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Recognition of the untapped potential of photosynthesis to improve crop yields has spurred research to identify targets for breeding. The CO2-fixing enzyme Rubisco is characterized by a number of inefficiencies, and frequently limits carbon assimilation at the top of the canopy, representing a clear target for wheat improvement. Two bread wheat lines with similar genetic backgrounds and contrasting in vivo maximum carboxylation activity of Rubisco per unit leaf nitrogen (V c,max,25/ N area) determined using high-throughput phenotyping methods were selected for detailed study from a panel of 80 spring wheat lines. Detailed phenotyping of photosynthetic traits in the two lines using glasshouse-grown plants showed no difference in V c,max,25/ N area determined directly via in vivo and in vitro methods. Detailed phenotyping of glasshouse-grown plants of the 80 wheat lines also showed no correlation between photosynthetic traits measured via high-throughput phenotyping of field-grown plants. Our findings suggest that the complex interplay between traits determining crop productivity and the dynamic environments experienced by field-grown plants needs to be considered in designing strategies for effective wheat crop yield improvement when breeding for particular environments. [ABSTRACT FROM AUTHOR]

Improvement of photosynthetic traits in crops to increase yield potential and crop resilience has recently become a major breeding target. Synthetic biology and genetic technologies offer unparalleled opportunities to create new genetics for photosynthetic traits driven by existing fundamental knowledge. However, large 'gene bank' collections of germplasm comprising historical collections of crop species and their relatives offer a wealth of opportunities to find novel allelic variation in the key steps of photosynthesis, to identify new mechanisms and to accelerate genetic progress in crop breeding programmes. Here we explore the available genetic resources in food and fibre crops, strategies to selectively target allelic variation in genes underpinning key photosynthetic processes, and deployment of this variation via gene editing in modern elite material. [ABSTRACT FROM AUTHOR]

Future climate scenarios suggest that crop plants will experience environmental changes capable of affecting their productivity. Among the most harmful environmental stresses is drought, defined as a total or partial lack of water availability. It is essential to study and understand both the damage caused by drought on crop plants and the mechanisms implemented to tolerate the stress. In this study, we focused on four cultivars of tomato, an economically important crop in the Mediterranean basin. We investigated the biochemical mechanisms of plant defense against drought by focusing on proteins specifically involved in this stress, such as osmotin, dehydrin, and aquaporin, and on proteins involved in the general stress response, such as HSP70 and cyclophilins. Since sugars are also known to act as osmoprotectants in plant cells, proteins involved in sugar metabolism (such as RuBisCO and sucrose synthase) were also analyzed. The results show crucial differences in biochemical behavior among the selected cultivars and highlight that the most tolerant tomato cultivars adopt quite specific biochemical strategies such as different accumulations of aquaporins and osmotins. The data set also suggests that RuBisCO isoforms and aquaporins can be used as markers of tolerance/susceptibility to drought stress and be used to select tomato cultivars within breeding programs. [ABSTRACT FROM AUTHOR]

Cyanobacteria evolved an inorganic carbon-concentrating mechanism (CCM) to perform effective oxygenic photosynthesis and prevent photorespiratory carbon losses. This process facilitates the acclimation of cyanobacteria to various habitats, particularly in CO2-limited environments. To date, there is limited information on the CCM of thermophilic cyanobacteria whose habitats limit the solubility of inorganic carbon. Here, genome-based approaches were used to identify the molecular components of CCM in 17 well-described thermophilic cyanobacteria. These cyanobacteria were from the genus Leptodesmis , Leptolyngbya , Leptothermofonsia , Thermoleptolyngbya , Thermostichus , and Thermosynechococcus. All the strains belong to β-cyanobacteria based on their β-carboxysome shell proteins with 1B form of Rubisco. The diversity in the Ci uptake systems and carboxysome composition of these thermophiles were analyzed based on their genomic information. For Ci uptake systems, two CO2 uptake systems (NDH-13 and NDH-14) and BicA for HCO3– transport were present in all the thermophilic cyanobacteria, while most strains did not have the Na+/HCO3– Sbt symporter and HCO3– transporter BCT1 were absent in four strains. As for carboxysome, the β-carboxysomal shell protein, ccmK2, was absent only in Thermoleptolyngbya strains, whereas ccmK3/K4 were absent in all Thermostichus and Thermosynechococcus strains. Besides, all Thermostichus and Thermosynechococcus strains lacked carboxysomal β-CA, ccaA, the carbonic anhydrase activity of which may be replaced by ccmM proteins as indicated by comparative domain analysis. The genomic distribution of CCM-related genes was different among the thermophiles, suggesting probably distinct expression regulation. Overall, the comparative genomic analysis revealed distinct molecular components and organization of CCM in thermophilic cyanobacteria. These findings provided insights into the CCM components of thermophilic cyanobacteria and fundamental knowledge for further research regarding photosynthetic improvement and biomass yield of thermophilic cyanobacteria with biotechnological potentials. [ABSTRACT FROM AUTHOR]

Abstract G protein γ subunit qPE9-1 plays multiple roles in rice growth and development. However, the role of qPE9-1 in rice exposed to elevated carbon dioxide concentration (eCO2) is unknown. Here, we investigated its role in the regulation of rice growth under eCO2 conditions using qPE9-1 overexpression (OE) lines, RNAi lines and corresponding WT rice. Compared to atmospheric carbon dioxide concentration (aCO2), relative expression of qPE9-1 in rice leaf was approximately tenfold higher under eCO2. Under eCO2, the growth of WT and qPE9-1-overexpressing rice was significantly higher than under aCO2. Moreover, there was no significant effect of eCO2 on the growth of qPE9-1 RNAi lines. Furthermore, WT and qPE9-1-overexpressing rice showed higher net photosynthetic rate and carbohydrate content under eCO2 than under aCO2. Moreover, the relative expression of some photosynthesis related genes in WT, but not in RNAi3 line, showed significant difference under eCO2 in RNA-seq analysis. Compared to WT and RNAi lines, the rbcL gene expression and Rubisco content of rice leaves in qPE9-1-overexpressors were higher under eCO2. Overall, these results suggest that qPE9-1 is involved in rice adaptation under elevated CO2 concentration by regulating leaf photosynthesis via moderating rice photosynthetic light reaction and Rubisco content.

