinone to the Qo site of the Cyt*b*6*f* complex [21,22] (**Figure 2**). Electron transfer from plasto‐ quinol to Cyt *f* is mediated by the Rieske protein and involves the movement of this protein from the proximal to distal position within the Cyt*b*6*f* complex [44,45]. Recent studies have revealed that the two conserved Cys residues of the Stt7/STN7 kinase form an intramolecular disulfide bridge which appears to be essential for kinase activity [43]. However, no change in the redox state of these Cys could be detected during state transitions. It is only under prolonged anaerobic conditions that this disulfide bridge was reduced but at a significantly slower pace than transition from state 1 to state 2 which occurs under anaerobiosis in *Chlamydomonas* [46]. In wild-type *Arabidopsis* plants, the STN7 kinase is only observed as a monomer under both state 1 and state 2 conditions although the dimer could be detected in plants overexpressing STN7 or in mutants with changes in either of the two luminal Cys of STN7 [47]. However, these results do not exclude the possibility of rapid and transient changes in the redox state of these two Cys. In fact, such changes were proposed to occur to accom‐ modate all the known features of the Stt7/STN7 kinase [43]. A transient change from an intrato intermolecular disulfide bond may occur which would activate the kinase and be coupled to the movement of the Rieske protein during electron transfer from plastoquinol to Cyt *f*. Moreover, it is interesting to note that the interaction site between the kinase and the Cyt*b*6*f* complex has been located close to the flexible glycine-rich hinge connecting the membrane anchor to the large head of the Rieske protein in the lumen [43]. A single chlorophyll *a* molecule with its phytyl tail close to the Qo site exists in Cyt*b*6*f* [48]. It is also possible that the kinase senses PQH2 binding to the Qo site through the single chlorophyll *a* molecule of the Cyt*b*6*f* complex whose phytyl tail is close to the Qo site [48,49]. It was proposed that this molecule may play a role in the activation of the Stt7/STN7 kinase based on site-directed mutagenesis of the chlorophyll *a* binding site [50]. The activation of the kinase would be triggered through the transient formation of a STN7 dimer with two intermolecular disulfide bridges which would transduce the signal to the catalytic domain on the stromal side of the thylakoid membrane [43].

in other electron transfer processes besides those in PSI. Although the two lumenal Cys are prime candidates for the redox control of the activity of the Stt7/STN7 kinase, high light treatment did not change the redox state of these Cys [43]. Another possibility is that high light affects the folding of the kinase in the thylakoid membrane, in particular through reactive

The Stt7/STN7 kinase is associated with the Cyt*b*6*f* complex. This kinase contains a transmem‐ brane domain connecting its N-terminus on the lumen side with two conserved Cys residues to the catalytic domain on the stromal side of the thylakoid membrane. The major substrates of this kinase are the LHCII proteins of the PSII antenna. The LHCII kinase is known to be inactivated by high light through the Fd/Trx system. This system could modulate the redox state of the two lumenal Cys through two proteins, CcdA and Hcf164, known to mediate the transfer of thiol-reducing equivalents across the thylakoid membrane. Another possibility is

It is known that the activation of the kinase is intimately connected to the docking of plasto‐ quinone to the Qo site of the Cyt*b*6*f* complex [21,22] (**Figure 2**). Electron transfer from plasto‐ quinol to Cyt *f* is mediated by the Rieske protein and involves the movement of this protein from the proximal to distal position within the Cyt*b*6*f* complex [44,45]. Recent studies have revealed that the two conserved Cys residues of the Stt7/STN7 kinase form an intramolecular disulfide bridge which appears to be essential for kinase activity [43]. However, no change in the redox state of these Cys could be detected during state transitions. It is only under prolonged anaerobic conditions that this disulfide bridge was reduced but at a significantly slower pace than transition from state 1 to state 2 which occurs under anaerobiosis in *Chlamydomonas* [46]. In wild-type *Arabidopsis* plants, the STN7 kinase is only observed as a monomer under both state 1 and state 2 conditions although the dimer could be detected in plants overexpressing STN7 or in mutants with changes in either of the two luminal Cys of

that this process is catalyzed by Lto1, the lumenal thiol oxidoreductase 1.

oxygen species (ROS) generated by the high light treatment.

