Structure of the multi-subunit chloroplast RNA polymerase.

Chloroplasts contain a dedicated genome that encodes subunits of the photosynthesis machinery. Transcription of photosynthesis genes is predominantly carried out by a plastid-encoded RNA polymerase (PEP), a nearly 1 MDa complex composed of core subunits with homology to eubacterial RNA polymerases (RNAPs) and at least 12 additional chloroplast-specific PEP-associated proteins (PAPs). However, the architecture of this complex and the functions of the PAPs remain unknown. Here, we report the cryo-EM structure of a 19-subunit PEP complex from Sinapis alba (white mustard). The structure reveals that the PEP core resembles prokaryotic and nuclear RNAPs but contains chloroplast-specific features that mediate interactions with the PAPs. The PAPs are unrelated to known transcription factors and arrange around the core in a unique fashion. Their structures suggest potential functions during transcription in the chemical environment of chloroplasts. These results reveal structural insights into chloroplast transcription and provide a framework for understanding photosynthesis gene expression.

Limiting etioplast gene expression induces apical hook twisting during skotomorphogenesis of Arabidopsis seedlings.

When covered by a layer of soil, seedling development follows a dark-specific program (skotomorphogenesis). In the dark, seedlings consist of small, non-green cotyledons, a long hypocotyl, and an apical hook to protect meristematic cells. We recently highlighted the role played by mitochondria in the high energy-consuming reprogramming of Arabidopsis skotomorphogenesis. Here, the role played by plastids, another energy-supplying organelle, in skotomorphogenesis is investigated. This study was conducted in dark conditions to exclude light signals so as to better focus on those produced by plastids. It was found that limitation of plastid gene expression (PGE) induced an exaggerated apical hook bending. Inhibition of PGE was obtained at the levels of transcription and translation using the antibiotics rifampicin (RIF) and spectinomycin, respectively, as well as plastid RPOTp RNA polymerase mutants. RIF-treated seedlings also showed expression induction of marker nuclear genes for mitochondrial stress, perturbation of mitochondrial metabolism, increased ROS levels, and an augmented capacity of oxygen consumption by mitochondrial alternative oxidases (AOXs). AOXs act to prevent overreduction of the mitochondrial electron transport chain. Previously, we reported that AOX1A, the main AOX isoform, is a key component in the developmental response to mitochondrial respiration deficiency. In this work, we suggest the involvement of AOX1A in the response to PGE dysfunction and propose the importance of signaling between plastids and mitochondria. Finally, it was found that seedling architecture reprogramming in response to RIF was independent of canonical organelle retrograde pathways and the ethylene signaling pathway.

Nucleo-plastidic PAP8/pTAC6 couples chloroplast formation with photomorphogenesis.

The initial greening of angiosperms involves light activation of photoreceptors that trigger photomorphogenesis, followed by the development of chloroplasts. In these semi-autonomous organelles, construction of the photosynthetic apparatus depends on the coordination of nuclear and plastid gene expression. Here, we show that the expression of PAP8, an essential subunit of the plastid-encoded RNA polymerase (PEP) in Arabidopsis thaliana, is under the control of a regulatory element recognized by the photomorphogenic factor HY5. PAP8 protein is localized and active in both plastids and the nucleus, and particularly required for the formation of late photobodies. In the pap8 albino mutant, phytochrome-mediated signalling is altered, degradation of the chloroplast development repressors PIF1/PIF3 is disrupted, HY5 is not stabilized, and the expression of the photomorphogenesis regulator GLK1 is impaired. PAP8 translocates into plastids via its targeting pre-sequence, interacts with the PEP and eventually reaches the nucleus, where it can interact with another PEP subunit pTAC12/HMR/PAP5. Since PAP8 is required for the phytochrome B-mediated signalling cascade and the reshaping of the PEP activity, it may coordinate nuclear gene expression with PEP-driven chloroplastic gene expression during chloroplast biogenesis.

Biogenic signals from plastids and their role in chloroplast development.