Rubisco is the key enzyme responsible for photosynthetic CO2 fixation. Previously, we succeeded to increase the catalytic rate (kcat) of Rubisco by the overexpression of sorghum C4-type Rubisco small subunit (RbcS) in rice. These Rubisco are present as a chimera of sorghum RbcS and rice RbcS. In this study, we introduced an RNAi construct for rice RbcS into a sorghum RbcS overexpression line (SS10). Obtained double-transgenic lines iSS4 and iSS5 dominantly expressed sorghum RbcS and accumulated 35% and 20% of Rubisco content of non-transgenic rice (WT), respectively. Rubisco in these double-transgenic lines showed higher kcat and Km for CO2 than WT and SS10. Moreover, iSS4 showed higher photosynthetic nitrogen use efficiency under high CO2 conditions. These results demonstrate the usefulness of C4-type RbcS to improve photosynthetic performance under elevated CO2 conditions.

Abstract The oxygenase activity of Ribulose-1,5-bisphosphate carboxylase/oxygenase (Rubisco) converts ribulose-1,5-bisphosphate (RuBP) into 2-phosphoglycolate, which in turn channels into photorespiration, resulting in carbon and energy loss in higher plants. We observed that glycolate can be accumulated extracellularly when two genes encoding the glycolate dehydrogenase of cyanobacteria Synechocystis sp. PCC 6803 were inactivated. This inspired us to explore the oxygenase function of Rubisco for production of glycolate, an important industrial chemical, from CO2 by engineered cyanobacteria. Since the oxygenase activity of Rubisco is generally low in CO2-rich carboxysome of cyanobacteria, we introduced Form II Rubisco, which cannot be assembled in carboxysome, into the cytoplasm of cyanobacteria. Heterologous expression of a Form II Rubisco from endosymbiont of tubeworm Riftia pachyptila (RPE Rubisco) significantly increased glycolate production. We show that the RPE Rubisco is expressed in the cytoplasm. Glycolate production increased upon addition of NaHCO3 but decreased upon supplying CO2. The titer of glycolate reached 2.8 g/L in 18 days, a 14-fold increase compared with the initial strain with glycolate dehydrogenase inactivated. This is also the highest glycolate titer biotechnologically produced from CO2 ever reported. Photosynthetic production of glycolate demonstrated the oxygenase activity of Form II Rubisco can be explored for production of chemicals from CO2.

Multi-walled carbon nanotubes (MWCNTs) are cylindrical rolled sheets with different applications in plant biotechnology due to their physicochemical characteristics. This study aimed to evaluate the in vitro physiological and biochemical responses of stevia (Stevia rebaudiana Bertoni) exposure to different concentrations of MWCNTs during multiplication phase in liquid medium using temporary immersion bioreactors. The MWCNTs were characterized regarding their properties by transmission electron microscope. Different treatments (0, 25, 50 and 100 mg/L) of MWCNTs were added to Murashige and Skoog semisolid culture medium at the proliferation phase. At 30 d of in vitro multiplication, the number of shoots, shoot length, number of leaves, dry matter, and antioxidant capacity were evaluated, as well as total chlorophyll, carotenoid, macro and micronutrient, phenolic, Phosphoenolpyruvate (PEP) and Rubisco contents. The results showed that adding MWCNTs at low doses (25 and 50 mg/L MWCNT) led to an increase in the in vitro multiplication rate and a higher number of leaves, while, at the highest dose (100 mg/L MWCNT), there was an inhibition of these variables. MWCNT treatments had an effect on Mg, Fe and C elements. The accumulation of photosynthetic pigments was associated with a higher PEP content in MWCNT treatments. Phenolic content increased at a concentration of 100 mg/L MWCNT, while antioxidant capacity increased at a concentration of 50 mg/L MWCNT. In conclusion, the usage of low MWCNT doses could be used to induce a hormetic response and to promote photosynthesis during in vitro propagation of stevia. • Multi-walled carbon nanotubes (MWCNTs) promote the in vitro multiplication rate of stevia shoots. • MWCNTs increase chlorophyll and carotenoid content. • MWCNTs increase phenolic content and antioxidant capacity. • Temporary immersion bioreactors and MWCNTs are an alternative for in vitro propagation of stevia. [ABSTRACT FROM AUTHOR]

Carbon capture technology is currently considered one of the promising technologies to mitigate atmospheric CO 2 concentration. CO 2 capture and utilization (CCU) captures anthropogenic waste CO 2 and valorizes it into useful products, supporting a circular transition pathway towards carbon neutrality. Unfortunately, the thermodynamic stability of CO 2 requires a high-energy input for its conversion, resulting in processes with a net positive carbon footprint. The incorporation of enzymes as biocatalysts in a process is attractive, as it facilitates CO 2 conversion under ambient conditions. In Nature, the conversion of CO 2 into organic compounds is done through photosynthesis, using an enzyme called ribulose-1,5-biphosphate carboxylase/oxygenase (RuBisCO). RuBisCO plays a central role in the natural assimilation of CO 2 , making it the enzyme chosen in Nature upon which all life forms depend. However, the slow carboxylation rate of RuBisCO (1–10/s) has caused it to be overlooked by faster enzymes such as carbonic anhydrase (CA), which has a carboxylation rate of 106/s. Despite this, RuBisCO has a rate enhancement of 108 to 1010 times higher than CA. Thus, this review aims to take a closer look at RuBisCO and examine its potential in CCU. Various aspects are considered, such as RuBisCO's performance in comparison to other enzymes, approaches to overcome its limitations, its applications and implications in CCU, the valuable chemicals that can be derived from it, recent developments in RuBisCO-integrated processes, and its economic and environmental considerations. Through this, RuBisCO's potential as one of the key enzymes in CCU will be explored. [ABSTRACT FROM AUTHOR]