**Figure 2.** Regulation of the Stt7/STN7 kinase.

28 Applied Photosynthesis - New Progress

Another proposal for the mechanism of activation of the Stt7/STN7 kinase is that hydrogen peroxide may be involved by oxidizing the luminal C1 and C2 to form intra- and/or intermo‐ lecular disulfide bridges. It is based on the observation that singlet oxygen generated by PSII can oxidize plastoquinol with concomitant production of hydrogen peroxide in the thylakoid membranes [51]. However, this proposal is difficult to reconcile with the observation that these Cys exist mostly in the oxidized form and the conversion from intra- to intermolecular disulfide bridges appears to be only transient [43].

State transitions involve remodeling of the antenna system of PSII within the thylakoid membranes. This poses a challenging problem especially considering the fact that among biological membranes, the thylakoid membrane is very crowded with 70% of the surface area of grana membranes occupied by proteins and 30% by lipids [11]. Light-induced architectural changes in the folding of the thylakoid membrane are induced at least partly by changes in phosphorylation of thylakoid proteins catalyzed by the protein kinases Stt7/STN7 and Stl1/ STN8, which most likely facilitate mobility of proteins in these membranes [52,53]. These two kinases appear to play an important role in chloroplast signaling in response to changing environmental conditions (**Figure 3**). Light irradiance, ambient CO2 level and the cellular ATP/ ADP ratio modulate the redox state of the plastoquinone pool of the electron transport chain which is sensed by the Stt7/STN7 kinase. Together with the Stl1/STN8 kinase and the two corresponding protein phosphatases PPH1/TAP38 and PBCP, Stt7/STN7 forms a central quartet which orchestrates the phosphorylation of the LHCII and the PSII core proteins (**Figure 3**). PTK is another chloroplast Ser/Thr kinase of the casein kinase II family which is associated with the plastid RNA polymerase and acts as a global regulator of chloroplast transcription [54,55]. The CSK kinase shares structural features with cyanobacterial sensor histidine kinases and is conserved in all major plant and algal lineages except *Chlamydomonas reinhardtii* [56]. Upon oxidation of the PQ pool, autophosphorylation of CSK occurs, an event which correlates with phosphorylation of the chloroplast σ factor Sig1 and the decrease of *psaAB* gene expression. Furthermore, CSK interacts with PTK and SIG1 in yeast two-hybrid assays. Based on these results, it was proposed that CSK is regulated by the redox state of the PQ pool through the STN7 kinase (**Figure 3**) [56]. However, how the different kinases are linked within this chloroplast signaling network shown in **Figure 3** is still unclear.

**Figure 3.** Chloroplast signaling.

The redox state of the PQ pool is modulated by the light irradiance, ATP/ADP ratio and ambient CO2 level. The protein kinases Stl1/STN8, Stt7/STN7, CSK, PTK and TAK1 [57] are shown with their targets indicated by arrows. *Broken arrows* indicate putative targets. *LTR*longterm response involving retrograde signaling is mediated through Stt7/STN7. Reproduced from Ref. 58 with permission.

#### **2.2. Non photochemical quenching**

While state transitions are mainly involved in low light responses through an extensive reorganization of the antenna systems, other mechanisms for the regulation of light-harvesting operate when oxygenic photosynthetic organisms are suddenly exposed to large and sudden changes in light intensity in their natural habitat. In the case of aquatic algae, even moderate water mixing can bring algae from full darkness to high light within minutes [59,60]. Under these conditions, increased electron flow along the electron transport chain generates a large proton gradient. The resulting acidification of the thylakoid lumen leads to the de-excitation of singlet excited light-harvesting pigments and is measured as non-photochemical quenching (NPQ of chlorophyll fluorescence). NPQ comprises several components; the major one is the high energy state quenching qE, which leads to the harmless heat dissipation of the absorbed excess light energy [20,61]. The other components which also contribute to fluorescence quenching are the photoinhibitory quenching qI and state transitions qT although qT is not associated with thermal dissipation of excitation energy. The qE mechanism occurs in all major algal taxa and land plants. However, the underlying molecular mechanisms of heat dissipation of excess excitation energy differ. The qE process involves both the xanthophyll cycle and the PsbS protein in plants. Another protein, LhcsR, has been shown to mediate qE in algae [19,62].