Plant seeds do not contain differentiated chloroplasts. Upon germination, the seedlings thus need to gain photoautotrophy before storage energies are depleted. This requires the coordinated expression of photosynthesis genes encoded in nuclear and plastid genomes. Chloroplast biogenesis needs to be additionally coordinated with the light regulation network that controls seedling development. This coordination is achieved by nucleus to plastid signals called anterograde and plastid to nucleus signals termed retrograde. Retrograde signals sent from plastids during initial chloroplast biogenesis are also called biogenic signals. They have been recognized as highly important for proper chloroplast biogenesis and for seedling development. The molecular nature, transport, targets, and signalling function of biogenic signals are, however, under debate. Several studies disproved the involvement of a number of key components that were at the base of initial models of retrograde signalling. New models now propose major roles for a functional feedback between plastid and cytosolic protein homeostasis in signalling plastid dysfunction as well as the action of dually localized nucleo-plastidic proteins that coordinate chloroplast biogenesis with light-dependent control of seedling development. This review provides a survey of the developments in this research field, summarizes the unsolved questions, highlights several recent advances, and discusses potential new working modes.

PAP8/pTAC6 Is Part of a Nuclear Protein Complex and Displays RNA Recognition Motifs of Viral Origin.

Chloroplast biogenesis depends on a complex transcriptional program involving coordinated expression of plastid and nuclear genes. In particular, photosynthesis-associated plastid genes are expressed by the plastid-encoded polymerase (PEP) that undergoes a structural rearrangement during chloroplast formation. The prokaryotic-type core enzyme is rebuilt into a larger complex by the addition of nuclear-encoded PEP-associated proteins (PAP1 to PAP12). Among the PAPs, some have been detected in the nucleus (PAP5 and PAP8), where they could serve a nuclear function required for efficient chloroplast biogenesis. Here, we detected PAP8 in a large nuclear subcomplex that may include other subunits of the plastid-encoded RNA polymerase. We have made use of PAP8 recombinant proteins in Arabidopsis thaliana to decouple its nucleus- and chloroplast-associated functions and found hypomorphic mutants pointing at essential amino acids. While the origin of the PAP8 gene remained elusive, we have found in its sequence a micro-homologous domain located within a large structural homology with a rhinoviral RNA-dependent RNA polymerase, highlighting potential RNA recognition motifs in PAP8. PAP8 in vitro RNA binding activity suggests that this domain is functional. Hence, we propose that the acquisition of PAPs may have occurred during evolution by different routes, including lateral gene transfer.

Three-Dimensional Envelope and Subunit Interactions of the Plastid-Encoded RNA Polymerase from Sinapis alba.

RNA polymerases (RNAPs) are found in all living organisms. In the chloroplasts, the plastid-encoded RNA polymerase (PEP) is a prokaryotic-type multimeric RNAP involved in the selective transcription of the plastid genome. One of its active states requires the assembly of nuclear-encoded PEP-Associated Proteins (PAPs) on the catalytic core, producing a complex of more than 900 kDa, regarded as essential for chloroplast biogenesis. In this study, sequence alignments of the catalytic core subunits across various chloroplasts of the green lineage and prokaryotes combined with structural data show that variations are observed at the surface of the core, whereas internal amino acids associated with the catalytic activity are conserved. A purification procedure compatible with a structural analysis was used to enrich the native PEP from Sinapis alba chloroplasts. A mass spectrometry (MS)-based proteomic analysis revealed the core components, the PAPs and additional proteins, such as FLN2 and pTAC18. MS coupled with crosslinking (XL-MS) provided the initial structural information in the form of protein clusters, highlighting the relative position of some subunits with the surfaces of their interactions. Using negative stain electron microscopy, the PEP three-dimensional envelope was calculated. Particles classification shows that the protrusions are very well-conserved, offering a framework for the future positioning of all the PAPs. Overall, the results show that PEP-associated proteins are firmly and specifically associated with the catalytic core, giving to the plastid transcriptional complex a singular structure compared to other RNAPs.

Working day and night: plastid casein kinase 2 catalyses phosphorylation of proteins with diverse functions in light- and dark-adapted plastids.

