Carbon-concentrating Mechanism Research Articles

Changes in the limiting nutrients and dominant phytoplankton communities of three major European rivers: Response to catchment lithologies and human activities

Catchment lithologies and human activities can cause variations in riverine carbon (C), nitrogen (N), and phosphorus (P) loads, and the ratio of these nutrients may affect the primary productivity of rivers and their phytoplankton communities. We conducted a comparative study of biogeochemical variations in three major European rivers—the Rhine, Danube, and Seine—which reveals the impacts of different carbonate outcrops and human impacts (agricultural activities and sewage input) on the changes in the limiting nutrients and phytoplankton community compositions of flowing water systems. N limitation was observed in the Rhine and Danube owing to their low average NO3– loads (4.04–6.45 mg/L). Although the extensive outcrops of carbonate rocks in the Seine catchment result in high average HCO3– concentrations of ∼ 4.68 mmol/L, the elevated inorganic C input did not match the high average NO3– loads (∼19.38 mg/L), driven by intensive agricultural activity and urban sewage input, demonstrating a potential C limitation on productivity. Our results show that the productivity of the Seine River, determined by the chlorophyll-a concentration, was significantly related to the dissolved inorganic carbon (DIC) concentration (coefficient of determination: R2 = 0.68, P < 0.05). Our results demonstrate that changes in DIC content can significantly alter the dominant phytoplankton species in the water column. The high inputs of DIC and aqueous CO2(aq) to the Seine lead to Bacillariophyta proportions that are 3.96–8.81 % and 19.87–22.56 % higher than those of the Rhine and Danube in the wet and dry seasons, respectively. This can be explained by the difference in the efficiency of carbon concentrating mechanisms (CCMs) among phytoplankton species. Therefore, to mitigate river eutrophication and improve the quality of the aquatic environment, we suggest that future water–carbon management in the flowing water systems with abundant nutrients should consider C-limitation alleviation and its potential effect for reducing the abundance of harmful algae.

Modification of Living Diatom, Thalassiosira weissflogii, with a Calcium Precursor through a Calcium Uptake Mechanism: A Next Generation Biomaterial for Advanced Delivery Systems.

The diatom's frustule, characterized by its rugged and porous exterior, exhibits a remarkable biomimetic morphology attributable to its highly ordered pores, extensive surface area, and unique architecture. Despite these advantages, the toxicity and nonbiodegradable nature of silica-based organisms pose a significant challenge when attempting to utilize these organisms as nanotopographically functionalized microparticles in the realm of biomedicine. In this study, we addressed this limitation by modulating the chemical composition of diatom microparticles by modulating the active silica metabolic uptake mechanism while maintaining their intricate three-dimensional architecture through calcium incorporation into living diatoms. Here, the diatom Thalassiosira weissflogii was chemically modified to replace its silica composition with a biodegradable calcium template, while simultaneously preserving the unique three-dimensional (3D) frustule structure with hierarchical patterns of pores and nanoscale architectural features, which was evident by the deposition of calcium as calcium carbonate. Calcium hydroxide is incorporated into the exoskeleton through the active mechanism of calcium uptake via a carbon-concentrating mechanism, without altering the microstructure. Our findings suggest that calcium-modified diatoms hold potential as a nature-inspired delivery system for immunotherapy through antibody-specific binding.

The redox-sensitive R-loop of the carbon control protein SbtB contributes to the regulation of the cyanobacterial CCM

SbtB is a PII-like protein that regulates the carbon-concentrating mechanism (CCM) in cyanobacteria. SbtB proteins can bind many adenyl nucleotides and possess a characteristic C-terminal redox sensitive loop (R-loop) that forms a disulfide bridge in response to the diurnal state of the cell. SbtBs also possess an ATPase/ADPase activity that is modulated by the redox-state of the R-loop. To investigate the R-loop in the cyanobacterium Synechocystis sp. PCC 6803, site-specific mutants, unable to form the hairpin and permanently in the reduced state, and a R-loop truncation mutant, were characterized under different inorganic carbon (Ci) and light regimes. Growth under diurnal rhythm showed a role of the R-loop as sensor for acclimation to changing light conditions. The redox-state of the R-loop was found to impact the binding of the adenyl-nucleotides to SbtB, its membrane association and thereby the CCM regulation, while these phenotypes disappeared after truncation of the R-loop. Collectively, our data imply that the redox-sensitive R-loop provides an additional regulatory layer to SbtB, linking the CO2-related signaling activity of SbtB with the redox state of cells, mainly reporting the actual light conditions. This regulation not only coordinates CCM activity in the diurnal rhythm but also affects the primary carbon metabolism.

