Role Of Chloroplasts Research Articles

The mechanisms of photoinhibition and repair in plants under high light conditions and interplay with abiotic stressors

This review comprehensively examines the phenomenon of photoinhibition in plants, focusing mainly on the intricate relationship between photodamage and photosystem II (PSII) repair and the role of PSII extrinsic proteins and protein phosphorylation in these processes. In natural environments, photoinhibition occurs together with a suite of concurrent stress factors, including extreme temperatures, drought and salinization. Photoinhibition, primarily caused by high irradiance, results in a critical imbalance between the rate of PSII photodamage and its repair. Central to this process is the generation of reactive oxygen species (ROS), which not only impair the photosynthetic apparatus first PSII but also play a signalling role in chloroplasts and other cellulular structures. ROS generated under stress conditions inhibit the repair of photodamaged PSII by suppressing D1 protein synthesis and affecting PSII protein phosphorylation. Furthermore, this review considers how environmental stressors exacerbate PSII damage by interfering with PSII repair primarily by reducing de novo protein synthesis. In addition to causing direct damage, these stressors also contribute to ROS production by restricting CO2 fixation, which also reduces the intensity of protein synthesis. This knowledge has significant implications for agricultural practices and crop improvement under stressful conditions.

CBSX2 is required for the efficient oxidation of chloroplast redox-regulated enzymes in darkness.

Thiol/disulfide-based redox regulation in plant chloroplasts is essential for controlling the activity of target proteins in response to light signals. One of the examples of such a role in chloroplasts is the activity of the chloroplast ATP synthase (CFoCF1), which is regulated by the redox state of the CF1γ subunit and involves two cysteines in its central domain. To investigate the mechanism underlying the oxidation of CF1γ and other chloroplast redox-regulated enzymes in the dark, we characterized the Arabidopsis cbsx2 mutant, which was isolated based on its altered NPQ (non-photochemical quenching) induction upon illumination. Whereas in dark-adapted WT plants CF1γ was completely oxidized, a small amount of CF1γ remained in the reduced state in cbsx2 under the same conditions. In this mutant, reduction of CF1γ was not affected in the light, but its oxidation was less efficient during a transition from light to darkness. The redox states of the Calvin cycle enzymes FBPase and SBPase in cbsx2 were similar to those of CF1γ during light/dark transitions. Affinity purification and subsequent analysis by mass spectrometry showed that the components of the ferredoxin-thioredoxin reductase/thioredoxin (FTR-Trx) and NADPH-dependent thioredoxin reductase (NTRC) systems as well as several 2-Cys peroxiredoxins (Prxs) can be co-purified with CBSX2. In addition to the thioredoxins, yeast two-hybrid analysis showed that CBSX2 also interacts with NTRC. Taken together, our results suggest that CBSX2 participates in the oxidation of the chloroplast redox-regulated enzymes in darkness, probably through regulation of the activity of chloroplast redox systems in vivo.

The tomato chloroplast stromal proteome compendium elucidated by leveraging a plastid protein-localization prediction Atlas.

