Generation and physiological characterization of genome-edited Nicotiana benthamiana plants containing zeaxanthin as the only leaf xanthophyll.

MAIN CONCLUSION: Simultaneous genome editing of the two homeologous LCYe and ZEP genes of Nicotiana benthamiana results in plants in which all xanthophylls are replaced by zeaxanthin. Plant carotenoids act both as photoreceptors and photoprotectants in photosynthesis and as precursors of apocarotenoids, which include signaling molecules such as abscisic acid (ABA). As dietary components, the xanthophylls lutein and zeaxanthin have photoprotective functions in the human macula. We developed transient and stable combinatorial genome editing methods, followed by direct LC-MS screening for zeaxanthin accumulation, for the simultaneous genome editing of the two homeologous Lycopene Epsilon Cyclase (LCYe) and the two Zeaxanthin Epoxidase (ZEP) genes present in the allopolyploid Nicotiana benthamiana genome. Editing of the four genes resulted in plants in which all leaf xanthophylls were substituted by zeaxanthin, but with different ABA levels and growth habits, depending on the severity of the ZEP1 mutation. In high-zeaxanthin lines, the abundance of the major photosystem II antenna LHCII was reduced with respect to wild-type plants and the LHCII trimeric state became unstable upon thylakoid solubilization. Consistent with the depletion in LHCII, edited plants underwent a compensatory increase in PSII/PSI ratios and a loss of the large-size PSII supercomplexes, while the level of PSI-LHCI supercomplex was unaffected. Reduced activity of the photoprotective mechanism NPQ was shown in high-zeaxanthin plants, while PSII photoinhibition was similar for all genotypes upon exposure to excess light, consistent with the antioxidant and photoprotective role of zeaxanthin in vivo.

A paler shade of green: engineering cellular chlorophyll content to enhance photosynthesis in crowded environments.

In natural ecosystems, plants compete for space, nutrients and light. The optically dense canopies limit the penetration of photosynthetically active radiation and light often becomes a growth-limiting factor for the understory. The reduced availability of photons in the lower leaf layers is also a major constraint for yield potential in canopies of crop monocultures. Traditionally, crop breeding has selected traits related to plant architecture and nutrient assimilation rather than light use efficiency. Leaf optical density is primarily determined by tissue morphology and by the foliar concentration of photosynthetic pigments (chlorophylls and carotenoids). Most pigment molecules are bound to light-harvesting antenna proteins in the chloroplast thylakoid membranes, where they serve photon capture and excitation energy transfer toward reaction centers of photosystems. Engineering the abundance and composition of antenna proteins has been suggested as a strategy to improve light distribution within canopies and reduce the gap between theoretical and field productivity. Since the assembly of the photosynthetic antennas relies on several coordinated biological processes, many genetic targets are available for modulating cellular chlorophyll levels. In this review, we outline the rationale behind the advantages of developing pale green phenotypes and describe possible approaches toward engineering light-harvesting systems.

The role of light-harvesting complex I in excitation energy transfer from LHCII to photosystem I in Arabidopsis.

Photosynthesis powers nearly all life on Earth. Light absorbed by photosystems drives the conversion of water and carbon dioxide into sugars. In plants, photosystem I (PSI) and photosystem II (PSII) work in series to drive the electron transport from water to NADP+. As both photosystems largely work in series, a balanced excitation pressure is required for optimal photosynthetic performance. Both photosystems are composed of a core and light-harvesting complexes (LHCI) for PSI and LHCII for PSII. When the light conditions favor the excitation of one photosystem over the other, a mobile pool of trimeric LHCII moves between both photosystems thus tuning their antenna cross-section in a process called state transitions. When PSII is overexcited multiple LHCIIs can associate with PSI. A trimeric LHCII binds to PSI at the PsaH/L/O site to form a well-characterized PSI-LHCI-LHCII supercomplex. The binding site(s) of the "additional" LHCII is still unclear, although a mediating role for LHCI has been proposed. In this work, we measured the PSI antenna size and trapping kinetics of photosynthetic membranes from Arabidopsis (Arabidopsis thaliana) plants. Membranes from wild-type (WT) plants were compared to those of the ΔLhca mutant that completely lacks the LHCI antenna. The results showed that "additional" LHCII complexes can transfer energy directly to the PSI core in the absence of LHCI. However, the transfer is about two times faster and therefore more efficient, when LHCI is present. This suggests LHCI mediates excitation energy transfer from loosely bound LHCII to PSI in WT plants.