An Evidence of Carbonic Anhydrase Activity in Native Microalgae for CO2 Capture Application.

A promising alternative for effective carbon capture has been found in microalgae because of their high photosynthetic capacity and quick growth. The carbon concentration mechanism of many microalgae is heavily reliant on the enzyme carbonic anhydrase (CA), which catalyze the production of bicarbonate from carbon dioxide. In this study, microalgal samples were collected, characterized, and cultured under controlled conditions for their optimal growth of cultures I-IX. The CA activity was investigated using a standard method; the Wilbur-Anderson assay was used to calculate CA activity in microalgal cultures. The comparative study was then used to measure the activity rate of the collected microalgae. Among the tested, culture I, VI, and IX showed a high enzyme activity rate of 4.15, 4.0, and 4.2µg·mL-1, respectively. To determine the rate of carbon dioxide hydration, the method involved tracking the pH change in a reaction mixture. In addition, genetic analysis facilitates the identification of key genes involved in CA activity and other metabolic processes, which enhance the knowledge of microalgal physiology, and enables genetic engineering efforts in the future studies. Overall, this investigation emphasizes the significance of studying unknown microalgal culture and their potential CA activity for industrial and bio-energy applications.

Salinity Mitigates the Negative Effect of Elevated Temperatures on Photosynthesis in the C3-C4 Intermediate Species Sedobassia sedoides.

The adaptation of plants to combined stresses requires unique responses capable of overcoming both the negative effects of each individual stress and their combination. Here, we studied the C3-C4 (C2) halophyte Sedobassia sedoides in response to elevated temperature (35 °C) and salinity (300 mM NaCl) as well as their combined effect. The responses we studied included changes in water-salt balance, light and dark photosynthetic reactions, the expression of photosynthetic genes, the activity of malate dehydrogenase complex enzymes, and the antioxidant system. Salt treatment led to altered water-salt balance, improved water use efficiency, and an increase in the abundance of key enzymes involved in intermediate C3-C4 photosynthesis (i.e., Rubisco and glycine decarboxylase). We also observed a possible increase in the activity of the C2 carbon-concentrating mechanism (CCM), which allowed plants to maintain high photosynthesis intensity and biomass accumulation. Elevated temperatures caused an imbalance in the dark and light reactions of photosynthesis, leading to stromal overreduction and the excessive generation of reactive oxygen species (ROS). In response, S. sedoides significantly activated a metabolic pathway for removing excess NADPH, the malate valve, which is catalyzed by NADP-MDH, without observable activation of the antioxidant system. The combined action of these two factors caused the activation of antioxidant defenses (i.e., increased activity of SOD and POX and upregulation of FDI), which led to a decrease in oxidative stress and helped restore the photosynthetic energy balance. Overall, improved PSII functioning and increased activity of PSI cyclic electron transport (CET) and C2 CCM led to an increase in the photosynthesis intensity of S. sedoides under the combined effect of salinity and elevated temperature relative to high temperature alone.

Characterization of a novel γ-type carbonic anhydrase, Sjγ-CA2, in Saccharina japonica: Insights into carbon concentration mechanism in macroalgae