Nitrogen is one of the most important nutrients for plant growth and a major constituent of proteins that regulate photosynthetic and respiratory processes. However, a comprehensive global analysis of nitrogen allocation in leaves for major processes with respect to different plant functional types (PFTs) is currently lacking. This study integrated observations from global databases with photosynthesis and respiration models to determine plant-functional-type-specific allocation patterns of leaf nitrogen for photosynthesis (Rubisco, electron transport, light absorption) and respiration (growth and maintenance), and by difference from observed total leaf nitrogen, an unexplained "residual" nitrogen pool. Based on our analysis, crops partition the largest fraction of nitrogen to photosynthesis (57%) and respiration (5%) followed by herbaceous plants (44% and 4%). Tropical broadleaf evergreen trees partition the least to photosynthesis (25%) and respiration (2%) followed by needle-leaved evergreen trees (28% and 3%). In trees (especially needle-leaved evergreen and tropical broadleaf evergreen trees) a large fraction (70% and 73%, respectively) of nitrogen was not explained by photosynthetic or respiratory functions. Compared to crops and herbaceous plants, this large residual pool is hypothesized to emerge from larger investments in cell wall proteins, lipids, amino acids, nucleic acid, CO2 fixation proteins (other than Rubisco), secondary compounds, and other proteins. Our estimates are different from previous studies due to differences in methodology and assumptions used in deriving nitrogen allocation estimates. Unlike previous studies, we integrate and infer nitrogen allocation estimates across multiple PFTs, and report substantial differences in nitrogen allocation across different PFTs. The resulting pattern of nitrogen allocation provides insights on mechanisms that operate at a cellular scale within leaves, and can be integrated with ecosystem models to derive emergent properties of ecosystem productivity at local, regional, and global scales.

Light quality plays an essential role in setting plant structural and functional traits, including antioxidant compounds. This paper aimed to assess how manipulating the light spectrum during growth may regulate the photosynthetic activity and fruit bioactive compound synthesis in Solanum lycopersicum L. cv. ‘Microtom’ to improve plant physiological performance and fruit nutritional value. Plants were cultivated under three light quality regimes: red-green-blue LEDs (RGB), red-blue LEDs (RB) and white fluorescent lamps (FL), from sowing to fruit ripening. Leaf functional traits, photosynthetic efficiency, Rubisco and D1 protein expression, and antioxidant production in fruits were analyzed. Compared to FL, RGB and RB regimes reduced height and increased leaf number and specific leaf area, enhancing plant dwarf growth. The RGB regime improved photosynthesis and stomatal conductance despite lower biomass, favoring Rubisco synthesis and carboxylation rate than RB and FL regimes. The RB light produced plants with fewer flowers and fruits with a lower ascorbic acid amount but the highest polyphenol content, antioxidant capacity and SOD and CAT activities. Our data indicate that the high percentage of the green wavelength in the RGB regime promoted photosynthesis and reduced plant reproductive capacity compared to FL and RB. Conversely, the RB regime was the best in favoring the production of health-promoting compounds in tomato berries.

Cyanobacteria evolved an inorganic carbon-concentrating mechanism (CCM) to perform effective oxygenic photosynthesis and prevent photorespiratory carbon losses. This process facilitates the acclimation of cyanobacteria to various habitats, particularly in CO2-limited environments. To date, there is limited information on the CCM of thermophilic cyanobacteria whose habitats limit the solubility of inorganic carbon. Here, genome-based approaches were used to identify the molecular components of CCM in 17 well-described thermophilic cyanobacteria. These cyanobacteria were from the genus Leptodesmis, Leptolyngbya, Leptothermofonsia, Thermoleptolyngbya, Thermostichus, and Thermosynechococcus. All the strains belong to β-cyanobacteria based on their β-carboxysome shell proteins with 1B form of Rubisco. The diversity in the Ci uptake systems and carboxysome composition of these thermophiles were analyzed based on their genomic information. For Ci uptake systems, two CO2 uptake systems (NDH-13 and NDH-14) and BicA for HCO3– transport were present in all the thermophilic cyanobacteria, while most strains did not have the Na+/HCO3– Sbt symporter and HCO3– transporter BCT1 were absent in four strains. As for carboxysome, the β-carboxysomal shell protein, ccmK2, was absent only in Thermoleptolyngbya strains, whereas ccmK3/K4 were absent in all Thermostichus and Thermosynechococcus strains. Besides, all Thermostichus and Thermosynechococcus strains lacked carboxysomal β-CA, ccaA, the carbonic anhydrase activity of which may be replaced by ccmM proteins as indicated by comparative domain analysis. The genomic distribution of CCM-related genes was different among the thermophiles, suggesting probably distinct expression regulation. Overall, the comparative genomic analysis revealed distinct molecular components and organization of CCM in thermophilic cyanobacteria. These findings provided insights into the CCM components of thermophilic cyanobacteria and fundamental knowledge for further research regarding photosynthetic improvement and biomass yield of thermophilic cyanobacteria with biotechnological potentials.

Abstract Background Mixotrophy can confer a higher growth rate than the sum of photoautotrophy and heterotrophy in many microalgal species. Thus, it has been applied to biodiesel production and wastewater utilization. However, its carbon and energy metabolic mechanism is currently poorly understood. Results To elucidate underlying carbon and energy metabolic mechanism of mixotrophy, Chromochloris zofingiensis was employed in the present study. Photosynthesis and glucose metabolism were found to operate in a dynamic balance during mixotrophic cultivation, the enhancement of one led to the lowering of the other. Furthermore, compared with photoautotrophy, non-photochemical quenching and photorespiration, considered by many as energy dissipation processes, were significantly reduced under mixotrophy. Comparative transcriptome analysis suggested that the intermediates of glycolysis could directly enter the chloroplast and replace RuBisCO-fixed CO2 to provide carbon sources for chloroplast organic carbon metabolism under mixotrophy. Therefore, the photosynthesis rate-limiting enzyme, RuBisCO, was skipped, allowing for more efficient utilization of photoreaction-derived energy. Besides, compared with heterotrophy, photoreaction-derived ATP reduced the need for TCA-derived ATP, so the glucose decomposition was reduced, which led to higher biomass yield on glucose. Based on these results, a mixotrophic metabolic mechanism was identified. Conclusions Our results demonstrate that the intermediates of glycolysis could directly enter the chloroplast and replace RuBisCO-fixed CO2 to provide carbon for photosynthesis in mixotrophy. Therefore, the photosynthesis rate-limiting enzyme, RuBisCO, was skipped in mixotrophy, which could reduce energy waste of photosynthesis while promote cell growth. This finding provides a foundation for future studies on mixotrophic biomass production and photosynthetic metabolism.

