Photosynthesis Enzymes Research Articles

Removal of redox-sensitive Rubisco Activase does not alter Rubisco regulation in soybean

Rubisco activase (Rca) facilitates the catalytic repair of Rubisco, the CO2-fixing enzyme of photosynthesis, following periods of darkness, low to high light transitions or stress. Removal of the redox-regulated isoform of Rubisco activase, Rca-α, enhances photosynthetic induction in Arabidopsis and has been suggested as a strategy for the improvement of crops, which may experience frequent light transitions in the field; however, this has never been tested in a crop species. Therefore, we used RNAi to reduce the Rca-α content of soybean (Glycinemax cv. Williams82) below detectable levels and then characterized the growth, photosynthesis, and Rubisco activity of the resulting transgenics, in both growth chamber and field conditions. Under a 16h sine wave photoperiod, the reduction of Rca-α contents had no impact on morphological characteristics, leaf expansion rate, or total biomass. Photosynthetic induction rates were unaltered in both chamber-grown and field-grown plants. Plants with reduced Rca-α content maintained the ability to regulate Rubisco activity in low light just as in control plants. This result suggests that in soybean, Rca-α is not as centrally involved in the regulation of Rca oligomer activity as it is in Arabidopsis. The isoform stoichiometry supports this conclusion, as Rca-α comprises only ~ 10% of the Rubisco activase content of soybean, compared to ~ 50% in Arabidopsis. This is likely to hold true in other species that contain a low ratio of Rca-α to Rca-ß isoforms.

Evaluation of remobilization rate, grain yield and antioxidant content of maize in reaction to biochar and humic acid amounts under water deficiency stress

In order to investigate the effect of biochar and humic acid on the rate of remobilization, grain yield and antioxidant content of maize under water deficiency stress, an experiment was conducted as asplit-split plot in the form of a randomized complete blocks design with three replications in Ahvaz, southwest of Iran. The main plot involved water stress with three irrigation levels after depleting 30, 40 and 50% of field capacity, non-stress, moderate stress, and severe stress, respectively, asubplot withbiochar with 2 control levels (no use of biochar) and application of 4 tons in biochar hectares and another subplot having humic acid with 4 control levels (no use of humic acid) and application of 2, 4 and 6 liters per hectare of humic acid. The results showed that the effect of water deficit stress, biochar and humic acid on grain yield, remobilization rate, current photosynthesis, share of current photosynthesis, catalase and superoxide dismutase enzymes was significant at the level of 1% probability. The highest grain yield was related to irrigation treatment after depleting 40% of field capacity and application of 4 tons per hectare of biochar. In total, the use of 4 liters per hectare of humic acid in moderate moisture stress conditions due to its positive role on the growth and hence on the 42% increase in the yield, compared to severe stress treatment and no consumption of humic acid, can be recommended under arid and semi-arid conditions to save water consumption, and reduce the effects of water deficit stress.

Exogenous DCPTA Treatment Increases Mung Bean Yield by Improving Carbon Metabolism Pathway and Up-Regulating Photosynthetic Capacity and Antioxidants.

Mung bean is characterized by having a good edible and medicinal value, while its flowers and pods have low production. Being a tertiary amine, DCPTA [2-(3,4-dichlorophenoxy) triethylamine] substantially regulates the growth and development of crops, maintaining production. Yet it is still limited in terms of the regulation of DCPTA on growth and development, including the yield and sugar metabolism of mung bean. In this study, DCPTA was sprayed at the beginning of mung flowering through a two-season cultivation, to assess its effects on the yield, leaf area per plant, plant height, seed setting rate, photosynthesis, chlorophyll content, and endogenous protective enzymes. Experimental results illustrated that relative to the control (CK), the DCPTA application significantly (p < 0.05) improved the yield of Bailv 11 mung bean, which rose to 6.9% in 2020 and 7.8% in 2021, respectively. This effect positively corresponded to a significant (p<0.05) increase in the number of pods and grains per plant and pod setting rate, but a non-significant difference in 1,000-grain weight. DCPA application also increased the area and fresh weight of leaf, mung height, and its organ dry weight (i.e., leaf, branch, and stem). During plant growth over DCPTA application, the increased activities of SOD, POD, and CAT improved the net photosynthetic rate, stomatal conductance, and transpiration. In addition, transcriptome sequencing further demonstrated that DCPTA treatment significantly (p < 0.05) up-regulated the sucrose synthase, invertase, and fructose kinase in all organs (i.e., leaves, pod skins, and grains) of the plant. In particular, this effect was much greater in the sucrose synthesis (i.e., sucrose content) in leaves. Our study, therefore, concludes that DCPTA application promotes the yield of mung bean via likely enhancing its photosynthetic capacity and sucrose synthase, fructokinase, and beta-fructofuranosidase expression regulation.