Casein kinase 2 is a ubiquitous protein kinase that has puzzled researchers for several decades because of its pleiotropic activity. Here, we set out to identify the in vivo targets of plastid casein kinase 2 (pCK2) in Arabidopsis thaliana. Survey phosphoproteome analyses were combined with targeted analyses with wild-type and pck2 knockdown mutants to identify potential pCK2 targets by their decreased phosphorylation state in the mutant. To validate potential substrates, we complemented the pck2 knockdown line with tandem affinity tag (TAP)-tagged pCK2 and found it to restore growth parameters, as well as many, but not all, putative pCK2-dependent phosphorylation events. We further performed a targeted analysis at the end-of-night to increase the specificity of target protein identification. This analysis confirmed light-independent phosphorylation of several pCK2 target proteins. Based on the aforementioned data, we define a set of in vivo pCK2-targets that span different chloroplast functions, such as metabolism, transcription, translation and photosynthesis. The pleiotropy of pCK2 functions is also manifested by altered state transition kinetics during short-term acclimation and significant alterations in the mutant metabolism, supporting its function in photosynthetic regulation. Thus, our data expand our understanding on chloroplast phosphorylation networks and provide insights into kinase networks in the regulation of chloroplast functions.

Occurrence of albinism during wheat androgenesis is correlated with repression of the key genes required for proper chloroplast biogenesis.

MAIN CONCLUSION: The phenomenon of albinism in wheat androgenesis is linked to the transcriptional repression of specific genes involved in chloroplast biogenesis during the first weeks of in vitro culture. Isolated microspore culture is widely used to accelerate breeding programs and produce new cultivars. However, in cereals and particularly in wheat, the use of this technique is limited due to the high proportion of regenerated albino plantlets. The causes and mechanisms leading to the formation of albino plantlets in wheat remain largely unknown and, to date, no concrete solution has been found to make it possible to overcome this barrier. We performed a molecular study of proplastid-to-chloroplast differentiation within wheat microspore cultures by analyzing the expression of 20 genes specifically involved in chloroplast biogenesis. Their expression levels were compared between two wheat genotypes that exhibit differential capacities to regenerate green plantlets, i.e., Pavon and Paledor, which produce high and low rates of green plants, respectively. We observed that chloroplast biogenesis within wheat microspores was affected as of the very early stages of the androgenesis process. A successful transition from a NEP- to a PEP-dependent transcription during early plastid development was found to be strongly correlated with the formation of green plantlets, while failure of this transition was strongly correlated with the regeneration of albino plantlets. The very low expression of plastid-encoded 16S and 23S rRNAs within plastids of the recalcitrant genotype Paledor suggests a low translation activity in albino plastids. Furthermore, a delay in the activation of the transcription of nuclear encoded key genes like GLK1 related to chloroplast biogenesis was observed in multicellular structures and pro-embryos of the genotype Paledor. These data help to understand the phenomenon of albinism in wheat androgenesis, which appears to be linked to the transcriptional activation of specific genes involved in the initial steps of chloroplast biogenesis that occurs between days 7 and 21 of in vitro culture.

PAP genes are tissue- and cell-specific markers of chloroplast development.

MAIN CONCLUSION: Expression of PAP genes is strongly coordinated and represents a highly selective cell-specific marker associated with the development of chloroplasts in photosynthetically active organs of Arabidopsis seedlings and adult plants. Transcription in plastids of plants depends on the activity of phage-type single-subunit nuclear-encoded RNA polymerases (NEP) and a prokaryotic multi-subunit plastid-encoded RNA polymerase (PEP). PEP is comprised of the core subunits α, ß, ß' and ß″ encoded by rpoA, rpoB/C1/C2 genes located on the plastome. This core enzyme needs to interact with nuclear-encoded sigma factors for proper promoter recognition. In chloroplasts, the core enzyme is surrounded by additional 12 nuclear-encoded subunits, all of eukaryotic origin. These PEP-associated proteins (PAPs) were found to be essential for chloroplast biogenesis as Arabidopsis inactivation mutants for each of them revealed albino or pale-green phenotypes. In silico analysis of transcriptomic data suggests that PAP genes represent a tightly controlled regulon, whereas wetlab data are sparse and correspond to the expression of individual genes mostly studied at the seedling stage. Using RT-PCR, transient, and stable expression assays of PAP promoter-GUS-constructs, we do provide, in this study, a comprehensive expression catalogue for PAP genes throughout the life cycle of Arabidopsis. We demonstrate a selective impact of light on PAP gene expression and uncover a high tissue specificity that is coupled to developmental progression especially during the transition from skotomorphogenesis to photomorphogenesis. Our data imply that PAP gene expression precedes the formation of chloroplasts rendering PAP genes a tissue- and cell-specific marker of chloroplast biogenesis.