A kaleidoscope of photosynthetic antenna proteins and their emerging roles.

Photosynthetic light-harvesting antennae are pigment-binding proteins that perform one of the most fundamental tasks on Earth, capturing light and transferring energy that enables life in our biosphere. Adaptation to different light environments led to the evolution of an astonishing diversity of light-harvesting systems. At the same time, several strategies have been developed to optimize the light energy input into photosynthetic membranes in response to fluctuating conditions. The basic feature of these prompt responses is the dynamic nature of antenna complexes, whose function readily adapts to the light available. High-resolution microscopy and spectroscopic studies on membrane dynamics demonstrate the crosstalk between antennae and other thylakoid membrane components. With the increased understanding of light-harvesting mechanisms and their regulation, efforts are focusing on the development of sustainable processes for effective conversion of sunlight into functional bio-products. The major challenge in this approach lies in the application of fundamental discoveries in light-harvesting systems for the improvement of plant or algal photosynthesis. Here, we underline some of the latest fundamental discoveries on the molecular mechanisms and regulation of light harvesting that can potentially be exploited for the optimization of photosynthesis.

Light-harvesting complex stress-related proteins play crucial roles in the acclimation of Physcomitrella patens under fluctuating light conditions.

Photosynthetic organisms have evolved photoprotective mechanisms to acclimate to light intensity fluctuations in their natural growth environments. Photosystem (PS) II subunit S (PsbS) and light-harvesting complex (LHC) stress-related proteins (LhcSR) are essential for triggering photoprotection in vascular plants and green algae, respectively. The activity of both proteins is strongly enhanced in the moss Physcomitrella patens under high-light conditions. However, their role in regulating photosynthesis acclimation in P. patens under fluctuating light (FL) conditions is still unknown. Here, we compare the responses of wild-type (WT) P. patens and mutants lacking PsbS (psbs KO) or LhcSR1 and 2 (lhcsr KO) to FL conditions in which the low-light phases were periodically interrupted with high-light pulses. lhcsr KO mutant showed a strong reduction in growth with respect to WT and psbs KO under FL conditions. The lack of LhcSR not only decreased the level of non-photochemical quenching, resulting in an over-reduced plastoquinone pool, but also significantly increased the PSI acceptor limitation values with respect to WT and psbs KO under FL conditions. Moreover, in lhcsr KO mutant, the abundance of PSI core and PSI-LHCI complex decreased greatly under FL conditions compared with the WT and psbs KO. We proposed that LhcSR in P. patens play a crucial role in moss acclimation to dynamic light changes.

Molecular mechanisms of light harvesting in the minor antenna CP29 in near-native membrane lipidic environment.