Regulation of electron transfer pathways in cytochromes with aromatic amino acids

Natural photosynthetic proteins are based upon on chains of reactions that start with sub-nanosecond light energy conversion into energy of electrical charges and followed by multi-step electron transfer (ET). Attempts to develop artificial enzymes for photosynthesis have been relatively futile due to the difficulty of generating sufficiently fast primary charge separation even with the smallest proteins. Here we report our results on attempts to accelerate ET by placing aromatic redox-active amino acids along the putative path of ET for the E39C mutant of PpcA, a 3-heme cytochrome.

Melatonin increases growth and salt tolerance of Limonium bicolor by improving photosynthetic and antioxidant capacity

BackgroundSoil salinization is becoming an increasingly serious problem worldwide, resulting in cultivated land loss and desertification, as well as having a serious impact on agriculture and the economy. The indoleamine melatonin (N-acetyl-5-methoxytryptamine) has a wide array of biological roles in plants, including acting as an auxin analog and an antioxidant. Previous studies have shown that exogenous melatonin application alleviates the salt-induced growth inhibition in non-halophyte plants; however, to our knowledge, melatonin effects have not been examined on halophytes, and it is unclear whether melatonin provides similar protection to salt-exposed halophytic plants.ResultsWe exposed the halophyte Limonium bicolor to salt stress (300 mM) and concomitantly treated the plants with 5 μM melatonin to examine the effect of melatonin on salt tolerance. Exogenous melatonin treatment promoted the growth of L. bicolor under salt stress, as reflected by increasing its fresh weight and leaf area. This increased growth was caused by an increase in net photosynthetic rate and water use efficiency. Treatment of salt-stressed L. bicolor seedlings with 5 μM melatonin also enhanced the activities of antioxidants (superoxide dismutase [SOD], peroxidase [POD], catalase [CAT], and ascorbate peroxidase [APX]), while significantly decreasing the contents of hydrogen peroxide (H2O2), superoxide anion (O2•−), and malondialdehyde (MDA). To screen for L. bicolor genes involved in the above physiological processes, high-throughput RNA sequencing was conducted. A gene ontology enrichment analysis indicated that genes related to photosynthesis, reactive oxygen species scavenging, the auxin-dependent signaling pathway and mitogen-activated protein kinase (MAPK) were highly expressed under melatonin treatment. These data indicated that melatonin improved photosynthesis, decreased reactive oxygen species (ROS) and activated MAPK-mediated antioxidant responses, triggering a downstream MAPK cascade that upregulated the expression of antioxidant-related genes. Thus, melatonin improves the salt tolerance of L. bicolor by increasing photosynthesis and improving cellular redox homeostasis under salt stress.ConclusionsOur results showed that melatonin can upregulate the expression of genes related to photosynthesis, reactive oxygen species scavenging and mitogen-activated protein kinase (MAPK) of L. bicolor under salt stress, which can improve photosynthesis and antioxidant enzyme activities. Thus melatonin can promote the growth of the species and maintain the homeostasis of reactive oxygen species to alleviate salt stress.