*psaAB* gene expression. Furthermore, CSK interacts with PTK and SIG1 in yeast two-hybrid assays. Based on these results, it was proposed that CSK is regulated by the redox state of the PQ pool through the STN7 kinase (**Figure 3**) [56]. However, how the different kinases are linked

The redox state of the PQ pool is modulated by the light irradiance, ATP/ADP ratio and ambient CO2 level. The protein kinases Stl1/STN8, Stt7/STN7, CSK, PTK and TAK1 [57] are shown with their targets indicated by arrows. *Broken arrows* indicate putative targets. *LTR*longterm response involving retrograde signaling is mediated through Stt7/STN7. Reproduced

While state transitions are mainly involved in low light responses through an extensive reorganization of the antenna systems, other mechanisms for the regulation of light-harvesting operate when oxygenic photosynthetic organisms are suddenly exposed to large and sudden changes in light intensity in their natural habitat. In the case of aquatic algae, even moderate water mixing can bring algae from full darkness to high light within minutes [59,60]. Under these conditions, increased electron flow along the electron transport chain generates a large proton gradient. The resulting acidification of the thylakoid lumen leads to the de-excitation of singlet excited light-harvesting pigments and is measured as non-photochemical quenching (NPQ of chlorophyll fluorescence). NPQ comprises several components; the major one is the high energy state quenching qE, which leads to the harmless heat dissipation of the absorbed excess light energy [20,61]. The other components which also contribute to fluorescence quenching are the photoinhibitory quenching qI and state transitions qT although qT is not associated with thermal dissipation of excitation energy. The qE mechanism occurs in all major

within this chloroplast signaling network shown in **Figure 3** is still unclear.

**Figure 3.** Chloroplast signaling.

30 Applied Photosynthesis - New Progress

from Ref. 58 with permission.

**2.2. Non photochemical quenching**

The proton gradient acts as a sensor of the state of the photosynthetic electron transport chain. The magnitude of this gradient is low under low light illumination and high under illumina‐ tion with high light especially when it exceeds the capacity of the photosynthetic apparatus. The resulting acidification of the thylakoid lumen activates the xanthophyll cycle in which violaxanthin is de-epoxidized to zeaxanthin, a reaction catalyzed by violaxanthin de-epoxidase (VDE) which has an acidic pH optimum [63]. The reverse reaction is catalyzed by zeaxanthin epoxidase with a broad pH optimum and which in contrast to VDE is active both in the dark and in the light. Because the turnover of this enzyme is considerably lower than that of VDE, zeaxanthin accumulates rapidly during high light illumination. The zeaxanthin-dependent NPQ depends greatly on the grana structure as unstacking of the membranes abolishes qE. It was proposed that the organization of LHCII in an aggregated state within the stacked grana region is essential for efficient qE [64]. Both high proton concentration in the lumen and accumulation of zeaxanthin promote not only aggregation of LHCII but also that of the minor PSII antenna proteins CP29, CP26 and CP24 [65,66]. In plants, qE occurs in the LHC proteins at multiple sites of the antenna system [67]. These proteins have the ability to switch from an efficient light-harvesting mode to a light energy dissipating state [68]. Several mechanisms have been proposed including excitonic coupling, charge transfer and energy transfer between carotenoids and chlorophylls as well as chlorophyll–chlorophyll charge transfer states (for review see Ref. 20).

Another important player involved in NPQ is PsbS, a four-helix member of the LHC protein family [69]. However, this protein does not appear to bind pigments although a chlorophyll molecule was detected at the dimer interface in the PsbS crystals [70]. This protein appears to act as a sensor of the lumen pH most likely through protonation of its acidic lumen residues which in turn induces a rearrangement of the light-harvesting system required for induction of NPQ [71–73]. In this sense, PsbS would act as an antenna organizer, a view which is further supported by the fact that it is mobile in the thylakoid membrane [74], and it is able to associate with both the PSII core complex and LHCII [75]. Moreover, qE can be switched on without the PsbS protein if the lumen pH is very low [76]. It thus appears that protonated PsbS allows for a fast and efficient rearrangement of the PSII antenna which is still possible in its absence but requires a longer time.

In *Chlamydomonas reinhardtii* and *Phaeodactylum tricornutum*, two representatives of green algae and diatoms, respectively, qE is mediated by Lhcsr, another three helix member of the LHC protein family [62]. In high light, most Lhcsr genes are up-regulated in contrast to the lightharvesting genes which are down-regulated. Recent studies reveal that Lhcsr binds chloro‐ phylls and xanthophylls *in vitro* and that it has a basal quenching activity associated with chlorophyll–xanthophyll charge transfer [77]. Its chlorophyll fluorescence lifetime is remark‐ ably short and even shorter at low pH suggesting that this protein has some quenching activity even in low light which is enhanced at low pH. It was proposed that these properties could explain the low expression of Lhcsr under low light when constitutive quenching would be wasteful [20]. The *Chlamydomonas* Lhcsr is bound to PSII where it may interact with the LHC proteins, especially Lhcbm1 which is known to be involved in thermal dissipation [78–80]. Interestingly, as several other LHC proteins, Lhcsr is phosphorylated by the Stt7 kinase and moves from PSII to PSI during a state 1 to state 2 transition [78]. This observation is particularly interesting with regard to chlorophyll fluorescence lifetime measurements which reveal two different kinetic components suggesting the existence of two underlying mechanisms [81]. It is noteworthy that although PsbS is also present in green algae, there is no evidence that it is involved in qE in these organisms. This is in contrast to the moss *Physcomitrella patens* that has both Lhcsr- and PsbS-dependent NPQ which operate independently and additively [82,83]. The maintenance of these two mechanisms in mosses may be linked to a greater need for inducible NPQ in these organisms [84].