Tomato (Solanum lycopersicum) is a model species for studying fruit development, wounding, herbivory, and pathogen attack. Despite tomato's world-wide economic importance and the role of chloroplasts as metabolic hubs and integrators of environmental cues, little is known about the stromal proteome of tomato. Using a high-yielding protocol for chloroplast and stromal protein isolation, MudPIT nano-LC-MS/MS analyses, a robust in-house protein database (the Atlas) for predicting the plastid localization of tomato proteins, and rigorous selection criteria for inclusion/exclusion in the stromal proteome, we identified 1,278 proteins of the tomato stromal proteome. We provide one of the most robust stromal proteomes available to date with empirical evidence for 545 and 92 proteins not previously described for tomato plastids and the Arabidopsis stroma, respectively. The relative abundance of tomato stromal proteins was determined using the exponentially modified protein abundance index (emPAI). Comparison of the abundance of tomato and Arabidopsis stromal proteomes provided evidence for the species-specific nature of stromal protein homeostasis. The manual curation of the tomato stromal proteome classified proteins into ten functional categories resulting in an accessible compendium of tomato chloroplast proteins. After curation, only 91 proteins remained as unknown, uncharacterized or as enzymes with unknown functions. The curation of the tomato stromal proteins also indicated that tomato has a number of paralogous proteins, not present in Arabidopsis, which accumulated to different levels in chloroplasts. As some of these proteins function in key metabolic pathways or in perceiving or transmitting signals critical for plant adaptation to biotic and abiotic stress, these data suggest that tomato may modulate the bidirectional communication between chloroplasts and nuclei in a novel manner. The stromal proteome provides a fertile ground for future mechanistic studies in the field of tomato chloroplast-nuclear signaling and are foundational for our goal of elucidating the dynamics of the stromal proteome controlled by the solanaceous-specific, stromal, and wound-inducible leucine aminopeptidase A of tomato.

Competition co-immunoprecipitation reveals the interactors of the chloroplast CPN60 chaperonin machinery.

The functionality of all metabolic processes in chloroplasts depends on a balanced integration of nuclear- and chloroplast-encoded polypeptides into the plastid's proteome. The chloroplast chaperonin machinery is an essential player in chloroplast protein folding under ambient and stressful conditions, with a more intricate structure and subunit composition compared to the orthologous GroEL/ES chaperonin of Escherichia coli. However, its exact role in chloroplasts remains obscure, mainly because of very limited knowledge about the interactors. We employed the competition immunoprecipitation method for the identification of the chaperonin's interactors in Chlamydomonas reinhardtii. Co-immunoprecipitation of the target complex in the presence of increasing amounts of isotope-labelled competitor epitope and subsequent mass spectrometry analysis specifically allowed to distinguish true interactors from unspecifically co-precipitated proteins. Besides known substrates such as RbcL and the expected complex partners, we revealed numerous new interactors with high confidence. Proteins that qualify as putative substrate proteins differ from bulk chloroplast proteins by a higher content of beta-sheets, lower alpha-helical conformation and increased aggregation propensity. Immunoprecipitations targeted against a subunit of the co-chaperonin lid revealed the ClpP protease as a specific partner complex, pointing to a close collaboration of these machineries to maintain protein homeostasis in the chloroplast.

The Crucial Role of Chloroplast-Related Proteins in Viral Genome Replication and Host Defense against Positive-Sense Single-Stranded RNA Viruses.

Plant viruses are responsible for worldwide production losses of numerous economically important crops. The most common plant RNA viruses are positivesense single-stranded RNA viruses [(+)ss RNA viruses]. These viruses have small genomes that encode a limited number of proteins. The viruses depend on their host's machinery for the replication of their RNA genome, assembly, movement, and attraction to the vectors for dispersal. Recently researchers have reported that chloroplast proteins are crucial for replicating (+)ss plant RNA viruses. Some chloroplast proteins, including translation initiation factor [eIF(iso)4E] and 75 DEAD-box RNA helicase RH8, help viruses fulfill their infection cycle in plants. In contrast, other chloroplast proteins such as PAP2.1, PSaC, and ATPsyn-α play active roles in plant defense against viruses. This is also consistent with the idea that reactive oxygen species, salicylic acid, jasmonic acid, and abscisic acid are produced in chloroplast. However, knowledge of molecular mechanisms and functions underlying these chloroplast host factors during the virus infection is still scarce and remains largely unknown. Our review briefly summarizes the latest knowledge regarding the possible role of chloroplast in plant virus replication, emphasizing chloroplast-related proteins. We have highlighted current advances regarding chloroplast-related proteins' role in replicating plant (+)ss RNA viruses.

ROS production and signalling in chloroplasts: cornerstones and evolving concepts.