Light and Plastid Signals Regulate Different Sets of Genes in the Albino Mutant Pap7-1.

Plants possessing dysfunctional plastids due to defects in pigment biosynthesis or translation are known to repress photosynthesis-associated nuclear genes via retrograde signals from the disturbed organelles toward the nucleus. These signals are thought to be essential for proper biogenesis and function of the plastid. Mutants lacking plastid-encoded RNA polymerase-associated proteins (PAPs) display a genetic arrest in eoplast-chloroplast transition leading to an albino phenotype in the light. Retrograde signaling in these mutants, therefore, could be expected to be similar as under conditions inducing plastid dysfunction. To answer this question, we performed plastome- and genomewide array analyses in the pap7-1 mutant of Arabidopsis (Arabidopsis thaliana). In parallel, we determined the potential overlap with light-regulated expression networks. To this end, we performed a comparative expression profiling approach using light- and dark-grown wild-type plants as relative control for the expression profiles obtained from light-grown pap7-1 mutants. Our data indicate a specific impact of retrograde signals on metabolism-related genes in pap7-1 mutants reflecting the starvation situation of the albino seedlings. In contrast, light regulation of PhANGs and other nuclear gene groups appears to be fully functional in this mutant, indicating that a block in chloroplast biogenesis per se does not repress expression of them as suggested by earlier studies. Only genes for light harvesting complex proteins displayed a significant repression indicating an exclusive retrograde impact on this gene family. Our results indicate that chloroplasts and arrested plastids each emit specific signals that control different target gene modules both in positive and negative manner.

Characterization of the psbH precursor RNAs reveals a precise endoribonuclease cleavage site in the psbT/psbH intergenic region that is dependent on psbN gene expression.

The plastid psbB operon harbours 5 genes, psbB, psbT, psbH, petB and petD. A sixth gene, the psbN gene, is located on the opposite DNA strand in the psbT/psbH intergenic region. Its transcription produces antisense RNA to a large part of the psbB pentacistronic mRNA. We have investigated whether transcription of the psbN gene, i.e. production of antisense RNA, influences psbT/psbH intergenic processing. Results reveal the existence of four different psbH precursor RNAs. Three of them result from processing and one is produced by transcription initiation. One of the processed RNAs is probably created by site-specific RNA cleavage. This RNA is absent in plants where the psbN gene is not transcribed suggesting that cleavage at this site is dependent on the formation of sense/antisense double-stranded RNA. In order to characterize the nuclease that might be responsible for double-stranded RNA cleavage, we analysed csp41a and csp41b knock-out mutants and the corresponding double mutant. Both CSP41 proteins are known to interact physically and CSP41a had been shown to cleave within 3'-untranslated region stem-loop structures, which contain double-stranded RNA, in vitro. We demonstrate that the psbH RNA, that is absent in plants where the psbN gene is not transcribed, is also strongly diminished in all csp41 plants. Altogether, results reveal a site-specific endoribonuclease cleavage event that seems to depend on antisense RNA and might implicate endoribonuclease activity of CSP41a.

ZmpTAC12 binds single-stranded nucleic acids and is essential for accumulation of the plastid-encoded polymerase complex in maize.