Carbonic anhydrase (CA) is a crucial component of CO2-concentrating mechanism (CCM) in macroalgae. In Saccharina japonica, an important brown seaweed, 11 CAs, including 5 α-, 3 β-, and 3 γ-CAs, have been documented. Among them, one α-CA and one β-CA were localized in the periplasmic space, one α-CA was found in the chloroplast, and one γ-CA was situated in mitochondria. Notably, the known γ-CAs have predominantly been identified in mitochondria. In this study, we identified a chloroplastic γ-type CA, Sjγ-CA2, in S. japonica. Based on the reported amino acid sequence of Sjγ-CA2, the epitope peptide for monoclonal antibody production was selected as 165 Pro-305. After purification and specificity identification, anti-SjγCA2 monoclonal antibody was employed in immunogold electron microscopy. The results illustrated that Sjγ-CA2 was localized in the chloroplasts of both gametophytes and sporophytes of S. japonica. Subsequently, immunoprecipitation coupled with LC-MS/MS analysis revealed that Sjγ-CA2 mainly interacted with photosynthesis-related proteins. Moreover, the first 65 amino acids at N-terminal of Sjγ-CA2 was identified as the chloroplast transit peptide by the transient expression of GFP-SjγCA2 fused protein in tabacco. Real-time PCR results demonstrated an up-regulation of the transcription of Sjγ-CA2 gene in response to high CO2 concentration. These findings implied that Sjγ-CA2 might contribute to minimizing the leakage of CO2 from chloroplasts and help maintaining a high concentration of CO2 around Rubisco.

The bHLH transcription factor OsPRI1 activates the Setaria viridisPEPC1 promoter in rice.

Photosynthetic efficiency is reduced by the dual role of Rubisco, which acts either as a carboxylase or as an oxygenase, the latter leading to photorespiration. C4 photosynthesis evolved as a carbon-concentrating mechanism to reduce photorespiration. To engineer C4 into a C3 plant, it is essential to understand how C4 genes, such as phosphoenolpyruvate carboxylase (PEPC1), are regulated to be expressed at high levels and in a cell-specific manner. Yeast one-hybrid screening was used to show that OsPRI1, a rice bHLH transcription factor involved in iron homeostasis, binds to the Setaria viridis PEPC1 promoter. This promoter drives mesophyll-specific gene expression in rice. The role of OsPRI1 in planta was characterized using a rice line harbouring SvPEPC1pro ::GUS. We show that OsPRI1 activates the S. viridis PEPC1 promoter by binding to an N-box in the proximal promoter, and that GUS activity is highly reduced in SvPEPC1pro ::GUS lines when OsPRI1 is mutated. Cross-species comparisons showed that the SvPRI1 homolog binds to the SvPEPC1 promoter but the maize ZmPRI1 does not bind to the ZmPEPC1 promoter. Our results suggest that elements of the iron homeostasis pathway were co-opted to regulate PEPC1 gene expression during the evolution of some but not all C4 species.

CAH3 from Chlamydomonas reinhardtii: Unique Carbonic Anhydrase of the Thylakoid Lumen.

CAH3 is the only carbonic anhydrase (CA) present in the thylakoid lumen of the green algae Chlamydomonas reinhardtii. The monomer of the enzyme has a molecular weight of ~29.5 kDa with high CA activity. Through its dehydration activity, CAH3 can be involved either in the carbon-concentrating mechanism supplying CO2 for RuBisCO in the pyrenoid or in supporting the maximal photosynthetic activity of photosystem II (PSII) by accelerating the removal of protons from the active center of the water-oxidizing complex. Both proposed roles are considered in this review, together with a description of the enzymatic parameters of native and recombinant CAH3, the crystal structure of the protein, and the possible use of lumenal CA as a tool for increasing biomass production in higher plants. The identified involvement of lumenal CAH3 in the function of PSII is still unique among green algae and higher plants and can be used to understand the mechanism(s) of the functional interconnection between PSII and the proposed CA(s) of the thylakoid lumen in other organisms.