Abstract Background The global demand for functional proteins is extensive, diverse, and constantly increasing. Medicine, agriculture, and industrial manufacturing all rely on high-quality proteins as major active components or process additives. Historically, these demands have been met by microbial bioreactors that are expensive to operate and maintain, prone to contamination, and relatively inflexible to changing market demands. Well-established crop cultivation techniques coupled with new advancements in genetic engineering may offer a cheaper and more versatile protein production platform. Chloroplast-engineered plants, like tobacco, have the potential to produce large quantities of high-value proteins, but often result in engineered plants with mutant phenotypes. This technology needs to be fine-tuned for commercial applications to maximize target protein yield while maintaining robust plant growth. Results Here, we show that a previously developed Nicotiana tabacum line, TetC-cel6A, can produce an industrial cellulase at levels of up to 28% of total soluble protein (TSP) with a slight dwarf phenotype but no loss in biomass. In seedlings, the dwarf phenotype is recovered by exogenous application of gibberellic acid. We also demonstrate that accumulating foreign protein represents an added burden to the plants’ metabolism that can make them more sensitive to limiting growth conditions such as low nitrogen. The biomass of nitrogen-limited TetC-cel6A plants was found to be as much as 40% lower than wildtype (WT) tobacco, although heterologous cellulase production was not greatly reduced compared to well-fertilized TetC-cel6A plants. Furthermore, cultivation at elevated carbon dioxide (1600 ppm CO2) restored biomass accumulation in TetC-cel6A plants to that of WT, while also increasing total heterologous protein yield (mg Cel6A plant−1) by 50–70%. Conclusions The work reported here demonstrates that well-fertilized tobacco plants have a substantial degree of flexibility in protein metabolism and can accommodate considerable levels of some recombinant proteins without exhibiting deleterious mutant phenotypes. Furthermore, we show that the alterations to protein expression triggered by growth at elevated CO2 can help rebalance endogenous protein expression and/or increase foreign protein production in chloroplast-engineered tobacco.

Protein extracts from green leaves, consisting of mainly RuBisCO, have potential applications for human consumption. However, their properties are highly influenced by the extraction process applied. This work assessed the impact of the extraction method on the solubility profile and emulsifying properties of two protein concentrates from perennial ryegrass, obtained using two different approaches. These two methods differed in the application (or absence) of a heating step (50 °C for 30 min) to improve the removal of chloroplast material. The functionality of both protein concentrates was highly influenced by the isoelectric point of the proteins. The extract containing less residual chloroplast material showed greater solubility, up to 40 % more, and superior emulsifying capacity and stability, at non-acidic pH, similar to WPI. At pH above 6, the protein extract which contained residual chloroplast material was significantly less soluble, which limited its performance as emulsifier. Therefore, the extraction process critically impacted the final product's functionality. This work demonstrated that effectively removing chloroplast material is essential to achieve targeted techno-functional properties of an ingredient and the food product in which it is used. [Display omitted] • Protein concentrates from perennial ryegrass were explored as food emulsifiers. • Extraction method impacted the functional properties of the proteins. • Heating step during protein extraction improved emulsification properties. • Interfacial properties of both concentrates were highly influenced by the pH. • Balance between extraction yield and functional properties should be considered. [ABSTRACT FROM AUTHOR]

Plants being the primary producers are the driving force of sustenance of the living world. However, plants derive the energy from Sun, to produce their own food via the process of photosynthesis. Depending on the plant's different photosynthetic pathways, it can be divided into prominent physiological groups. The main groups are C3 and C4. C4 plants are able to produce the highest rate of biological energy from solar energy. C4 plants are basically another form of C3 plants to survive in a hot and arid climate. The major drastic problem is photorespiration in C3 plants, which reduces the production of biological energy from solar energy. Therefore, C4 plants reduce photorespiration by concentrating the CO 2 concentration near RUBISCO. Hence, a long-standing goal is the conversion of C3 crops to C4 photosynthetic crops by employing different bio-engineering technologies and pathways. In this era of synthetic biology, the conversion has remained very tough owing to its specialized anatomy and biochemical cycles. This study reviewed the basic molecular concepts, the main requirements of C3-C4 conversion and advances in system biology methods. This review further emphasises on the various possible theoretical and practical knowledge with bio-engineering approaches and genetic engineering tools. Moreover, this study discussed different approaches and various future prospects in this field. [ABSTRACT FROM AUTHOR]

Microplastics (MPs) pollution and their impact on plants have become a global threat, but their effect at the molecular level remains scarce. This study aims to gain insight into the effects of polyvinylchloride microplastic (PVC-MP) on tomato plants at the genetic and protein levels. In this study, we found that increasing concentrations of PVC-MP (2.5, 5,7.5, and 10% w/w) in the soil did not cause any phytotoxic (chlorosis or necrosis) symptoms but it did result in a dose-dependent reduction in plant growth-related parameters, such as height, leaf area, stem diameter, and plant fresh and dry weight. Additionally, the number of secondary roots was reduced while the primary roots were elongated. Furthermore, PVC-MP also caused a significant decrease in light-harvesting pigments chlorophylls, and carotenoids while increasing the level of reactive oxygen species (ROS) and lipid peroxidation in plants. Microscopic analysis of the roots revealed the uptake of PVC-MP of size less than 10 μm. Micro- and macro-element analysis showed changes in concentrations of Ca, Cu, Fe, Mg, Mn, Ni, and Zn, upon PVC-MP exposure. Results from western blotting and q-PCR showed that higher doses of PVC-MP significantly reduced the CO 2 -fixing enzyme RuBisCO and D1 proteins of PSII at both protein and transcript levels. These findings suggest that lower levels of light-harvesting pigments, D1 protein, RuBisCO, and modulation of nutrient absorption are among the factors responsible for growth suppression in tomato plants upon exposure to PVC-MP. As tomato plants are economically significant crops, an increase in PVC-MP in agricultural fields may have a detrimental influence on crop production, resulting in economic loss. [Display omitted] • PVC-MP reduced photo-pigment content whereas increased ROS and lipid peroxidation. • PVC-MP damaged the secondary roots while elongating the primary roots. • Microplastic size less than 10 μm can enter into the roots of tomato plants. • PVC-MP reduced RuBisCO enzyme and D1 protein at both protein and transcript levels. • Absorption of certain nutrients was modulated in tomato plants upon PVC-MP exposure. [ABSTRACT FROM AUTHOR]