CP29, a chlorophyll a/b-xanthophyll binding protein, bridges energy transfer between the major LHCII antenna complexes and photosystem II reaction centers. It hosts one of the two identified quenching sites, making it crucial for regulated photoprotection mechanisms. Until now, the photophysics of CP29 has been studied on the purified protein in detergent solutions since spectrally overlapping signals affect in vivo measurements. However, the protein in detergent assumes non-native conformations compared to its physiological state in the thylakoid membrane. Here, we report a detailed photophysical study on CP29 inserted in discoidal lipid bilayers, known as nanodiscs, which mimic the native membrane environment. Using picosecond time-resolved fluorescence and femtosecond transient absorption (TA), we observed shortening of the Chl fluorescence lifetime with a decrease of the carotenoid triplet formation yield for CP29 in nanodiscs as compared to the protein in detergent. Global analysis of TA data suggests a 1Chl* quenching mechanism dependent on excitation energy transfer to a carotenoid dark state, likely the proposed S*, which is believed to be formed due to a carotenoid conformational change affecting the S1 state. We suggest that the accessibility of the S* state in different local environments plays a key role in determining the quenching of Chl excited states. In vivo, non-photochemical quenching is activated by de-epoxidation of violaxanthin into zeaxanthin. CP29-zeaxanthin in nanodiscs further shortens the Chl lifetime, which underlines the critical role of zeaxanthin in modulating photoprotection activity.

A new function for the xanthophyll zeaxanthin: glueing chlorophyll biosynthesis to thylakoid protein assembly.

Xanthophylls are coloured isoprenoid metabolites synthesized in many organisms with a variety of functions from the attraction of animals for impollination to absorption of light energy for photosynthesis to photoprotection against photooxidative stress. The finding by Proctor and co-workers makes a new addition to the last type of functions by showing that zeaxanthin is instrumental in coordinating chlorophyll biosynthesis with the insertion of pigment-binding proteins into the photosynthetic membrane by glueing the protein components catalyzing these functions into a supercomplex and regulating its activity.

LHCSR3 is a nonphotochemical quencher of both photosystems in Chlamydomonas reinhardtii.

Photosynthetic organisms prevent oxidative stress from light energy absorbed in excess through several photoprotective mechanisms. A major component is thermal dissipation of chlorophyll singlet excited states and is called nonphotochemical quenching (NPQ). NPQ is catalyzed in green algae by protein subunits called LHCSRs (Light Harvesting Complex Stress Related), homologous to the Light Harvesting Complexes (LHC), constituting the antenna system of both photosystem I (PSI) and PSII. We investigated the role of LHCSR1 and LHCSR3 in NPQ activation to verify whether these proteins are involved in thermal dissipation of PSI excitation energy, in addition to their well-known effect on PSII. To this aim, we measured the fluorescence emitted at 77 K by whole cells in a quenched or unquenched state, using green fluorescence protein as the internal standard. We show that NPQ activation by high light treatment in Chlamydomonas reinhardtii leads to energy quenching in both PSI and PSII antenna systems. By analyzing quenching properties of mutants affected on the expression of LHCSR1 or LHCSR3 gene products and/or state 1-state 2 transitions or zeaxanthin accumulation, namely, npq4, stt7, stt7 npq4, npq4 lhcsr1, lhcsr3-complemented npq4 lhcsr1 and npq1, we showed that PSI undergoes NPQ through quenching of the associated LHCII antenna. This quenching event is fast-reversible on switching the light off, is mainly related to LHCSR3 activity, and is dependent on thylakoid luminal pH. Moreover, PSI quenching could also be observed in the absence of zeaxanthin or STT7 kinase activity.

Microsecond and millisecond dynamics in the photosynthetic protein LHCSR1 observed by single-molecule correlation spectroscopy.

Biological systems are subjected to continuous environmental fluctuations, and therefore, flexibility in the structure and function of their protein building blocks is essential for survival. Protein dynamics are often local conformational changes, which allows multiple dynamical processes to occur simultaneously and rapidly in individual proteins. Experiments often average over these dynamics and their multiplicity, preventing identification of the molecular origin and impact on biological function. Green plants survive under high light by quenching excess energy, and Light-Harvesting Complex Stress Related 1 (LHCSR1) is the protein responsible for quenching in moss. Here, we expand an analysis of the correlation function of the fluorescence lifetime by improving the estimation of the lifetime states and by developing a multicomponent model correlation function, and we apply this analysis at the single-molecule level. Through these advances, we resolve previously hidden rapid dynamics, including multiple parallel processes. By applying this technique to LHCSR1, we identify and quantitate parallel dynamics on hundreds of microseconds and tens of milliseconds timescales, likely at two quenching sites within the protein. These sites are individually controlled in response to fluctuations in sunlight, which provides robust regulation of the light-harvesting machinery. Considering our results in combination with previous structural, spectroscopic, and computational data, we propose specific pigments that serve as the quenching sites. These findings, therefore, provide a mechanistic basis for quenching, illustrating the ability of this method to uncover protein function.