Inorganic carbon sensing and signalling in cyanobacteria

AbstractCyanobacteria utilize CO2 and HCO3− as inorganic carbon (Ci) sources. In low Ci, like in ambient air, cyanobacteria efficiently collect Ci using a carbon concentrating mechanism (CCM). The CCM includes bicarbonate transporters SbtA, BicA and BCT1; the specialized NDH complexes NDH‐13 and NDH‐14, which convert CO2 to HCO3− in the cytoplasm; and carboxysomes that are protein shell encapsulated ribulose‐1,5‐bisphosphate carboxylase/oxygenase (RuBisCo) and carbonic anhydrase containing bodies in which the first reaction of carbon fixation occurs. Ci‐dependent regulation of bicarbonate transporters and specialized NDH complexes, especially the regulation of the SbtA transporter, are well understood. CcmR (also called NdhR), CyAbrB2, CmpR and RbcR act as transcription factors regulating CCM genes. Ci signalling molecules detecting the metabolic status of the cells include 2‐oxoglutarate, which accumulates when the Ci/nitrogen ratio of the cell is high, and 2‐phosphoglycolate, the first intermediate of the photorespiration pathway, whose accumulation indicates low Ci. These signalling molecules act as corepressors and coactivators of the CcmR repressor protein, whereas 2‐phosphoglycolate and ribulose‐1,5‐bisphosphate activate transcription activator CmpR. In addition, bicarbonate or CO2 activates the adenylyl cyclase that produces cAMP, and ATP/ADP/AMP provide information about the energy status of the cell. Less is known about the molecular mechanisms regulating carboxysome dynamics or how production, activity and degradation of photosynthetic complexes are regulated by prevailing Ci conditions or which mechanisms adjust cell division according to Ci. This minireview summarizes the present knowledge about molecular mechanisms regulating cyanobacterial acclimation to prevailing Ci.

Measurement of Algal Photosynthesis Using a Clark-Type O2 Electrode.

The Clark-type electrode can be used to assess the rates of photosynthesis by detecting changes in O2 concentration in a culture. This chapter describes a method for a liquid phase measurement of light and dissolved inorganic carbon-dependent photosynthesis using the model green alga Chlamydomonas reinhardtii. The technique can be used to evaluate the presence or efficiency of carbon-concentrating mechanisms.

Single-species dinoflagellate cyst carbon isotope fractionation in core-top sediments: environmental controls, CO2 dependency and proxy potential

Abstract. Sedimentary bulk organic matter and various molecular organic components exhibit strong CO2-dependent carbon isotope fractionation relative to dissolved inorganic carbon sources. This fractionation (εp) has been employed as a proxy for paleo-pCO2. Yet, culture experiments indicate that CO2-dependent εp is highly specific at genus and even species level, potentially hampering the use of bulk organic matter and non-species-specific organic compounds. In recent years, significant progress has been made towards a CO2 proxy using controlled growth experiments with dinoflagellate species, also showing highly species-specific εp values. These values were, however, based on motile specimens, and it remains unknown whether these relations also hold for the organic-walled resting cysts (dinocysts) produced by these dinoflagellate species in their natural environment. We here analyze dinocysts isolated from core tops from the Atlantic Ocean and Mediterranean Sea, representing several species (Spiniferites elongatus, S. (cf.) ramosus, S. mirabilis, Operculodinium centrocarpum sensu Wall and Dale (1966) (hereafter referred to as O. centrocarpum) and Impagidinium aculeatum) using laser ablation–nano-combustion–gas-chromatography–isotope ratio mass spectrometry (LA/nC/GC-IRMS). We find that the dinocysts produced in the natural environment are all appreciably more 13C-depleted compared to the cultured motile dinoflagellate cells, implying higher overall εp values, and, moreover, exhibit large isotope variability. Where several species could be analyzed from a single location, we often record significant differences in isotopic variance and offsets in mean δ13C values between species, highlighting the importance of single-species carbon isotope analyses. The most geographically expanded dataset, based on O. centrocarpum, shows that εp correlates significantly with various environmental parameters. Importantly, O. centrocarpum shows a CO2-dependent εp above ∼ 240 µatm pCO2. Similar to other marine autotrophs, relative insensitivity at low pCO2 is in line with active carbon-concentrating mechanisms at low pCO2, although we here cannot fully exclude that we partly underestimated εp sensitivity at low pCO2 values due to the relatively sparse sampling in that range. Finally, we use the relation between εp and pCO2 in O. centrocarpum to propose a first pCO2 proxy based on a single dinocyst species.

Multiple highly expressed phosphoenolpyruvate carboxylase genes have divergent enzyme kinetic properties in two C4 grasses.