Form I rubiscos evolved in Cyanobacteria ≥ 2.5 billion years ago and are enzymatically unique due to the presence of small subunits (RbcS) capping both ends of an octameric large subunit (RbcL) rubisco assembly to form a hexadecameric (L8S8) holoenzyme. Although RbcS was previously thought to be integral to Form I rubisco stability, the recent discovery of a closely related sister clade of octameric rubiscos (Form I’; L8) demonstrates that the L8 complex can assemble without small subunits (Banda et al. 2020). Rubisco also displays a kinetic isotope effect (KIE) where the 3PG product is depleted in 13C relative to 12C. In Cyanobacteria, only two Form I KIE measurements exist, making interpretation of bacterial carbon isotope data difficult. To aid comparison, we measured in vitro the KIEs of Form I’ (Candidatus Promineofilum breve) and Form I (Synechococcus elongatus PCC 6301) rubiscos and found the KIE to be smaller in the L8 rubisco (16.25 ± 1.36‰ vs. 22.42 ± 2.37‰, respectively). Therefore, while small subunits may not be necessary for protein stability, they may affect the KIE. Our findings may provide insight into the function of RbcS and allow more refined interpretation of environmental carbon isotope data.

The overexpression of the RuBisCO (rbc) gene has recently become an achievable strategy for increasing cyanobacterial biomass and overcoming the biocompound production restriction. We successfully constructed two rbc-overexpressing Synechocystis sp. PCC 6803 strains (OX), including a strain overexpressing a large subunit of RuBisCO (OXrbcL) and another strain overexpressing all large, chaperone, and small subunits of RuBisCO (OXrbcLXS), resulting in higher and faster growth than wild type under sodium bicarbonate supplementation. This increased biomass of OX strains significantly contributed to the higher polyhydroxybutyrate (PHB) production induced by nutrient-deprived conditions, in particular nitrogen (N) and phosphorus (P). As a result of higher PHB contents in OX strains occurring at days 7 and 9 of nutrient deprivation, this enhancement was apparently made possible by cells preferentially maintaining their internal lipids while accumulating less glycogen. The OXrbcLXS strain, with the highest level of PHB at about 39 %w/dry cell weight (DCW) during 7 days of BG11-NP treatment, contained a lower glycogen level (31.9 %w/DCW) than wild type control (40 %w/DCW). In contrast, the wild type control strain exposed to N- and NP-stresses tended to retain lipid levels and store more glycogen than PHB. In this model, we, for the first time, implemented a RuBisCO-overexpressing cyanobacterial factory for overproducing PHB, destined for biofuel and biomaterial biotechnology.

A Chlamydomonas reinhardtii RuBisCO-less mutant, Δ rbcL , was used to study carbohydrate metabolism without fixation of atmospheric carbon. The regulatory mechanism(s) that control linear electron flow, known as "photosynthetic control," are amplified in Δ rbcL at the onset of illumination. With the aim to understand the metabolites that control this regulatory response, we have correlated the kinetics of primary carbon metabolites to chlorophyll fluorescence induction curves. We identify that Δ rbcL in the absence of acetate generates adenosine triphosphate (ATP) via photosynthetic electron transfer reactions. Also, metabolites of the Calvin Benson Bassham (CBB) cycle are responsive to the light. Indeed, ribulose 1,5-bisphosphate (RuBP), the last intermediate before carboxylation by Ribulose-1,5-bisphosphate carboxylase-oxygenase, accumulates significantly with time, and CBB cycle intermediates for RuBP regeneration, dihydroxyacetone phosphate (DHAP), pentose phosphates and ribose-5-phosphate (R5P) are rapidly accumulated in the first seconds of illumination, then consumed, showing that although the CBB is blocked, these enzymes are still transiently active. In opposition, in the presence of acetate, consumption of CBB cycle intermediates is strongly diminished, suggesting that the link between light and primary carbon metabolism is almost lost. Phosphorylated hexoses and starch accumulate significantly. We show that acetate uptake results in heterotrophic metabolism dominating phototrophic metabolism, with glyoxylate and tricarboxylic acid (TCA) cycle intermediates being the most highly represented metabolites, specifically succinate and malate. These findings allow us to hypothesize which metabolites and metabolic pathways are relevant to the upregulation of processes like cyclic electron flow that are implicated in photosynthetic control mechanisms. [ABSTRACT FROM AUTHOR]

Fragility of photosystem I has been observed in transgenic rice plants that overproduce Rubisco activase (RCA). In this study, we examined the effects of RCA overproduction on the sensitivity of PSI to photoinhibition in three lines of plants overexpressing RCA (RCA -ox). In all the RCA -ox plants the quantum yield of PSI [ Y (I)] decreased whilst in contrast the quantum yield of acceptor-side limitation of PSI [ Y (NA)] increased, especially under low light conditions. In the transgenic line with the highest RCA content (RCA -ox 1), the quantum yield of PSII [ Y (II)] and CO2 assimilation also decreased under low light. When leaves were exposed to high light (2000 μmol photon m−2 s−1) for 60 min, the maximal activity of PSI (P m) drastically decreased in RCA -ox 1. These results suggested that overproduction of RCA disturbs PSI electron transport control, thus increasing the susceptibility of PSI to photoinhibition. When flavodiiron protein (FLV), which functions as a large electron sink downstream of PSI, was expressed in the RCA -ox 1 background (RCA - FLV), PSI and PSII parameters, and CO2 assimilation were recovered to wild-type levels. Thus, expression of FLV restored the robustness of PSI in RCA -ox plants. [ABSTRACT FROM AUTHOR]