Violaxanthin and Zeaxanthin May Replace Lutein at the L1 Site of LHCII, Conserving the Interactions with Surrounding Chlorophylls and the Capability of Triplet-Triplet Energy Transfer.

Carotenoids represent the first line of defence of photosystems against singlet oxygen (1O2) toxicity, because of their capacity to quench the chlorophyll triplet state (3Chl) through a physical mechanism based on the transfer of triplet excitation (triplet-triplet energy transfer, TTET). In previous works, we showed that the antenna LHCII is characterised by a robust photoprotective mechanism, able to adapt to the removal of individual chlorophylls while maintaining a remarkable capacity for 3Chl quenching. In this work, we investigated the effects on this quenching induced in LHCII by the replacement of the lutein bound at the L1 site with violaxanthin and zeaxanthin. We studied LHCII isolated from the Arabidopsis thaliana mutants lut2-in which lutein is replaced by violaxanthin-and lut2 npq2, in which all xanthophylls are replaced constitutively by zeaxanthin. We characterised the photophysics of these systems via optically detected magnetic resonance (ODMR) and time-resolved electron paramagnetic resonance (TR-EPR). We concluded that, in LHCII, lutein-binding sites have conserved characteristics, and ensure efficient TTET regardless of the identity of the carotenoid accommodated.

Protein-Protein Interactions Induce pH-Dependent and Zeaxanthin-Independent Photoprotection in the Plant Light-Harvesting Complex, LHCII.

Plants use energy from the sun yet also require protection against the generation of deleterious photoproducts from excess energy. Photoprotection in green plants, known as nonphotochemical quenching (NPQ), involves thermal dissipation of energy and is activated by a series of interrelated factors: a pH drop in the lumen, accumulation of the carotenoid zeaxanthin (Zea), and formation of arrays of pigment-containing antenna complexes. However, understanding their individual contributions and their interactions has been challenging, particularly for the antenna arrays, which are difficult to manipulate in vitro. Here, we achieved systematic and discrete control over the array size for the principal antenna complex, light-harvesting complex II, using near-native in vitro membranes called nanodiscs. Each of the factors had a distinct influence on the level of dissipation, which was characterized by measurements of fluorescence quenching and ultrafast chlorophyll-to-carotenoid energy transfer. First, an increase in array size led to a corresponding increase in dissipation; the dramatic changes in the chlorophyll dynamics suggested that this is due to an allosteric conformational change of the protein. Second, a pH drop increased dissipation but exclusively in the presence of protein-protein interactions. Third, no Zea dependence was identified which suggested that Zea regulates a distinct aspect of NPQ. Collectively, these results indicate that each factor provides a separate type of control knob for photoprotection, which likely enables a flexible and tunable response to solar fluctuations.

A microalgal-based preparation with synergistic cellulolytic and detoxifying action towards chemical-treated lignocellulose.