The plastid-encoded plastid RNA polymerase (PEP) represents the major transcription machinery in mature chloroplasts. Proteomic studies identified four plastome- and at least ten nuclear-encoded proteins making up this multimeric enzyme. Depletion of single subunits is known to result in strongly diminished PEP activity causing severe defects in chloroplast biogenesis. Here, we characterized one PEP subunit in maize, ZmpTAC12, and investigated the molecular basis underlying PEP-deficiency in Zmptac12 mutants. We show that the ZmpTAC12 gene encodes two different protein isoforms, both of which localize dually in chloroplasts and nuclei. Moreover, both variants assemble into the PEP-complex. Analysis of PEP-complex assembly in various maize mutants lacking different PEP-complex components demonstrates that ZmpTAC12, ZmpTAC2, ZmpTAC10 and ZmMurE are each required to accumulate a fully assembled PEP-complex. Antibodies to ZmpTAC12 coimmunoprecipitate a subset of plastid RNAs that are synthesized by PEP-dependent transcription. Gel mobility shift analyses with recombinant ZmpTAC12 revealed binding capabilities with ssRNA and ssDNA, but not dsDNA. Collectively these data demonstrate that ZmpTAC12 is required for the proper build-up of the PEP-complex and that it interacts with single-stranded nucleic acids.

Plastid RNA polymerases: orchestration of enzymes with different evolutionary origins controls chloroplast biogenesis during the plant life cycle.

Chloroplasts are the sunlight-collecting organelles of photosynthetic eukaryotes that energetically drive the biosphere of our planet. They are the base for all major food webs by providing essential photosynthates to all heterotrophic organisms including humans. Recent research has focused largely on an understanding of the function of these organelles, but knowledge about the biogenesis of chloroplasts is rather limited. It is known that chloroplasts develop from undifferentiated precursor plastids, the proplastids, in meristematic cells. This review focuses on the activation and action of plastid RNA polymerases, which play a key role in the development of new chloroplasts from proplastids. Evolutionarily, plastids emerged from the endosymbiosis of a cyanobacterium-like ancestor into a heterotrophic eukaryote. As an evolutionary remnant of this process, they possess their own genome, which is expressed by two types of plastid RNA polymerase, phage-type and prokaryotic-type RNA polymerase. The protein subunits of these polymerases are encoded in both the nuclear and plastid genomes. Their activation and action therefore require a highly sophisticated regulation that controls and coordinates the expression of the components encoded in the plastid and nucleus. Stoichiometric expression and correct assembly of RNA polymerase complexes is achieved by a combination of developmental and environmentally induced programmes. This review highlights the current knowledge about the functional coordination between the different types of plastid RNA polymerases and provides working models of their sequential expression and function for future investigations.

A novel proteinase, SNOWY COTYLEDON4, is required for photosynthetic acclimation to higher light intensities in Arabidopsis.

Excess light can have a negative impact on photosynthesis; thus, plants have evolved many different ways to adapt to different light conditions to both optimize energy use and avoid damage caused by excess light. Analysis of the Arabidopsis (Arabidopsis thaliana) mutant snowy cotyledon4 (sco4) revealed a mutation in a chloroplast-targeted protein that shares limited homology with CaaX-type endopeptidases. The SCO4 protein possesses an important function in photosynthesis and development, with point mutations rendering the seedlings and adult plants susceptible to photooxidative stress. The sco4 mutation impairs the acclimation of chloroplasts and their photosystems to excess light, evidenced in a reduction in photosystem I function, decreased linear electron transfer, yet increased nonphotochemical quenching. SCO4 is localized to the chloroplasts, which suggests the existence of an unreported type of protein modification within this organelle. Phylogenetic and yeast complementation analyses of SCO4-like proteins reveal that SCO4 is a member of an unknown group of higher plant-specific proteinases quite distinct from the well-described CaaX-type endopeptidases RAS Converting Enzyme1 (RCE1) and zinc metallopeptidase STE24 and lacks canonical CaaX activity. Therefore, we hypothesize that SCO4 is a novel endopeptidase required for critical protein modifications within chloroplasts, influencing the function of proteins involved in photosynthesis required for tolerance to excess light.