In tropical no-till systems, applying lime (L) and phosphogypsum (PG) on the soil surface may be a potential strategy for reducing soil acidification and improving soybean root growth, thereby enhancing plant nutrition and physiological responses and, in turn, crop resistance to dry spells. This study evaluated the impact of long-term (17 years) surface soil amendment on soil fertility and soybean root development, nutrition, gas exchange, carbon and antioxidant enzyme activity, and grain yield in a tropical region subject to dry spells. The treatments consisted of the following long-term soil amendments: control (no soil amendment); L alone; PG alone; and L + PG (LPG). Liming, especially when combined with PG, improved soil fertility, as evidenced by increases in pH and P, Ca2+, and Mg2+ levels throughout the soil profile, but reduced Al3+ and micronutrients (Fe, Mn, Cu, and Zn). The improvements in soil fertility were associated with increased root development throughout the profile. Long-term application of LPG reduced the negative impacts of dry spells on pigment concentrations, gas exchange, Rubisco and sucrose synthase activities and antioxidant metabolism, and increased soybean grain yield. Our results reveal that long-term application of LPG is an important approach for increasing the vertical movement of cationic bases and roots in no-till systems to improve soybean nutrition. Long-term amendment with LPG enhanced both carbon and antioxidant metabolism in soybean plants, resulting in higher soybean grain yield, despite the predisposition of this tropical region to dry spells. [ABSTRACT FROM AUTHOR]

Functional and active Rubisco is essential for CO2 fixation and is a primary target for engineering approaches to increasing crop yields. However, the assembly and maintenance of active Rubisco are dependent on the coordinated biosynthesis of at least 11 nuclear-encoded proteins, termed the 'Rubiscosome'. Using publicly available gene expression data for wheat (Triticum aestivum L.), we show that the expression of Rubiscosome genes is balanced across the three closely related subgenomes that form the allohexaploid genome. Each subgenome contains a near complete set of homoeologous genes and contributes equally to overall expression, both under optimal and under heat stress conditions. The expression of the wheat thermo-tolerant Rubisco activase isoform 1β increases under heat stress and remains balanced across the subgenomes, albeit with a slight shift towards greater contribution from the D subgenome. The findings show that the gene copies in all three subgenomes need to be accounted for when designing strategies for crop improvement. [ABSTRACT FROM AUTHOR]

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The present human population is more than three times what it was in 1950. With that, there is an increasing demand for the consumption of fossil fuels for various anthropogenic activities. This consumption is the major source of carbon dioxide emission causing greenhouse effects leading to global warming. The dependency on fossil fuels around the globe is such that it would be hard to move away from it any time soon. Hence, we must work on strategies to improve carbon dioxide fixation as we are making advancements in clean energy technology. This review explores the natural carbon dioxide fixation pathways in plants and various microorganisms and discusses their limitations and alternative strategies. It explains what necessitates the exploration of synthetic pathways and discusses strategies and matrices to consider while evaluating various pathways. This review also discusses the recent breakthroughs in the field of nanosciences that could accelerate chemical methods of carbon dioxide fixation. [ABSTRACT FROM AUTHOR]

Background: Improvement in photosynthesis is one of the most promising approaches to increase grain yields. Transgenic rice plants overproducing Rubisco by 30% (RBCS-sense rice plants) showed up to 28% increase in grain yields under sufficient nitrogen (N) fertilization using an isolated experimental paddy field (Yoon et al. in Nat Food 1:134–139, 2020). The plant N contents above-ground sections and Rubisco contents of the flag leaves were higher in the RBCS-sense plants than in the wild-type rice plants during the ripening period, which may be reasons for the increased yields. However, some imprecise points were left in the previous research, such as contributions of photosynthesis of leaves below the flag leaves to the yield, and maintenance duration of high photosynthesis of RBCS-sense rice plants during ripening periods. Result: In this research, the photosynthetic capacity and canopy architecture were analyzed to explore factors for the increased yields of RBCS-sense rice plants. It was found that N had already been preferentially distributed into the flag leaves at the early ripening stage, contributing to maintaining higher Rubisco content levels in the enlarged flag leaves and extending the lifespan of the flag leaves of RBCS-sense rice plants throughout ripening periods under sufficient N fertilization. The higher amounts of Rubisco also improved the photosynthetic activity in the flag leaves throughout the ripening period. Although the enlarged flag leaves of the RBCS-sense rice plants occupied large spatial areas of the uppermost layer in the canopy, no significant prevention of light penetration to leaves below the flag leaves was observed. Additionally, since the CO2 assimilation rates of lower leaves between wild-type and RBCS-sense rice plants were the same at the early ripening stage, the lower leaves did not contribute to an increase in yields of the RBCS-sense rice plants. Conclusion: We concluded that improvements in the photosynthetic capacity by higher leaf N and Rubisco contents, enlarged leaf area and extended lifespan of flag leaves led to an increase in grain yields of RBCS-sense rice plants grown under sufficient N fertilization. [ABSTRACT FROM AUTHOR]