High-temperature bioconversion of lignocellulose into fermentable sugars has drawn attention for efficient production of renewable chemicals and biofuels, because competing microbial activities are inhibited at elevated temperatures and thermostable cell wall degrading enzymes are superior to mesophilic enzymes. Here, we report on the development of a platform to produce four different thermostable cell wall degrading enzymes in the chloroplast of Chlamydomonas reinhardtii. The enzyme blend was composed of the cellobiohydrolase CBM3GH5 from C. saccharolyticus, the ß-glucosidase celB from P. furiosus, the endoglucanase B and the endoxylanase XynA from T. neapolitana. In addition, transplastomic microalgae were engineered for the expression of phosphite dehydrogenase D from Pseudomonas stutzeri, allowing for growth in non-axenic media by selective phosphite nutrition. The cellulolytic blend composed of the glycoside hydrolase (GH) domain GH12/GH5/GH1 allowed the conversion of alkaline-treated lignocellulose into glucose with efficiencies ranging from 14% to 17% upon 48h of reaction and an enzyme loading of 0.05% (w/w). Hydrolysates from treated cellulosic materials with extracts of transgenic microalgae boosted both the biogas production by methanogenic bacteria and the mixotrophic growth of the oleaginous microalga Chlorella vulgaris. Notably, microalgal treatment suppressed the detrimental effect of inhibitory by-products released from the alkaline treatment of biomass, thus allowing for efficient assimilation of lignocellulose-derived sugars by C. vulgaris under mixotrophic growth.

Optimized Cas9 expression systems for highly efficient Arabidopsis genome editing facilitate isolation of complex alleles in a single generation.

Genetic resources for the model plant Arabidopsis comprise mutant lines defective in almost any single gene in reference accession Columbia. However, gene redundancy and/or close linkage often render it extremely laborious or even impossible to isolate a desired line lacking a specific function or set of genes from segregating populations. Therefore, we here evaluated strategies and efficiencies for the inactivation of multiple genes by Cas9-based nucleases and multiplexing. In first attempts, we succeeded in isolating a mutant line carrying a 70 kb deletion, which occurred at a frequency of ~ 1.6% in the T2 generation, through PCR-based screening of numerous individuals. However, we failed to isolate a line lacking Lhcb1 genes, which are present in five copies organized at two loci in the Arabidopsis genome. To improve efficiency of our Cas9-based nuclease system, regulatory sequences controlling Cas9 expression levels and timing were systematically compared. Indeed, use of DD45 and RPS5a promoters improved efficiency of our genome editing system by approximately 25-30-fold in comparison to the previous ubiquitin promoter. Using an optimized genome editing system with RPS5a promoter-driven Cas9, putatively quintuple mutant lines lacking detectable amounts of Lhcb1 protein represented approximately 30% of T1 transformants. These results show how improved genome editing systems facilitate the isolation of complex mutant alleles, previously considered impossible to generate, at high frequency even in a single (T1) generation.

Plants with less chlorophyll: A global change perspective.

The necessary reduction of greenhouse gas (GHG) emissions may lead in the future to an increase in solar irradiance (solar brightening). Anthropogenic aerosols (and their precursors) that cause solar dimming are in fact often co-emitted with GHGs. While the reduction of GHG emissions is expected to slow down the ongoing increase in the greenhouse effect, an increased surface irradiance due to reduced atmospheric aerosol load might occur in the most populated areas of the earth. Increased irradiance may lead to air warming, favour the occurrence of heatwaves and increase the evaporative demand of the atmosphere. This is why effective and sustainable solar radiation management strategies to reflect more light back to space should be designed, tested and implemented together with GHG emission mitigation. Here we propose that new plants (crops, orchards and forests) with low-chlorophyll (Chl) content may provide a realistic, sustainable and relatively simple solution to increase surface reflectance of large geographical areas via changes in surface albedo. This may finally offset all or part of the expected local solar brightening. While high-Chl content provides substantial competitive advantages to plants growing in their natural environment, new plants with low-Chl content may be successfully used in agriculture and silviculture and be as productive as the green wildtypes (or even more). The most appropriate strategies to obtain highly productive and highly reflective plants are discussed in this paper and their mitigation potential is examined together with the challenges associated with their introduction in agriculture.

LHC-like proteins involved in stress responses and biogenesis/repair of the photosynthetic apparatus.