SUMMARYReactive oxygen species (ROS) such as singlet oxygen, superoxide (O2 ●−) and hydrogen peroxide (H2O2) are the markers of living cells. Oxygenic photosynthesis produces ROS in abundance, which act as a readout of a functional electron transport system and metabolism. The concept that photosynthetic ROS production is a major driving force in chloroplast to nucleus retrograde signalling is embedded in the literature, as is the role of chloroplasts as environmental sensors. The different complexes and components of the photosynthetic electron transport chain (PETC) regulate O2 ●− production in relation to light energy availability and the redox state of the stromal Cys‐based redox systems. All of the ROS generated in chloroplasts have the potential to act as signals and there are many sulphhydryl‐containing proteins and peptides in chloroplasts that have the potential to act as H2O2 sensors and function in signal transduction. While ROS may directly move out of the chloroplasts to other cellular compartments, ROS signalling pathways can only be triggered if appropriate ROS‐sensing proteins are present at or near the site of ROS production. Chloroplast antioxidant systems serve either to propagate these signals or to remove excess ROS that cannot effectively be harnessed in signalling. The key challenge is to understand how regulated ROS delivery from the PETC to the Cys‐based redox machinery is organised to transmit redox signals from the environment to the nucleus. Redox changes associated with stromal carbohydrate metabolism also play a key role in chloroplast signalling pathways.

Whole-Genome Sequence Resource of Fusarium oxysporum Strain TH15, a Plant Growth Promoting Endophytic Fungus Isolated from Tetrastigma hemsleyanum

Whole-Genome Sequence Resource of <i>Fusarium oxysporum</i> Strain TH15, a Plant Growth Promoting Endophytic Fungus Isolated from <i>Tetrastigma hemsleyanum</i>

Chemical Perturbation of Chloroplast Ca2+ Dynamics in Arabidopsis thaliana Suspension Cell Cultures and Seedlings.

Ca2+ signaling is part of universal signal transduction pathways to respond to external and internal stimuli or stress and in plants plays a central role in chloroplasts, such as in the regulation of photosynthetic enzymes or the transition from light to dark. Only recently, the underlying molecular machinery, e.g., transporters and channels that enable chloroplast Ca2+ fluxes, has started to be elucidated. However, chemical tools to specifically perturb these chloroplast Ca2+ fluxes are largely lacking. Here, we describe an efficient aequorin-based system in Arabidopsis thaliana suspension cell cultures to screen for chemicals that alter light-to-dark-induced chloroplast stroma Ca2+ signals. Subsequently, the effect of the hits on chloroplast Ca2+ signals is validated in Arabidopsis seedlings. The research lays a foundation for the identification of novel proteins involved in Ca2+ transport in chloroplast stroma under light-to-dark transition and for investigating the interaction of chloroplast Ca2+ signaling with photosynthesis in general.

Osj10gBTF3-Mediated Import of Chloroplast Protein Is Essential for Pollen Development in Rice.

Chloroplasts are crucial organelles for the generation of fatty acids and starch required for plant development. Nascent polypeptide-associated complex (NAC) proteins have been implicated in development as transcription factors. However, their chaperone roles in chloroplasts and their relationship with pollen development in plants remain to be elucidated. Here, we demonstrated that Osj10gBTF3, a NAC protein, regulates pollen and chloroplast development in rice by coordinating with a Hsp90 family chaperone OsHSP82 to mediate chloroplast import. Knockout of Osj10gBTF3 affects pollen and chloroplast development and significantly reduces the accumulation of fertility-related chloroplast protein OsPPR676. Both Osj10gBTF3 and OsHSP82 interact with OsPPR676. Interestingly, the interaction between OsHSP82 and OsPPR676 is only found in the cytoplasm, while the interaction between Osj10gBTF3 and OsPPR676 also occurs inside the chloroplast. The chloroplast stroma chaperone OsCpn60 can also be co-precipitated with Osj10gBTF3, but not with OsHSP82. Further investigation indicates that Osj10gBTF3 enters the chloroplast stroma possibly through the inner chloroplast membrane channel protein Tic110 and then recruits OsCpn60 for the folding or assembly of OsPPR676. Our results reveal a chaperone role of Osj10gBTF3 in chloroplast import different from Hsp90 and provide a link between chloroplast transport and pollen development in rice.