Partial gene sequencing of phosphoenol pyruvate carboxylase (PEPc) gene from Saccharum spp. hybrid CoLk 94184

Phosphoenol pyruvate carboxylase (PEPc), a key enzyme in photosynthesis, an important carboxylase for plant cell metabolism, fixes CO2 in mesophyll cells to form dicarboxylic acids, and being more specific for inorganic carbon, improves the photosynthetic rate of C4 plants. In the present study, partial gene sequencing of PEPc was done using Saccharum spp. hybrid, CoLk 94184 of sugarcane, one of the most popular variety released by ICAR-IISR, Lucknow. Utilizing PEPc of different cultivars of sugarcane species, the primers were designed for PEPc gene and the first ever nucleotide sequence was determined, especially for Saccharum spp. hybrid CoLk 94184 (accession no: MW766370.1). Sequence based phylogenetic tree showed close relatedness between the PEPc sequences for various Saccharum cultivars, thus pointing to little genetic change, while that for sorghum and maize were relatively distantly related. Results may help in understanding the photosynthetic process using sugarcane hybrid specific genes

Rubisco and abiotic stresses in plants: Current assessment

Abiotic stresses are serious environmental factors militating against the production of many crops around the world. The consequence of this, is the difficulty of meeting the demands of the increasing world population. Aside from other negative effects, reduction in photosynthesis is an important feature of abiotic stresses. Abiotic stresses limit photosynthesis in a number of ways. The reduction in ribulose 1, 5-bisphosphate carboxylase/oxygenase (Rubisco) content and activity is one of the paramount ways through which abiotic stresses affect photosynthesis. Rubisco is the CO2 fixing enzyme of photosynthesis and also catalyses the photo-respiratory carbon oxidation. The enzyme has low turnover and also copes with competitive inhibition by O2 . Hence, manipulating the enzyme in order to boost photosynthesis has been the target of scientists, especially in stressed environments. Based on recent studies, the mechanism of the harmful effects of abiotic stresses on Rubisco is examined in this review. In addition, the prevalent ways through which Rubisco can be made to thrive well despite the various abiotic stresses are evaluated. This review paper also outlines practicable approaches to promote existing ways of enhancing Rubisco tolerance to abiotic stresses in order to produce more crops with higher stress resilience.

Chromatin and regulatory differentiation between bundle sheath and mesophyll cells in maize.

C4 plants partition photosynthesis enzymes between the bundle sheath (BS) and the mesophyll (M) cells for the better delivery of CO2 to RuBisCO and to reduce photorespiration. To better understand how C4 photosynthesis is regulated at the transcriptional level, we performed RNA-seq, ATAC-seq, ChIP-seq and Bisulfite-seq (BS-seq) on BS and M cells isolated from maize leaves. By integrating differentially expressed genes with chromatin features, we found that chromatin accessibility coordinates with epigenetic features, especially H3K27me3 modification and CHH methylation, to regulate cell type-preferentially enriched gene expression. Not only the chromatin-accessible regions (ACRs) proximal to the genes (pACRs) but also the distal ACRs (dACRs) are determinants of cell type-preferentially enriched expression. We further identified cell type-preferentially enriched motifs, e.g. AAAG for BS cells and TGACC/T for M cells, and determined their corresponding transcription factors: DOFs and WRKYs. The complex interaction between cis and trans factors in the preferential expression of C4 genes was also observed. Interestingly, cell type-preferentially enriched gene expression can be fine-tuned by the coordination of multiple chromatin features. Such coordination may be critical in ensuring the cell type-specific function of key C4 genes. Based on the observed cell type-preferentially enriched expression pattern and coordinated chromatin features, we predicted a set of functionally unknown genes, e.g. Zm00001d042050 and Zm00001d040659, to be potential key C4 genes. Our findings provide deep insight into the architectures associated with C4 gene expression and could serve as a valuable resource to further identify the regulatory mechanisms present in C4 species.

Integrated physiological and comparative proteomics analysis of contrasting genotypes of pearl millet reveals underlying salt-responsive mechanisms.