LHC (light-harvesting complex) proteins of plants and algae are known to be involved both in collecting light energy for driving the primary photochemical reactions of photosynthesis and in photoprotection when the absorbed light energy exceeds the capacity of the photosynthetic apparatus. These proteins usually contain three transmembrane (TM) helices which span the thylakoid membranes and bind several chlorophyll, carotenoid and lipid molecules. In addition, the LHC protein family includes LHC-like proteins containing one, two, three or even four TM domains. One-helix proteins are not only present in eukaryotic photosynthetic organisms but also in cyanobacteria where they have been named high light-inducible proteins. These small proteins are probably the ancestors of the members of the extant LHC protein family which arouse through gene duplications, deletions and fusions. During evolution, some of these proteins have diverged and acquired novel functions. In most cases, LHC-like proteins are induced in response to various stress conditions including high light, high salinity, elevated temperature and nutrient limitation. Many of these proteins play key roles in photoprotection, notably in non-photochemical quenching of absorbed light energy. Moreover, some of these proteins appear to be involved in the regulation of chlorophyll synthesis and in the assembly and repair of Photosystem II and also of Photosystem I possibly by mediating the insertion of newly synthesized pigments into the photosynthetic reaction centers.

The Physcomitrella patens gene atlas project: large-scale RNA-seq based expression data.

High-throughput RNA sequencing (RNA-seq) has recently become the method of choice to define and analyze transcriptomes. For the model moss Physcomitrella patens, although this method has been used to help analyze specific perturbations, no overall reference dataset has yet been established. In the framework of the Gene Atlas project, the Joint Genome Institute selected P. patens as a flagship genome, opening the way to generate the first comprehensive transcriptome dataset for this moss. The first round of sequencing described here is composed of 99 independent libraries spanning 34 different developmental stages and conditions. Upon dataset quality control and processing through read mapping, 28 509 of the 34 361 v3.3 gene models (83%) were detected to be expressed across the samples. Differentially expressed genes (DEGs) were calculated across the dataset to permit perturbation comparisons between conditions. The analysis of the three most distinct and abundant P. patens growth stages - protonema, gametophore and sporophyte - allowed us to define both general transcriptional patterns and stage-specific transcripts. As an example of variation of physico-chemical growth conditions, we detail here the impact of ammonium supplementation under standard growth conditions on the protonemal transcriptome. Finally, the cooperative nature of this project allowed us to analyze inter-laboratory variation, as 13 different laboratories around the world provided samples. We compare differences in the replication of experiments in a single laboratory and between different laboratories.

Dynamic Changes between Two LHCX-Related Energy Quenching Sites Control Diatom Photoacclimation.

Marine diatoms are prominent phytoplankton organisms that perform photosynthesis in extremely variable environments. Diatoms possess a strong ability to dissipate excess absorbed energy as heat via nonphotochemical quenching (NPQ). This process relies on changes in carotenoid pigment composition (xanthophyll cycle) and on specific members of the light-harvesting complex family specialized in photoprotection (LHCXs), which potentially act as NPQ effectors. However, the link between light stress, NPQ, and the existence of different LHCX isoforms is not understood in these organisms. Using picosecond fluorescence analysis, we observed two types of NPQ in the pennate diatom Phaeodactylum tricornutum that were dependent on light conditions. Short exposure of low-light-acclimated cells to high light triggers the onset of energy quenching close to the core of photosystem II, while prolonged light stress activates NPQ in the antenna. Biochemical analysis indicated a link between the changes in the NPQ site/mechanism and the induction of different LHCX isoforms, which accumulate either in the antenna complexes or in the core complex. By comparing the responses of wild-type cells and transgenic lines with a reduced expression of the major LHCX isoform, LHCX1, we conclude that core complex-associated NPQ is more effective in photoprotection than is the antenna complex. Overall, our data clarify the complex molecular scenario of light responses in diatoms and provide a rationale for the existence of a degenerate family of LHCX proteins in these algae.