An Arabidopsis GluTR binding protein mediates spatial separation of 5-aminolevulinic acid synthesis in chloroplasts.

5-Aminolevulinic acid (ALA) is the universal precursor for tetrapyrrole biosynthesis and is synthesized in plants in three enzymatic steps: ligation of glutamate (Glu) to tRNA(Glu) by glutamyl-tRNA synthetase, reduction of activated Glu to Glu-1-semialdehyde by glutamyl-tRNA reductase (GluTR), and transamination to ALA by Glu 1-semialdehyde aminotransferase. ALA formation controls the metabolic flow into the tetrapyrrole biosynthetic pathway. GluTR is proposed to be the key regulatory enzyme that is tightly controlled at transcriptional and posttranslational levels. We identified a GluTR binding protein (GluTRBP; previously called PROTON GRADIENT REGULATION7) that is localized in chloroplasts and part of a 300-kD protein complex in the thylakoid membrane. Although the protein does not modulate activity of ALA synthesis, the knockout of GluTRBP is lethal in Arabidopsis thaliana, whereas mutants expressing reduced levels of GluTRBP contain less heme. GluTRBP expression correlates with a function in heme biosynthesis. It is postulated that GluTRBP contributes to subcompartmentalized ALA biosynthesis by maintaining a portion of GluTR at the plastid membrane that funnels ALA into the heme biosynthetic pathway. These results regarding GluTRBP support a model of plant ALA synthesis that is organized in two separate ALA pools in the chloroplast to provide appropriate substrate amounts for balanced synthesis of heme and chlorophyll.

Photosystem II supercomplex remodeling serves as an entry mechanism for state transitions in Arabidopsis.

Within dense plant populations, strong light quality gradients cause unbalanced excitation of the two photosystems resulting in reduced photosynthetic efficiency. Plants redirect such imbalances by structural rearrangements of the photosynthetic apparatus via state transitions and photosystem stoichiometry adjustments. However, less is known about the function of photosystem II (PSII) supercomplexes in this context. Here, we show in Arabidopsis thaliana that PSII supercomplex remodeling precedes and facilitates state transitions. Intriguingly, the remodeling occurs in the short term, paralleling state transitions, but is also present in a state transition-deficient mutant, indicating that PSII supercomplex generation is independently regulated and does not require light-harvesting complex phosphorylation and movement. Instead, PSII supercomplex remodeling involves reversible phosphorylation of PSII core subunits (preferentially of CP43) and requires the luminal PSII subunit Psb27 for general formation and structural stabilization. Arabidopsis knockout mutants lacking Psb27 display highly accelerated state transitions, indicating that release of PSII supercomplexes is required for phosphorylation and subsequent movement of the antenna. Downregulation of PSII supercomplex number by physiological light treatments also results in acceleration of state transitions confirming the genetic analyses. Thus, supercomplex remodeling is a prerequisite and an important kinetic determinant of state transitions.

Photosynthesis in the Biomass Model Species Lemna minor Displays Plant-Conserved and Species-Specific Features.

Lemnaceae are small freshwater plants with extraordinary high growth rates. We aimed to test whether this correlates with a more efficient photosynthesis, the primary energy source for growth. To this end, we compared photosynthesis properties of the duckweed Lemna minor and the terrestrial model plant Arabidopsis thaliana. Chlorophyll fluorescence analyses revealed high similarity in principle photosynthesis characteristics; however, Lemna exhibited a more effective light energy transfer into photochemistry and more stable photosynthesis parameters especially under high light intensities. Western immunoblot analyses of representative photosynthesis proteins suggested potential post-translational modifications in Lemna proteins that are possibly connected to this. Phospho-threonine phosphorylation patterns of thylakoid membrane proteins displayed a few differences between the two species. However, phosphorylation-dependent processes in Lemna such as photosystem II antenna association and the recovery from high-light-induced photoinhibition were not different from responses known from terrestrial plants. We thus hypothesize that molecular differences in Lemna photosynthesis proteins are associated with yet unidentified mechanisms that improve photosynthesis and growth efficiencies. We also developed a high-magnification video imaging approach for Lemna multiplication which is useful to assess the impact of external factors on Lemna photosynthesis and growth.