Phosphoenolpyruvate (PEP) carboxylase (PEPC) catalyses the irreversible carboxylation of PEP with bicarbonate to produce oxaloacetate. This reaction powers the carbon-concentrating mechanism (CCM) in plants that perform C4 photosynthesis. This CCM is generally driven by a single PEPC gene product that is highly expressed in the cytosol of mesophyll cells. We found two C4 grasses, Panicum miliaceum and Echinochloa colona, that each have two highly expressed PEPC genes. We characterized the kinetic properties of the two most abundant PEPCs in E. colona and P. miliaceum to better understand how the enzyme's amino acid structure influences its function. Coding sequences of the two most abundant PEPC proteins in E. colona and P. miliaceum were synthesized by GenScript and were inserted into bacteria expression plasmids. Point mutations resulting in substitutions at conserved amino acid residues (e.g. N-terminal serine and residue 890) were created via site-directed PCR mutagenesis. The kinetic properties of semi-purified plant PEPCs from Escherichia coli were analysed using membrane-inlet mass spectrometry and a spectrophotometric enzyme-coupled reaction. The two most abundant P. miliaceum PEPCs (PmPPC1 and PmPPC2) have similar sequence identities (>95 %), and as a result had similar kinetic properties. The two most abundant E. colona PEPCs (EcPPC1 and EcPPC2) had identities of ~78 % and had significantly different kinetic properties. The PmPPCs and EcPPCs had different responses to allosteric inhibitors and activators, and substitutions at the conserved N-terminal serine and residue 890 resulted in significantly altered responses to allosteric regulators. The two, significantly expressed C4Ppc genes in P. miliaceum were probably the result of genomes combining from two closely related C4Panicum species. We found natural variation in PEPC's sensitivity to allosteric inhibition that seems to bypass the conserved 890 residue, suggesting alternative evolutionary pathways for increased malate tolerance and other kinetic properties.

Polyphasic characterization of a novel hot-spring cyanobacterium Thermocoleostomius sinensis gen et sp. nov. and genomic insights into its carbon concentration mechanism.

Thermophilic cyanobacteria play a crucial role as primary producers in hot spring ecosystems, yet their microbiological, taxonomic, and ecological characteristics are not extensively studied. This study aimed to characterize a novel strain of thermophilic cyanobacteria, PKUAC-SCTA174 (A174), using a combination of traditional polyphasic methods and modern genomic-based approaches. The study included 16S rRNA-based phylogeny, ITS secondary structure prediction, morphological and habitat analyses, as well as high-quality genome sequencing with corresponding phylogenomic analyses. The results of the 16S rRNA, 16S-23S ITS secondary structure, morphology, and habitat analyses supported the classification of the strain as a member of a novel genus within the family Oculatellaceae, closely related to Albertania and Trichotorquatus. Genomic analysis revealed the presence of a sophisticated carbon-concentrating mechanism (CCM) in the strain, involving two CO2 uptake systems NDH-I3, and NDH-I4, three types of bicarbonate transporters (BCT1, bicA, sbtA,) and two distinct putative carboxysomal carbonic anhydrases (ccaA1 and ccaA2). The expression of CCM genes was investigated with a CO2 shift experiment, indicating varying transcript abundance among different carbon uptake systems. Based on the comprehensive characterization, the strain was delineated as Thermocoleostomius sinensis, based on the botanical code. The study of the complete genome of strain A174 contributes valuable insights into the genetic characteristics of the genus Thermocoleostomius and related organisms and provides a systematic understanding of thermophilic cyanobacteria. The findings presented here offer valuable data that can be utilized for future research in taxogenomics, ecogenomics, and geogenomics.

Novel protein CcmS is required for stabilization of the assembly of β-carboxysome in Synechocystis sp. strain PCC 6803.

The carboxysome plays an essential role in the carbon concentration mechanism in cyanobacteria. Although significant progress has been made in the structural analysis of the carboxysome, little is still known about its biosynthesis. We identified slr1911, a gene encoding a protein of unknown function in cyanobacterium Synechocystis sp. Strain PCC 6803 (Syn6803), which we termed ccmS by screening a low CO2 -sensitive mutant. CcmS interacts with CcmK1 and CcmM. The former is a shell protein of the β-carboxysome and the latter is a crucial component of the β-carboxysome, which is responsible for aggregating RuBisCO and recruiting shell proteins. The deletion of ccmS lowers the accumulation and assembly of CcmK1, resulting in aberrant carboxysomes, suppressed photosynthetic capacities, and leads to a slow growth phenotype, especially under CO2 -limited conditions. These observations suggest that CcmS stabilizes the assembly of the β-carboxysome shell and likely connects the carboxysome core with the shell. Our results provide a molecular view of the role played by CcmS in the formation of the β-carboxysome and its function in Syn6803.