NPQ has also been investigated in diatoms, a ubiquitous group of unicellular marine algae which make an important contribution to the global carbon assimilation [85]. Diatoms acquired their chloroplast through secondary endosymbiosis from a red algal ancestor [86]. In these organisms, similar to plants and green algae, qE relies on three interacting components, the light-induced proton gradient across the thylakoid membrane, the conversion of the xantho‐ phyll diadinoxanthin (Dd) to diatoxanthin (Dt) catalyzed by the enzyme Dd-de-epoxidase which depends on a transthylakoid proton gradient and the Lhcx antenna proteins (for review see Ref. 87). Among these, Lhcx1 appears to play a major role in qE as changes in its level are directly related to the quenching of light energy [88]. Lhcx1 also plays an important general role in light responses in diatoms as it accumulates in different amounts in ecotypes originating from different latitudes. In contrast to land plants, the proton gradient is unable to induce NPQ on its own in diatoms. It is only required to activate the de-epoxidation of Dd. The qE process represents an important photoprotective mechanism and involves a reorganization of the antenna complexes of diatoms [87]. However, the quenching sites within the antenna systems of these organisms have not yet been precisely determined.

Another original feature of diatoms is the way they adjust the ATP/NADPH ratio which is important for proper carbon assimilation by the CBC and for optimal growth. In plants and green algae, this ratio is mainly set by the relative contributions of LEF and CEF and by the water-to-water cycles [89], whereas in diatoms this ratio relies principally on energetic exchanges between plastids and mitochondria [90]. These bidirectional organellar interactions involve the rerouting of reducing power generated by photosynthesis in the plastids to mitochondria and the import of ATP produced in the mitochondria to the plastids.

#### **2.3. PSII repair cycle**

Water splitting by PSII is one of the strongest oxidizing reactions which occurs in living organisms. As a result, photodamage to PSII is unavoidable. A remarkable feature of this system is that it is efficiently repaired [91]. PSII exists as a dimer in which each monomer consists of 28 subunits generally associated with two LHCII trimers in a supercomplex [8]. The PSII core consists of the two reaction center polypeptides D1 and D2 which form a central heterodimer which acts as ligand for the chlorophyll dimer P680 and the other redox compo‐ nents including the quinones QA and QB, the primary and secondary electron acceptors. Among all PSII subunits, D1 is the major target of photodamage and needs to be specifically replaced. This process, called PSII repair cycle, involves the partial disassembly of the PSII supercom‐ plex, the removal and degradation of the damaged D1 protein, its replacement by a newly synthesized copy and the reassembly of the PSII complex (**Figure 4**) [92]. An important feature of this repair cycle is that it is compartmentalized within the crowded thylakoid membrane [93]. Whereas damage of D1 occurs in the stacked grana region where most of PSII is located, the replacement of this protein takes place in the stroma lamellae. Although the exact role of phosphorylation is not fully understood, the current view is that the PSII repair cycle starts with phosphorylation of the PSII core subunits D1, D2, CP43 and PsbH mediated by the STN8 kinase [94,95] which leads to the disassembly of the PSII-LHCII supercomplex thereby allowing PSII to move to the grana margins and stroma lamellae [96,97]. Dephosphorylation by the PSII core phosphatase Pbcp [53] and by other unknown phosphatases is followed by the degradation of D1 by the FtsH and Deg proteases and a newly synthesized D1 is cotrans‐ lationally inserted into the PSII complex [98]. Finally, the reassembled PSII complex moves back to the grana and reforms a supercomplex with LHCII. To make this cycle efficient, it is essential that the enzymes involved are confined to distinct thylakoid membrane subcom‐ partments. Thus, protein degradation occurs on the grana margins and protein synthesis on the stroma lamellae. Additionally, partial conversion of grana stacks to grana margins allows the proteases to access PSII [93].