Multiple cellular compartments engagement in Nicotiana benthamiana-peanut stunt virus-satRNA interactions revealed by systems biology approach

Key messagePSV infection changed the abundance of host plant’s transcripts and proteins associated with various cellular compartments, including ribosomes, chloroplasts, mitochondria, the nucleus and cytosol, affecting photosynthesis, translation, transcription, and splicing.Virus infection is a process resulting in numerous molecular, cellular, and physiological changes, a wide range of which can be analyzed due to development of many high-throughput techniques. Plant RNA viruses are known to replicate in the cytoplasm; however, the roles of chloroplasts and other cellular structures in the viral replication cycle and in plant antiviral defense have been recently emphasized. Therefore, the aim of this study was to analyze the small RNAs, transcripts, proteins, and phosphoproteins affected during peanut stunt virus strain P (PSV-P)–Nicotiana benthamiana interactions with or without satellite RNA (satRNA) in the context of their cellular localization or functional connections with particular cellular compartments to elucidate the compartments most affected during pathogenesis at the early stages of infection. Moreover, the processes associated with particular cell compartments were determined. The ‘omic’ results were subjected to comparative data analyses. Transcriptomic and small RNA (sRNA)–seq data were obtained to provide new insights into PSV-P–satRNA–plant interactions, whereas previously obtained proteomic and phosphoproteomic data were used to broaden the analysis to terms associated with cellular compartments affected by virus infection. Based on the collected results, infection with PSV-P contributed to changes in the abundance of transcripts and proteins associated with various cellular compartments, including ribosomes, chloroplasts, mitochondria, the nucleus and the cytosol, and the most affected processes were photosynthesis, translation, transcription, and mRNA splicing. Furthermore, sRNA-seq and phosphoproteomic analyses indicated that kinase regulation resulted in decreases in phosphorylation levels. The kinases were associated with the membrane, cytoplasm, and nucleus components.

Chloroplast: The Emerging Battlefield in Plant-Microbe Interactions.

Higher plants and some algae convert the absorbed light into chemical energy through one of the most important organelles, chloroplast, for photosynthesis and store it in the form of organic compounds to supply their life activities. However, more and more studies have shown that the role of chloroplasts is more than a factory for photosynthesis. In the process of light conversion to chemical energy, any damage to the components of chloroplast may affect the photosynthesis efficiency and promote the production of by-products, reactive oxygen species, that are mainly produced in the chloroplasts. Substantial evidence show that chloroplasts are also involved in the battle of plants and microbes. Chloroplasts are important in integrating a variety of external environmental stimuli and regulate plant immune responses by transmitting signals to the nucleus and other cell compartments through retrograde signaling pathways. Besides, chloroplasts can also regulate the biosynthesis and signal transduction of phytohormones, including salicylic acid and jasmonic acid, to affect the interaction between the plants and microbes. Since chloroplasts play such an important role in plant immunity, correspondingly, chloroplasts have become the target of pathogens. Different microbial pathogens target the chloroplast and affect its functions to promote their colonization in the host plants.