Salinity stress poses a significant risk to plant development and agricultural yield. Therefore, elucidation of stress-response mechanisms has become essential to identify salt-tolerance genes in plants. In the present study, two genotypes of pearl millet (Pennisetum glaucum L.) with contrasting tolerance for salinity exhibited differential morpho-physiological and proteomic responses under 150 mM NaCl. The genotype IC 325825 was shown to withstand the stress better than IP 17224. The salt-tolerance potential of IC 325825 was associated with its ability to maintain intracellular osmotic, ionic, and redox homeostasis and membrane integrity under stress. The IC 325825 genotype exhibited a higher abundance of C4 photosynthesis enzymes, efficient enzymatic and non-enzymatic antioxidant system, and lower Na+ /K+ ratio compared with IP 17224. Comparative proteomics analysis revealed greater metabolic perturbation in IP 17224 under salinity, in contrast to IC 325825 that harbored pro-active stress-responsive machinery, allowing its survival and better adaptability under salt stress. The differentially abundant proteins were in silico characterized for their functions, subcellular-localization, associated pathways, and protein-protein interaction. These proteins were mainly involved in photosynthesis/response to light stimulus, carbohydrate and energy metabolism, and stress responses. Proteomics data were validated through expression profiling of the selected genes, revealing a poor correlation between protein abundance and their relative transcript levels. This study has provided novel insights into salt adaptive mechanisms in P. glaucum, demonstrating the power of proteomics-based approaches. The critical proteins identified in the present study could be further explored as potential objects for engineering stress tolerance in salt-sensitive major crops.

Topological Analysis of the Carbon-Concentrating CETCH Cycle and a Photorespiratory Bypass Reveals Boosted CO2-Sequestration by Plants.

Synthetically designed alternative photorespiratory pathways increase the biomass of tobacco and rice plants. Likewise, some in planta–tested synthetic carbon-concentrating cycles (CCCs) hold promise to increase plant biomass while diminishing atmospheric carbon dioxide burden. Taking these individual contributions into account, we hypothesize that the integration of bypasses and CCCs will further increase plant productivity. To test this in silico, we reconstructed a metabolic model by integrating photorespiration and photosynthesis with the synthetically designed alternative pathway 3 (AP3) enzymes and transporters. We calculated fluxes of the native plant system and those of AP3 combined with the inhibition of the glycolate/glycerate transporter by using the YANAsquare package. The activity values corresponding to each enzyme in photosynthesis, photorespiration, and for synthetically designed alternative pathways were estimated. Next, we modeled the effect of the crotonyl-CoA/ethylmalonyl-CoA/hydroxybutyryl-CoA cycle (CETCH), which is a set of natural and synthetically designed enzymes that fix CO₂ manifold more than the native Calvin–Benson–Bassham (CBB) cycle. We compared estimated fluxes across various pathways in the native model and under an introduced CETCH cycle. Moreover, we combined CETCH and AP3-w/plgg1RNAi, and calculated the fluxes. We anticipate higher carbon dioxide–harvesting potential in plants with an AP3 bypass and CETCH–AP3 combination. We discuss the in vivo implementation of these strategies for the improvement of C3 plants and in natural high carbon harvesters.

Discovery of a readily heterologously expressed Rubisco from the deep sea with potential for CO2 capture

Ribulose-1,5-bisphosphate carboxylase/oxygenase (Rubisco), the key CO2-fixing enzyme in photosynthesis, is notorious for its low carboxylation. We report a highly active and assembly-competent Form II Rubisco from the endosymbiont of a deep-sea tubeworm Riftia pachyptila (RPE Rubisco), which shows a 50.5% higher carboxylation efficiency than that of a high functioning Rubisco from Synechococcus sp. PCC7002 (7002 Rubisco). It is a simpler hexamer with three pairs of large subunit homodimers around a central threefold symmetry axis. Compared with 7002 Rubisco, it showed a 3.6-fold higher carbon capture efficiency in vivo using a designed CO2 capture model. The simple structure, high carboxylation efficiency, easy heterologous soluble expression/assembly make RPE Rubisco a ready-to-deploy enzyme for CO2 capture that does not require complex co-expression of chaperones. The chemosynthetic CO2 fixation machinery of chemolithoautotrophs, CO2-fixing endosymbionts, may be more efficient than previously realized with great potential for next-generation microbial CO2 sequestration platforms.