Giant clams are ecologically and economically relevant reef inhabitants that host photosynthetic dinoflagellates inside tubules located mainly in their colorful outer mantle. This study examined the effects of exposure to darkness for 30 days and the subsequent 11 days of recovery under a normal photoperiod on the outer mantle of the fluted giant clam Tridacna squamosa. Changes in the abundance of iridocytes and symbionts were assessed by fluorescence microscopy. Light microscopy was applied to quantify symbionts isolated from the outer mantle, while chlorophyll was extracted and analyzed by spectroscopy. The transcript levels of the host's vacuolar-type H+-ATPase subunit A (ATP6V1A) and symbionts' form II ribulose-1,5-bisphosphate carboxylase/oxygenase (Zoox-rbcII) were determined by quantitative real-time PCR (qPCR) and used as proxies for iridocyte abundance and phototrophic potential, respectively. The protein abundance of ATP6V1A and Zoox-RBCII was quantified by western blotting. After exposure to darkness for 30 days, the outer mantle of T. squamosa individuals lost the distinct multiple color patterns, and the gene and protein expression levels of ATP6V1A/ATP6V1A and Zoox-rbcII/Zoox-RBCII decreased dramatically. Microscopy assessment confirmed the reduction in iridocyte and symbiont populations, and the chlorophyll level also decreased considerably. However, just 11 days after returning to a normal light:dark regimen, the quantity of coccoid dinoflagellates and the expression of Zoox-rbcII/Zoox-RBCII in the outer mantle increased significantly to levels higher than those of the individuals prior to exposure to darkness (control), while the chlorophyll content returned to the control level. Additionally, the outer mantle regained most of its coloration with partial recovery of the iridocyte population. These results are relevant not only for understanding the phenomena of light deprivation and symbiont loss in dinoflagellate-associated reef organisms, but also signify that the giant clam-coccoid dinoflagellate holobiont is phototrophically plastic and particularly tolerant to bleaching. [ABSTRACT FROM AUTHOR]

The aim of this work was to evaluate the photosynthetic performance of Pear trees (cv. ‘Rocha’) infected with Erwinia amylovora, three months after suffering a pruning of infected branches (P-trees) compared with asymptomatic trees (C-trees) of the same orchard. Three months after pruning, P-trees looked healthy and were negative for the presence of E. amylovora. In September of 2018, fully expanded leaves of both P- and C- trees were sampled and analysed for photosynthetic parameters related to chlorophyll a fluorescence and gas exchange, alongside with pigments, total soluble sugars, starch, and ribulose-1,5-bisphosphate carboxylase/oxygenase (RuBisCO) contents. No significant differences were found in chlorophyll and carotenoids levels, but anthocyanins significantly decreased in P-trees. Also, despite the maximum quantum yield (Fv/Fm) significantly decreased in P-trees, the effective quantum yield of the PSII was maintained, paralleled with no changes in gas exchange parameters (PN, gs, Ci, E, iWUE, PN/gs), nor in RuBisCO relative content. Finally, the maintenance of the levels of total soluble sugars and starch also supports that the photosynthetic performance of P-trees, three months after pruning, reached values similar to those of the C-trees, contributing to the normal development and ripening of the fruit. Data support that pruning represents a reliable control measure against this quarantine pathogen. This work is the first evaluation of pruning in fire blight management regarding carbon metabolism in P. communis trees.

Abstract Background The ratio of CO2 mesophyll conductance (g m) to Ribulose-1, 5-bisphosphate carboxylase/oxygenase (Rubisco) content has been suggested to positively affect photosynthetic nitrogen use efficiency (PNUE). The anatomical basis of g m has been quantified, but information on the relationship between cell-level anatomies and PNUE is less advanced. Here, hydroponic experiments were conducted in rice plants supplied with ammonium (NH4 +) and nitrate (NO3 −) under three N levels (low, 0.71 mM; intermediate, 2.86 mM; high, 7.14 mM) to investigate the gas exchange parameters, leaf anatomical structure and PNUE. Results The results showed a lower PNUE in plants supplied with high nitrogen and NH4 +, which was positively correlated with the g m/Rubisco ratio. A one-dimensional within-leaf model revealed that the resistance to CO2 diffusion in the liquid phase (r liq) dominated the overall mesophyll resistance (r m), in which CO2 transfer resistance in the cell wall, cytoplasm and stroma were significantly affected by nitrogen supply. The chloroplast surface area exposed to intercellular space (S c) per Rubisco rather than the g m/S c ratio was positively correlated with PNUE and was thus considered a key component influencing PNUE. Conclusion In conclusion, our study emphasized that S c was the most important anatomical trait in coordinating g m and PNUE with contrasting N supply.

A Chlamydomonas reinhardtii RuBisCO-less mutant, ΔrbcL, was used to study carbohydrate metabolism without fixation of atmospheric carbon. The regulatory mechanism(s) that control linear electron flow, known as “photosynthetic control,” are amplified in ΔrbcL at the onset of illumination. With the aim to understand the metabolites that control this regulatory response, we have correlated the kinetics of primary carbon metabolites to chlorophyll fluorescence induction curves. We identify that ΔrbcL in the absence of acetate generates adenosine triphosphate (ATP) via photosynthetic electron transfer reactions. Also, metabolites of the Calvin Benson Bassham (CBB) cycle are responsive to the light. Indeed, ribulose 1,5-bisphosphate (RuBP), the last intermediate before carboxylation by Ribulose-1,5-bisphosphate carboxylase-oxygenase, accumulates significantly with time, and CBB cycle intermediates for RuBP regeneration, dihydroxyacetone phosphate (DHAP), pentose phosphates and ribose-5-phosphate (R5P) are rapidly accumulated in the first seconds of illumination, then consumed, showing that although the CBB is blocked, these enzymes are still transiently active. In opposition, in the presence of acetate, consumption of CBB cycle intermediates is strongly diminished, suggesting that the link between light and primary carbon metabolism is almost lost. Phosphorylated hexoses and starch accumulate significantly. We show that acetate uptake results in heterotrophic metabolism dominating phototrophic metabolism, with glyoxylate and tricarboxylic acid (TCA) cycle intermediates being the most highly represented metabolites, specifically succinate and malate. These findings allow us to hypothesize which metabolites and metabolic pathways are relevant to the upregulation of processes like cyclic electron flow that are implicated in photosynthetic control mechanisms.

Carbon dioxide is essential for life. It is at the beginning of every life process as a substrate of photosynthesis. It is at the end of every life process as the product of post-mortem decay. Therefore, it is not surprising that this gas regulates such diverse processes as cellular chemical reactions, transport, maintenance of the cellular environment, and behaviour. Carbon dioxide is a strategically important research target relevant to crop responses to environmental change, insect vector-borne disease and public health. However, we know little of carbon dioxide’s direct interactions with the cell. The carbamate post-translational modification, mediated by the nucleophilic attack by carbon dioxide on N-terminal α-amino groups or the lysine ɛ-amino groups, is one mechanism by which carbon dioxide might alter protein function to form part of a sensing and signalling mechanism. We detail known protein carbamates, including the history of their discovery. Further, we describe recent studies on new techniques to isolate this problematic post-translational modification.