Carotenoid accumulation in the eyespot apparatus required for phototaxis is independent of chloroplast development in Euglena gracilis

Euglena gracilis exhibits photomovements in response to various light stimuli, such as phototactic and photophobic responses. Our recent study revealed that carotenoids in the eyespot apparatus are required for triggering phototaxis in this alga. However, the role of chloroplasts in eyespot formation is not understood. Here, we isolated carotenoid-less (cl) strains of E. gracilis from cells silenced gene expression of phytoene synthase (EgcrtB). Unlike WT, the culture colors of cl1, cl3, and the non-photosynthetic mutant SM-ZK were orange, while that of cl4 was white. Electron microscope observations showed that SM-ZK, cl1, and cl3 had no developed chloroplast and formed a normal eyespot apparatus, similar to that of WT, but this was not the case for cl4. Carotenoids detected in WT were diadinoxanthin, neoxanthin, and β-carotene. However, the most abundant species of SM-ZK, cl1, and cl3 was zeaxanthin, and there was no diadinoxanthin or neoxanthin. Photomovement analysis showed that SM-ZK, cl1, and cl3 exhibited negative phototactic and photophobic responses, similar to those of WT, whereas cl4 lacked negative phototaxis. Taken together, the formation of the eyespot apparatus required for phototaxis is independent of chloroplast development in E. gracilis, suggesting that this property is different from other photosynthetic flagellates.

Plastid ribosomal protein LPE2 is involved in photosynthesis and the response to C/N balance in Arabidopsis thaliana.

The balance between cellular carbon (C) and nitrogen (N) must be tightly coordinated to sustain optimal growth and development in plants. In chloroplasts, photosynthesis converts inorganic C to organic C, which is important for maintenance of C content in plant cells. However, little is known about the role of chloroplasts in C/N balance. Here, we identified a nuclear‐encoded protein LOW PHOTOSYNTHETIC EFFICIENCY2 (LPE2) that it is required for photosynthesis and C/N balance in Arabidopsis. LPE2 is specifically localized in the chloroplast. Both loss‐of‐function mutants, lpe2‐1 and lpe2‐2, showed lower photosynthetic activity, characterized by slower electron transport and lower PSII quantum yield than the wild type. Notably, LPE2 is predicted to encode the plastid ribosomal protein S21 (RPS21). Deficiency of LPE2 significantly perturbed the thylakoid membrane composition and plastid protein accumulation, although the transcription of plastid genes is not affected obviously. More interestingly, transcriptome analysis indicated that the loss of LPE2 altered the expression of C and N response related genes in nucleus, which is confirmed by quantitative real‐time‐polymerase chain reaction. Moreover, deficiency of LPE2 suppressed the response of C/N balance in physiological level. Taken together, our findings suggest that LPE2 plays dual roles in photosynthesis and the response to C/N balance.

Plant Chloroplast Stress Response: Insights from Thiol Redox Proteomics.

Significance: Plant chloroplasts generate reactive oxygen species (ROS) during photosynthesis, especially under stresses. The sulfhydryl groups of protein cysteine residues are susceptible to redox modifications, which regulate protein structure and function, and thus different signaling and metabolic processes. The ROS-governed protein thiol redox switches play important roles in chloroplasts. Recent Advances: Various high-throughput thiol redox proteomic approaches have been developed, and they have enabled the improved understanding of redox regulatory mechanisms in chloroplasts. For example, the thioredoxin-modulated antioxidant enzymes help to maintain cellular ROS homeostasis. The light- and dark-dependent redox regulation of photosynthetic electron transport, the Calvin/Benson cycle, and starch biosynthesis ensures metabolic coordination and efficient energy utilization. In addition, redox cascades link the light with the dynamic changes of metabolites in nitrate and sulfur assimilation, shikimate pathway, and biosynthesis of fatty acid hormone as well as purine, pyrimidine, and thiamine. Importantly, redox regulation of tetrapyrrole and chlorophyll biosynthesis is critical to balance the photodynamic tetrapyrrole intermediates and prevent oxidative damage. Moreover, redox regulation of diverse elongation factors, chaperones, and kinases plays an important role in the modulation of gene expression, protein conformation, and posttranslational modification that contribute to photosystem II (PSII) repair, state transition, and signaling in chloroplasts. Critical Issues: This review focuses on recent advances in plant thiol redox proteomics and redox protein networks toward understanding plant chloroplast signaling, metabolism, and stress responses. Future Directions: Using redox proteomics integrated with biochemical and molecular genetic approaches, detailed studies of cysteine residues, their redox states, cross talk with other modifications, and the functional implications will yield a holistic understanding of chloroplast stress responses.