Autotoxic compounds from rhizosphere soil of lanzhou lily extracts: Identification and biological activity

We investigated the autotoxicity of Lanzhou lily rhizosphere soil with different cultivation years. The allelochemicals in such soils were isolated and identified by gas chromatography-mass spectrometry (GC-MS). Based on our earlier studies, the antioxidant 2246 and dioctyl terephthalate were found in lily sick soil were used in pot experiments determine to their autotoxic effects on the Lanzhou lily seedlings growth, photosynthetic parameters and antioxidant enzyme activities in seedling leaves. The content of antioxidant 2246 and dioctyl terephthalate was determined by GC-MS in rhizosphere soil of different cultivation years. The aqueous extracts of Lanzhou lily rhizosphere soil promoted the growth of its own seedlings at 0.2 mg·mL-1, however, the concentrations &gt; 2 mg·mL-1 were inhibitory. The longer the cultivation period (1-yr, 2-yr and 4-yr), the stronger were the inhibitory effects. In rhizosphere soils of 1-yr, 2-yr and 4-yr ; 8,: 15 and 18 compounds were identified, respectively. The identified compounds were camphor, 3-hydroxy-4-methoxy-benzaldehyde, 2,4-bis(1,1- dimethylethyl)-phenol, tributyl phosphate, dibutyl phthalate, antioxidant 2246 and dioctyl terephthalater and these are reported as allelochemicals. The pot experiment results showed that low concentrations of antioxidant 2246 stimulated the seedling growth but high concentrations were inhibitory, while all concentrations of dioctyl terephthalater inhibited the seedling growth. At 100 mg·mL-1, the antioxidant 2246 and dioctyl terephthalate significantly inhibited the photosynthesis and antioxidant enzyme activity in leaves (P &lt;0.05). Furthermore, the content of these two compounds in soils increased with the increase of cultivation years. These results suggested that allelochemicals accumulated in replanted soil contributed to the autotoxicity of Lanzhou lily in rhizosphere soil.

Tunneling and Nonadiabatic Effects on a Proton-Coupled Electron Transfer Model for the Qo Site in Cytochrome bc1.

Cytochrome bc1 is a fundamental enzyme for cellular respiration and photosynthesis. This dimeric protein complex catalyzes a proton-coupled electron transfer (PCET) from the reduced coenzyme-Q substrate (Q) to a bimetallic iron-sulfur cluster in the Qo active site. Herein, we combine molecular dynamics simulations of the complete cytochrome bc1 protein with electronic-structure calculations of truncated models and a semiclassical tunneling theory to investigate the electron-proton adiabaticity of the initial reaction catalyzed in the Qo site. After sampling possible orientations between the Q substrate and a histidine side chain that functions as hydrogen acceptor, we find that a truncated model composed by ubiquinol-methyl and imidazole-iron(III)-sulfide captures the expected changes in oxidation and spin states of the electron donor and acceptor. Diabatic electronic surfaces obtained for this model with multiconfigurational wave function calculations demonstrate that this reaction is electronic nonadiabatic, and proton tunneling is faster than mixing of electronic configurations. These results indicate the formalism that should be used to calculate vibronic couplings and kinetic parameters for the initial reaction in the Qo site of cytochrome bc1. This framework for molecular simulation may also be applied to investigate other PCET reactions in the Q-cycle or in various metalloproteins that catalyze proton translocation coupled to redox processes.

Quantitative proteomic analysis of Xanthoceras sorbifolium Bunge seedlings in response to drought and heat stress

Yellowhorn (Xanthoceras sorbifolium Bunge) is a woody oil species that is widely distributed in northwestern China. To investigate the molecular mechanisms underlying the drought and heat tolerance response of yellowhorn seedlings, changes in protein abundance were analyzed via comparative proteomics. Drought and heat treatment of seedlings was applied in growth chamber, and the leaves were harvested after 7 days of treatment. The total protein was extracted, and comparative proteomic analysis was performed via isobaric tag for relative and absolute quantitation (iTRAQ). The abundance of most of the proteins associated with oxidative phosphorylation, NADH dehydrogenase and superoxide dismutase (SOD) was reduced. The differential proteins associated with photosynthesis enzymes indicated that stress had different effects on photosystem I (PSI) and photosystem II (PSII). After comprehensively analyzing the results, we speculated that drought and heat stress could hinder the synthesis of riboflavin, reducing NADH dehydrogenase content, which might further have an impact on energy utilization. Yellowhorn seedlings relied on Fe–Mn SOD enzymes rather than Cu/Zn SOD enzymes to remove reactive oxygen species (ROS). In addition, heat-shock proteins (HSPs) had significant increase and played a key role in stress response, which could be divided into two categories according to their transcription and translation efficiency. Over all, the results can provide a basis for understanding the molecular mechanism underlying resistance to drought and heat stress in yellowhorn and for subsequent research of posttranslational modification-related omics of key proteins.