Identification of essential subunits in the plastid-encoded RNA polymerase complex reveals building blocks for proper plastid development.

The major RNA polymerase activity in mature chloroplasts is a multisubunit, Escherichia coli-like protein complex called PEP (for plastid-encoded RNA polymerase). Its subunit structure has been extensively investigated by biochemical means. Beside the "prokaryotic" subunits encoded by the plastome-located RNA polymerase genes, a number of additional nucleus-encoded subunits of eukaryotic origin have been identified in the PEP complex. These subunits appear to provide additional functions and regulation modes necessary to adapt transcription to the varying functional situations in chloroplasts. However, despite the enormous progress in genomic data and mass spectrometry techniques, it is still under debate which of these subunits belong to the core complex of PEP and which ones represent rather transient or peripheral components. Here, we present a catalog of true PEP subunits that is based on comparative analyses from biochemical purifications, protein mass spectrometry, and phenotypic analyses. We regard reproducibly identified protein subunits of the basic PEP complex as essential when the corresponding knockout mutants reveal an albino or pale-green phenotype. Our study provides a clearly defined subunit catalog of the basic PEP complex, generating the basis for a better understanding of chloroplast transcription regulation. In addition, the data support a model that links PEP complex assembly and chloroplast buildup during early seedling development in vascular plants.

Dynamic plastid redox signals integrate gene expression and metabolism to induce distinct metabolic states in photosynthetic acclimation in Arabidopsis.

Plants possess acclimation responses in which structural reconfigurations adapt the photosynthetic apparatus to fluctuating illumination. Long-term acclimation involves changes in plastid and nuclear gene expression and is controlled by redox signals from photosynthesis. The kinetics of these signals and the adjustments of energetic and metabolic demands to the changes in the photosynthetic apparatus are currently poorly understood. Using a redox signaling system that preferentially excites either photosystem I or II, we measured the time-dependent impact of redox signals on the transcriptome and metabolome of Arabidopsis thaliana. We observed rapid and dynamic changes in nuclear transcript accumulation resulting in differential and specific expression patterns for genes associated with photosynthesis and metabolism. Metabolite pools also exhibited dynamic changes and indicate readjustments between distinct metabolic states depending on the respective illumination. These states reflect reallocation of energy resources in a defined and reversible manner, indicating that structural changes in the photosynthetic apparatus during long-term acclimation are additionally supported at the level of metabolism. We propose that photosynthesis can act as an environmental sensor, producing retrograde redox signals that trigger two parallel adjustment loops that coordinate photosynthesis and metabolism to adapt plant primary productivity to the environment.

Photosystem II core phosphorylation and photosynthetic acclimation require two different protein kinases.

Illumination changes elicit modifications of thylakoid proteins and reorganization of the photosynthetic machinery. This involves, in the short term, phosphorylation of photosystem II (PSII) and light-harvesting (LHCII) proteins. PSII phosphorylation is thought to be relevant for PSII turnover, whereas LHCII phosphorylation is associated with the relocation of LHCII and the redistribution of excitation energy (state transitions) between photosystems. In the long term, imbalances in energy distribution between photosystems are counteracted by adjusting photosystem stoichiometry. In the green alga Chlamydomonas and the plant Arabidopsis, state transitions require the orthologous protein kinases STT7 and STN7, respectively. Here we show that in Arabidopsis a second protein kinase, STN8, is required for the quantitative phosphorylation of PSII core proteins. However, PSII activity under high-intensity light is affected only slightly in stn8 mutants, and D1 turnover is indistinguishable from the wild type, implying that reversible protein phosphorylation is not essential for PSII repair. Acclimation to changes in light quality is defective in stn7 but not in stn8 mutants, indicating that short-term and long-term photosynthetic adaptations are coupled. Therefore the phosphorylation of LHCII, or of an unknown substrate of STN7, is also crucial for the control of photosynthetic gene expression.