Disruption of ζ-Carotene Desaturase Protein ALE1 Leads to Chloroplast Developmental Defects and Seedling Lethality.

ζ-carotene desaturase (ZDS) is a key enzyme in carotenoid biosynthesis and plays an important role in plant photosynthesis. We characterized an albino leaf-color mutant obtained from ethyl methanesulfonate treatment: albino and seedling lethality 1 (ale1). The material contains a chloroplast thylakoid defect where photosynthetic pigments declined and reactive oxygen species accumulated resulting in ale1 death within 3 weeks. Positional cloning and sequencing revealed that there was a single base substitution in ALE1, which encoded a ZDS involved in carotenoid biosynthesis. RNAi and complementation tests confirmed the identity of ALE1. Subcellular localization showed that the ALE1 protein is localized in the chloroplast. Expression analysis indicated that the genes involved in chlorophyll and carotenoid biosynthesis were downregulated. We conclude that ALE1 plays an important role in chloroplast and plant growth in rice.

The role of chloroplasts in the oxidative stress that is induced by zearalenone in wheat plants – The functions of 24-epibrassinolide and selenium in the protective mechanisms

This study focused on the idea that the toxic effect of zearalenone (ZEA) and the protective actions of the brassinosteroid - 24-epibrassinolide (EBR) as well as selenium are dependent on its accumulation in chloroplasts to a high degree. These organelles were isolated from the leaves of oxidative stress-sensitive and stress-tolerant wheat cultivars that had been grown from grains that had been incubated in a solution of ZEA (30 μM), Na2SeO4 (Se, 10 μM), EBR (0.1 μM) or in a mixture of ZEA with Se or EBR. Ultra-high performance liquid chromatography techniques indicated that ZEA was adsorbed in higher amounts in the chloroplasts in the sensitive rather than tolerant cultivar. Although the brassinosteroids and Se were also accumulated in the chloroplasts, higher levels were only found in the tolerant cultivar. The application of EBR increased the homocastasterone content, especially in the chloroplasts of the tolerant plant and after the addition of ZEA. The presence of both protectants caused a decrease in the ZEA content in studied organelles and resulted in diminishing of the oxidative stress (i.e. changes in the activity of the antioxidative enzymes). Moreover, a recovery of photosystem II and decrease in the negative impact of ZEN on Hsp90 transcript accumulation was observed in plants.

Chlorophyll deficient 3, Encoding a Putative Potassium Efflux Antiporter, Affects Chloroplast Development Under High Temperature Conditions in Rice (Oryza sativa L.)

Potassium efflux antiporters (KEAs) play important roles in the regulation of monovalent cation efflux to maintain ion homeostasis in plant cells; however, details of their function in chloroplast development remain elusive, especially under stress conditions. Here, we identified a KEA family gene based on three chlorophyll-deficient mutants, cde3-1, 2, 3. These cde3 mutants exhibit serious white stripe leaf and an albino midrib phenotype, reduced level of photosynthetic pigments, and arrested chloroplast development under high-temperature conditions. Map-based cloning and allelism tests showed that CDE3 encodes a putative KEA protein in rice. CDE3 is mainly expressed in green tissues, with maximum transcript abundance in young leaves. Subcellular localization analysis showed that CDE3 is localized in chloroplasts. Quantitative real-time PCR analysis showed that expression levels of genes associated with chlorophyll biosynthesis, photosynthesis, and chloroplast development are affected in cde3-1, which suggest that CDE3 plays an important role in chloroplast K+/H+ homeostasis and chloroplast development under high-temperature conditions.