Morphological characteristics and production of Xaraes and Zuri grass fertilized with combinations of sulfur and potassium

Sulfur and potassium are essential elements for the development of forages as part of protein synthesis, nutrient transport, activation of photosynthesis enzymes and stomatal activity. Thus, this study aimed was to evaluate the production and morphology of Xaraes grass (Urochloa brizantha cv. Xaraes) and Zuri grass (Megathyrsus maximus cv. Zuri) fertilized with combinations of sulfur (S) and potassium (K) (only S, only K, with S and K, and without both), and identify which of these nutrients should be prioritized in maintenance fertilization. Two experiments were conducted in the greenhouse, in a completely randomized design. The first experiment was utilized the Xaraes grass and the second experiment the Zuri grass. Each experimental plot consisted of 5 dm3 pots with five plants per pot. All treatments were fertilized 200 mg/dm3 per pot with nitrogen. On treatments fertilized with sulfur it were applied ammonium sulfate (21% N and 24% S) as nitrogen supply, thus, it was applied 228 mg/dm3 of sulfur per plot. On treatments without sulfur, the nitrogen source was urea (46% N). The treatments with potassium were fertilized 100 mg/dm3 with potassium, utilizing potassium chloride (58% K2O). The harvest was performed when the plants reached the height of 35 and 75 cm, for xaraes and zuri respectively. For Xaraes grass, the fertilization with S and K in different combinations had a significant effect (P&lt;0.05) on forage mass, leaves number and tillers per pot, individual tiller mass, leaf blade, stem + sheath, and dead material, as well as relative forage mass. For Zuri grass, the combinations of S and K showed statistical difference (P&lt;0.05) just on leaf blade mass and dead material, and the number of leaves per pot, as well as on relative leaf blade mass. Based on the results obtained in the two experiments, the S and K combination in the maintenance fertilization of xaraes grass improves the forage development, however, S does not increase the productive indexes of zuri grass.

Biochar Administration to San Marzano Tomato Plants Cultivated Under Low-Input Farming Increases Growth, Fruit Yield, and Affects Gene Expression.

Biochar is a rich-carbon charcoal obtained by pyrolysis of biomasses, which was used since antiquity as soil amendant. Its storage in soils was demonstrated contributing to abate the effects of climate changes by sequestering carbon, also providing bioenergy, and improving soil characteristics and crop yields. Despite interest in this amendant, there is still poor information on its effects on soil fertility and plant growth. Considerable variation in the plant response has been reported, depending on biomass source, pyrolysis conditions, crop species, and cultivation practices. Due to these conflicting evidences, this work was aimed at studying the effects of biochar from pyrolyzed wood at 550°C, containing 81.1% carbon and 0.91% nitrogen, on growth and yield of tomato plants experiencing low-input farming conditions. San Marzano ecotype from Southern Italy was investigated, due to its renowned quality and adaptability to sustainable farming practices. Biochar administration improved vegetative growth and berry yield, while affecting gene expression and protein repertoire in berries. Different enzymes of carbon metabolism and photosynthesis were over-represented, whereas various stress-responsive and defense proteins were down-represented. Molecular results are here discussed in relation to estimated agronomic parameters to provide a rationale justifying the growth-promoting effect of this soil amendant.

CYP20-3 deglutathionylates 2-CysPRX A and suppresses peroxide detoxification during heat stress.