**Figure 4.** PSII repair cycle.

wasteful [20]. The *Chlamydomonas* Lhcsr is bound to PSII where it may interact with the LHC proteins, especially Lhcbm1 which is known to be involved in thermal dissipation [78–80]. Interestingly, as several other LHC proteins, Lhcsr is phosphorylated by the Stt7 kinase and moves from PSII to PSI during a state 1 to state 2 transition [78]. This observation is particularly interesting with regard to chlorophyll fluorescence lifetime measurements which reveal two different kinetic components suggesting the existence of two underlying mechanisms [81]. It is noteworthy that although PsbS is also present in green algae, there is no evidence that it is involved in qE in these organisms. This is in contrast to the moss *Physcomitrella patens* that has both Lhcsr- and PsbS-dependent NPQ which operate independently and additively [82,83]. The maintenance of these two mechanisms in mosses may be linked to a greater need for

NPQ has also been investigated in diatoms, a ubiquitous group of unicellular marine algae which make an important contribution to the global carbon assimilation [85]. Diatoms acquired their chloroplast through secondary endosymbiosis from a red algal ancestor [86]. In these organisms, similar to plants and green algae, qE relies on three interacting components, the light-induced proton gradient across the thylakoid membrane, the conversion of the xantho‐ phyll diadinoxanthin (Dd) to diatoxanthin (Dt) catalyzed by the enzyme Dd-de-epoxidase which depends on a transthylakoid proton gradient and the Lhcx antenna proteins (for review see Ref. 87). Among these, Lhcx1 appears to play a major role in qE as changes in its level are directly related to the quenching of light energy [88]. Lhcx1 also plays an important general role in light responses in diatoms as it accumulates in different amounts in ecotypes originating from different latitudes. In contrast to land plants, the proton gradient is unable to induce NPQ on its own in diatoms. It is only required to activate the de-epoxidation of Dd. The qE process represents an important photoprotective mechanism and involves a reorganization of the antenna complexes of diatoms [87]. However, the quenching sites within the antenna systems

Another original feature of diatoms is the way they adjust the ATP/NADPH ratio which is important for proper carbon assimilation by the CBC and for optimal growth. In plants and green algae, this ratio is mainly set by the relative contributions of LEF and CEF and by the water-to-water cycles [89], whereas in diatoms this ratio relies principally on energetic exchanges between plastids and mitochondria [90]. These bidirectional organellar interactions involve the rerouting of reducing power generated by photosynthesis in the plastids to

Water splitting by PSII is one of the strongest oxidizing reactions which occurs in living organisms. As a result, photodamage to PSII is unavoidable. A remarkable feature of this system is that it is efficiently repaired [91]. PSII exists as a dimer in which each monomer consists of 28 subunits generally associated with two LHCII trimers in a supercomplex [8]. The PSII core consists of the two reaction center polypeptides D1 and D2 which form a central heterodimer which acts as ligand for the chlorophyll dimer P680 and the other redox compo‐ nents including the quinones QA and QB, the primary and secondary electron acceptors. Among

mitochondria and the import of ATP produced in the mitochondria to the plastids.

inducible NPQ in these organisms [84].

32 Applied Photosynthesis - New Progress

**2.3. PSII repair cycle**

of these organisms have not yet been precisely determined.

High light illumination leads to photooxidative damage of the PSII reaction center, especially the D1 protein. The PSII core proteins are phosphorylated and the damaged complex migrates from the grana (G) to the stromal lamellae (SL). The D1 protein is degraded by the FtsH and Deg proteases and upon its removal from the PSII reaction center a newly synthesized D1 protein is inserted cotranslationally into the complex which moves back to the grana and thereby completes the repair cycle. Reproduced from Ref. 5 with permission.

D1 degradation is significantly retarded in the *stn8* and *stn7 stn8* mutants of *Arabidopsis* in which the thylakoid membrane architecture is affected [52]. In the absence of STN8, grana diameter is increased and there are fewer grana stacks. This observation is particularly intriguing as it suggests that PSII core phosphorylation is important for maintaining grana size, a parameter which is highly conserved in land plants and algae [11]. Loss of STN8 also affects partitioning of FtsH between grana and stromal membranes and limits its access to the grana, and migration of D1 from the grana to the stroma lamellae is slowed down during the PSII repair cycle [52]. These observations are thus compatible with the view that PSII core phosphorylation has a strong impact on thylakoid membrane folding and architecture mediated most likely by electrostatic repulsion between membrane layers as proposed earlier [99].