The role of chloroplasts in plant pathology.

Plants have evolved complex tolerance systems to survive abiotic and biotic stresses. Central to these programmes is a sophisticated conversation of signals between the chloroplast and the nucleus. In this review, we examine the antagonism between abiotic stress tolerance (AST) and immunity: we propose that to generate immunogenic signals, plants must disable AST systems, in particular those that manage reactive oxygen species (ROS), while the pathogen seeks to reactivate or enhance those systems to achieve virulence. By boosting host systems of AST, pathogens trick the plant into suppressing chloroplast immunogenic signals and steer the host into making an inappropriate immune response. Pathogens disrupt chloroplast function, both transcriptionally-by secreting effectors that alter host gene expression by interacting with defence-related kinase cascades, with transcription factors, or with promoters themselves-and post-transcriptionally, by delivering effectors that enter the chloroplast or alter the localization of host proteins to change chloroplast activities. These mechanisms reconfigure the chloroplast proteome and chloroplast-originating immunogenic signals in order to promote infection.

Reactivation of electron flow in chloroplasts of in vitro shootlets of apple through elimination of carbon source and evaluation of its activity by inhibitors of electron transport chain and chlorophyll fluorescence quenching

Electron transport chains (ETC) are drastically inactivated in chloroplasts of in vitro shootlets due to presence of carbon source in the growth media. This inactivation consequently creates limitations for in vitro studies on the role of chloroplasts in biotic and abiotic stresses. This research was carried out to evaluate reactivation of electron flow in chloroplasts ETC by elimination of carbon source and subsequent comparison of the effects of ETC inhibitors in the presence and absence of carbon source (i.e., sucrose) on the in vitro shootlets of apple rootstock, MM-111. All the tested ETC inhibitors, including uracil; glutaraldehyde, methyl viologen (MV) and DCMU triggered necrosis appearance on the in vitro apple shootlets exclusively on the carbon source-free media. Moreover, exposure of the in vitro shootlets to the various concentrations of ETC inhibitors demonstrated different severity of ETC inhibition by these inhibitors and their concentration-dependent effects on chloroplasts ETC. Uracil, the inhibitor of photosystem II, triggered the weakest shootlets necrosis, while DCMU exhibited no necrogenic effects at 50 mg/L and lower concentrations. MV, contrary to glutaraldehyde, did not have necrogenesis effects at 1 mg/L, but both of them caused sharp necrosis trigger on the shootlets as soon as 48–96 h after exposure to their effective concentrations. Evaluation of the shootlets H+-ATPase activity revealed that proton extrusion and pH decline in the growth media were mainly associated with the sucrose uptake by the shootlets, while no considerable pH decline observed in the presence of carbon source in the media. Finally, chloroplasts ETC activity of the shootlets was confirmed by higher percentage of O2 and less CO2 quantity in the atmospheres of the jars containing sucrose-free media, as well as by higher initial fluorescence (Fo) and maximum fluorescence (Fm) quenching parameters in the shootlets, in comparison with the sucrose-enriched condition.

Recombinant ACHT1 from Arabidopsis thaliana: crystallization and X-ray crystallographic analysis.

Thioredoxins (Trxs) play important roles in chloroplasts by linking photosynthetic light reactions to a series of plastid functions. They execute their function by regulating the oxidation and reduction of disulfide bonds. ACHT1 (atypical cysteine/histidine-rich Trx1) is a thylakoid-associated thioredoxin-type protein found in the Arabidopsis thaliana chloroplast. Recombinant ACHT1 protein was overexpressed in Escherichia coli, purified and crystallized by the vapour-diffusion method. The crystal diffracted to 1.7 Å resolution and a complete X-ray data set was collected. Preliminary crystallographic analysis suggested that the crystals belonged to space group C2221, with unit-cell parameters a=102.7, b = 100.6, c = 92.8 Å.