In plants, growth-defense trade-offs occur because of limited resources, which demand prioritization towards either of them depending on various external and internal factors. However, very little is known about molecular mechanisms underlying their occurrence. Here, we describe that cyclophilin 20-3 (CYP20-3), a 12-oxo-phytodienoic acid (OPDA)-binding protein, crisscrosses stress responses with light-dependent electron reactions, which fine-tunes activities of key enzymes in plastid sulfur assimilations and photosynthesis. Under stressed states, OPDA, accumulates in the chloroplasts, binds and stimulates CYP20-3 to convey electrons towards serine acetyltransferase 1 (SAT1) and 2-Cys peroxiredoxin A (2CPA). The latter is a thiol-based peroxidase, protecting and optimizing photosynthesis by reducing its toxic byproducts (e.g., H2O2). Reduction of 2CPA then inactivates its peroxidase activity, suppressing the peroxide detoxification machinery, whereas the activation of SAT1 promotes thiol synthesis and builds up reduction capacity, which in turn triggers the retrograde regulation of defense gene expressions against abiotic stress. Thus, we conclude that CYP20-3 is a unique metabolic hub conveying resource allocations between plant growth and defense responses (trade-offs), ultimately balancing optimal growth phonotype.

Rubisco accumulation factor 1 (Raf1) plays essential roles in mediating Rubisco assembly and carboxysome biogenesis

Carboxysomes are membrane-free organelles for carbon assimilation in cyanobacteria. The carboxysome consists of a proteinaceous shell that structurally resembles virus capsids and internal enzymes including ribulose 1,5-bisphosphate carboxylase/oxygenase (Rubisco), the primary carbon-fixing enzyme in photosynthesis. The formation of carboxysomes requires hierarchical self-assembly of thousands of protein subunits, initiated from Rubisco assembly and packaging to shell encapsulation. Here we study the role of Rubisco assembly factor 1 (Raf1) in Rubisco assembly and carboxysome formation in a model cyanobacterium, Synechococcus elongatus PCC7942 (Syn7942). Cryo-electron microscopy reveals that Raf1 facilitates Rubisco assembly by mediating RbcL dimer formation and dimer-dimer interactions. Syn7942 cells lacking Raf1 are unable to form canonical intact carboxysomes but generate a large number of intermediate assemblies comprising Rubisco, CcaA, CcmM, and CcmN without shell encapsulation and a low abundance of carboxysome-like structures with reduced dimensions and irregular shell shapes and internal organization. As a consequence, the Raf1-depleted cells exhibit reduced Rubisco content, CO2-fixing activity, and cell growth. Our results provide mechanistic insight into the chaperone-assisted Rubisco assembly and biogenesis of carboxysomes. Advanced understanding of the biogenesis and stepwise formation process of the biogeochemically important organelle may inform strategies for heterologous engineering of functional CO2-fixing modules to improve photosynthesis.

Molecular changes in Mesembryanthemum crystallinum guard cells underlying the C3 to CAM transition.

KEY MESSAGE: The timing and transcriptomic changes during the C3 to CAM transition of common ice plant support the notion that guard cells themselves can shift from C3 to CAM. Crassulacean acid metabolism (CAM) is a specialized type of photosynthesis: stomata close during the day, enhancing water conservation, and open at night, allowing CO2 uptake. Mesembryanthemum crystallinum (common ice plant) is a facultative CAM species that can shift from C3 photosynthesis to CAM under salt or drought stresses. However, the molecular mechanisms underlying the stress induced transition from C3 to CAM remain unknown. Here we determined the transition time from C3 to CAM in M. crystallinum under salt stress. In parallel, single-cell-type transcriptomic profiling by 3'-mRNA sequencing was conducted in isolated stomatal guard cells to determine the molecular changes in this key cell type during the transition. In total, 495 transcripts showed differential expression between control and salt-treated samples during the transition, including 285 known guard cell genes, seven CAM-related genes, 18 transcription factors, and 185 other genes previously not found to be expressed in guard cells. PEPC1 and PPCK1, which encode key enzymes of CAM photosynthesis, were up-regulated in guard cells after seven days of salt treatment, indicating that guard cells themselves can shift from C3 to CAM. This study provides important information towards introducing CAM stomatal behavior into C3 crops to enhance water use efficiency.

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