Biological chemical production has gained traction in recent years as a promising renewable alternative to traditional petrochemical based synthesis. Of particular interest in the field of metabolic engineering are photosynthetic microorganisms capable of sequestering atmospheric carbon dioxide. CO2 levels have continued to rise at alarming rates leading to an increasingly uncertain climate. CO2 can be sequestered by engineered photosynthetic microorganisms and used for chemical production, representing a renewable production method for valuable chemical commodities such as biofuels, plastics, and food additives. The main challenges in using photosynthetic microorganisms for chemical production stem from the seemingly inherent limitations of carbon fixation and photosynthesis resulting in slower growth and lower average product titers compared to heterotrophic organisms. Recently, there has been an increase in research around improving photosynthetic microorganisms as renewable chemical production hosts. This review will discuss the various efforts to overcome the intrinsic inefficiencies of carbon fixation and photosynthesis, including rewiring carbon fixation and photosynthesis, investigating alternative carbon fixation pathways, installing sugar catabolism to supplement carbon fixation, investigating newly discovered fast growing photosynthetic species, and using new synthetic biology tools such as CRISPR to radically alter metabolism.

Carboxylation, chloroplast, crop improvement, dynamic environments, metabolic regulation, photosynthesis, Rubisco, Rubisco activase, protein biochemistry Rubisco and its regulation - major advances to improve carbon assimilation and productivity Keywords: Carboxylation; chloroplast; crop improvement; dynamic environments; metabolic regulation; photosynthesis; protein biochemistry; Rubisco; Rubisco activase EN Carboxylation chloroplast crop improvement dynamic environments metabolic regulation photosynthesis protein biochemistry Rubisco Rubisco activase 507 509 3 01/13/23 20230111 NES 230111 B Significant advances in Rubisco research over the past decade have highlighted the intricate nature of the CO b SB B 2 b sb B -fixing enzyme and the complexity of environmental and cellular factors that affect its activity in photosynthetic organisms. Removal of sugar phosphate derivatives such as CA1P from inhibited catalytic sites requires the action of the AAA+ protein Rca, discovered by [15], which modulates activation and activity of Rubisco. [Extracted from the article]

In 2019 the Biology 100 class of Wenatchee Valley College at Omak (WVCO) worked on a DNA barcoding project with Tabitha Graves's huckleberry research project for bear habitat in Glacier National Park, Montana. Students isolated DNA from huckleberry leaf samples from the National Park. They then ran a PCR with an rbcL primer pair to target the rubisco gene in plant chloroplasts. The PCR product was sequenced by a private company, Genewiz, and the DNA sequence was analyzed through DNA Subway. Twelve student groups, one or two students per group, isolated DNA from huckleberry leaf samples that was sequenced and analyzed. Twelve samples were determined to be of the genus Vaccinium. One of the twelve samples distinguished between the five species of huckleberry, identified the sample as Vaccinium membranaceum, and was published in GenBank. They showed that DNA barcoding can be used successfully to aid in the identification of this species of huckleberry. There were many student outcomes, including hands-on skills with the tools of DNA technology, contributing to a real-world project, and analyzing data for DNA sequence matches. This is a great lab exercise to use for AP biology classes, two-year community college biology classes, and four-year college biology classes at the 100 to 200 level. [ABSTRACT FROM AUTHOR]

SUMMARY: The green alga Chlamydomonas reinhardtii is one of the most studied microorganisms in photosynthesis research and for biofuel production. A detailed understanding of the dynamic regulation of its carbon metabolism is therefore crucial for metabolic engineering. Post‐translational modifications can act as molecular switches for the control of protein function. Acetylation of the ɛ‐amino group of lysine residues is a dynamic modification on proteins across organisms from all kingdoms. Here, we performed mass spectrometry‐based profiling of proteome and lysine acetylome dynamics in Chlamydomonas under varying growth conditions. Chlamydomonas liquid cultures were transferred from mixotrophic (light and acetate as carbon source) to heterotrophic (dark and acetate) or photoautotrophic (light only) growth conditions for 30 h before harvest. In total, 5863 protein groups and 1376 lysine acetylation sites were identified with a false discovery rate of <1%. As a major result of this study, our data show that dynamic changes in the abundance of lysine acetylation on various enzymes involved in photosynthesis, fatty acid metabolism, and the glyoxylate cycle are dependent on acetate and light. Exemplary determination of acetylation site stoichiometries revealed particularly high occupancy levels on K175 of the large subunit of RuBisCO and K99 and K340 of peroxisomal citrate synthase under heterotrophic conditions. The lysine acetylation stoichiometries correlated with increased activities of cellular citrate synthase and the known inactivation of the Calvin–Benson cycle under heterotrophic conditions. In conclusion, the newly identified dynamic lysine acetylation sites may be of great value for genetic engineering of metabolic pathways in Chlamydomonas. [ABSTRACT FROM AUTHOR]

Photosynthesis is an important remaining opportunity for further improvement in the genetic yield potential of our major crops. Measurement, analysis, and improvement of leaf CO2 assimilation (A) have focused largely on photosynthetic rates under light-saturated steady-state conditions. However, in modern crop canopies of several leaf layers, light is rarely constant, and the majority of leaves experience marked light fluctuations throughout the day. It takes several minutes for photosynthesis to regain efficiency in both sun-shade and shade-sun transitions, costing a calculated 10–40% of potential crop CO2 assimilation. Transgenic manipulations to accelerate the adjustment in sun-shade transitions have already shown a substantial productivity increase in field trials. Here, we explore means to further accelerate these adjustments and minimize these losses through transgenic manipulation, gene editing, and exploitation of natural variation. Measurement andanalysis of photosynthesis in sun-shade and shade-sun transitions are explained. Factors limiting speeds of adjustment and how they could be modified to effect improved efficiency are reviewed, specifically nonphotochemical quenching (NPQ), Rubisco activation, and stomatal responses. [ABSTRACT FROM AUTHOR]