Functional analysis of LHCSR1, a protein catalyzing NPQ in mosses, by heterologous expression in Arabidopsis thaliana.

Non-photochemical quenching, NPQ, of chlorophyll fluorescence regulates the heat dissipation of chlorophyll excited states and determines the efficiency of the oxygenic photosynthetic systems. NPQ is regulated by a pH-sensing protein, responding to the chloroplast lumen acidification induced by excess light, coupled to an actuator, a chlorophyll/xanthophyll subunit where quenching reactions are catalyzed. In plants, the sensor is PSBS, while the two pigment-binding proteins Lhcb4 (also known as CP29) and LHCII are the actuators. In algae and mosses, stress-related light-harvesting proteins (LHCSR) comprise both functions of sensor and actuator within a single subunit. Here, we report on expressing the lhcsr1 gene from the moss Physcomitrella patens into several Arabidopsis thaliana npq4 mutants lacking the pH sensing PSBS protein essential for NPQ activity. The heterologous protein LHCSR1 accumulates in thylakoids of A. thaliana and NPQ activity can be partially restored. Complementation of double mutants lacking, besides PSBS, specific xanthophylls, allowed analyzing chromophore requirement for LHCSR-dependent quenching activity. We show that the partial recovery of NPQ is mostly due to the lower levels of Zeaxanthin in A. thaliana in comparison to P. patens. Complemented npq2npq4 mutants, lacking besides PSBS, Zeaxanthin Epoxidase, showed an NPQ recovery of up to 70% in comparison to A. thaliana wild type. Furthermore, we show that Lutein is not essential for the folding nor for the quenching activity of LHCSR1. In short, we have developed a system to study the function of LHCSR proteins using heterologous expression in a variety of A. thaliana mutants.

The STN8 kinase-PBCP phosphatase system is responsible for high-light-induced reversible phosphorylation of the PSII inner antenna subunit CP29 in rice.

Reversible phosphorylation of thylakoid light-harvesting proteins is a mechanism to compensate for unbalanced excitation of photosystem I (PSI) versus photosystem II (PSII) under limiting light. In monocots, an additional phosphorylation event on the PSII antenna CP29 occurs upon exposure to excess light, enhancing resistance to light stress. Different from the case of the major LHCII antenna complex, the STN7 kinase and its related PPH1 phosphatase were proven not to be involved in CP29 phosphorylation, indicating that a different set of enzymes act in the high-light (HL) response. Here, we analyze a rice stn8 mutant in which both PSII core proteins and CP29 phosphorylation are suppressed in HL, implying that STN8 is the kinase catalyzing this reaction. In order to identify the phosphatase involved, we produced a recombinant enzyme encoded by the rice ortholog of AtPBCP, antagonist of AtSTN8, which catalyzes the dephosphorylation of PSII core proteins. The recombinant protein was active in dephosphorylating P-CP29. Based on these data, we propose that the activities of the OsSTN8 kinase and the antagonistic OsPBCP phosphatase, in addition to being involved in the repair of photo-damaged PSII, are also responsible for the HL-dependent reversible phosphorylation of the inner antenna CP29.

Molecular mechanisms involved in plant photoprotection.

Photosynthesis uses sunlight to convert water and carbon dioxide into biomass and oxygen. When in excess, light can be dangerous for the photosynthetic apparatus because it can cause photo-oxidative damage and decreases the efficiency of photosynthesis because of photoinhibition. Plants have evolved many photoprotective mechanisms in order to face reactive oxygen species production and thus avoid photoinhibition. These mechanisms include quenching of singlet and triplet excited states of chlorophyll, synthesis of antioxidant molecules and enzymes and repair processes for damaged photosystem II and photosystem I reaction centers. This review focuses on the mechanisms involved in photoprotection of chloroplasts through dissipation of energy absorbed in excess.