Isotopic and biochemical composition of Western Mediterranean macroalgae

ABSTRACT Among marine primary producers, macroalgae support complex and productive coastal food webs, but coastal primary production relies on terrigenous inputs and remineralized organic matter which both vary seasonally. An approach combining stable isotope and biochemical analyses enables a better characterization of macroalgae specificities and highlights environmental influences on their chemical signature. This study compared the isotopic signature and biochemical composition of 22 Mediterranean macroalgae belonging to Rhodophyta (red algae), Phaeophyceae (brown algae) and Chlorophyta (green algae) between March and November 2010 to capture the differences in species chemical signatures potentially driven by metabolic traits or environmental drivers. Carbon stable isotope values were evidenced as a good proxy of specific carbon metabolism: low values observed in red algae could be related to the reported absence of carbon concentrating mechanisms (CCMs) in this group while higher values were driven by strong CCM activity in green algae. Biochemical patterns also differed between groups: soluble carbohydrates were a major component for red algae, while lipids and proteins dominated in brown algae, and insoluble carbohydrate concentrations were high in green algae. Variation within species across two collection times could be related to environmental changes and algal metabolism. δ15N values confirm the efficiency of this parameter as a proxy of the impact of human influence in the Bay of Marseille.

The V-type ATPase enhances photosynthesis in marine phytoplankton and further links phagocytosis to symbiogenesis

Diatoms, dinoflagellates, and coccolithophores are dominant groups of marine eukaryotic phytoplankton that are collectively responsible for the majority of primary production in the ocean.1 These phytoplankton contain additional intracellular membranes around their chloroplasts, which are derived from ancestral engulfment of red microalgae by unicellular heterotrophic eukaryotes that led to secondary and tertiary endosymbiosis.2 However, the selectable evolutionary advantage of these membranes and the physiological significance for extant phytoplankton remain poorly understood. Since intracellular digestive vacuoles are ubiquitously acidified by V-type H+-ATPase (VHA),3 proton pumps were proposed to acidify the microenvironment around secondary chloroplasts to promote the dehydration of dissolved inorganic carbon (DIC) into CO2, thus enhancing photosynthesis.4,5 We report that VHA is localized around the chloroplasts of centric diatoms and that VHA significantly contributes to their photosynthesis across a wide range of oceanic irradiances. Similar results in a pennate diatom, dinoflagellate, and coccolithophore, but not green or red microalgae, imply the co-option of phagocytic VHA activity into a carbon-concentrating mechanism (CCM) is common to secondary endosymbiotic phytoplankton. Furthermore, analogous mechanisms in extant photosymbiotic marine invertebrates6,7,8 provide functional evidence for an adaptive advantage throughout the transition from endosymbiosis to symbiogenesis. Based on the contribution of diatoms to ocean biogeochemical cycles, VHA-mediated enhancement of photosynthesis contributes at least 3.5 Gtons of fixed carbon per year (or 7% of primary production in the ocean), providing an example of a symbiosis-derived evolutionary innovation with global environmental implications.

Research Progress in Improving Photosynthetic Efficiency.

Photosynthesis is the largest mass- and energy-conversion process on Earth, and it is the material basis for almost all biological activities. The efficiency of converting absorbed light energy into energy substances during photosynthesis is very low compared to theoretical values. Based on the importance of photosynthesis, this article summarizes the latest progress in improving photosynthesis efficiency from various perspectives. The main way to improve photosynthetic efficiency is to optimize the light reactions, including increasing light absorption and conversion, accelerating the recovery of non-photochemical quenching, modifying enzymes in the Calvin cycle, introducing carbon concentration mechanisms into C3 plants, rebuilding the photorespiration pathway, de novo synthesis, and changing stomatal conductance. These developments indicate that there is significant room for improvement in photosynthesis, providing support for improving crop yields and mitigating changes in climate conditions.

Adaptive diversity in structure and function of C4 photosynthetic components.

Many tropical and subtropical plant lineages have independently evolved C4 photosynthesis. The convergent evolution of this complex functional trait from different ancestors is reflected in variations in the structural and biochemical characteristics of C4 components such as enzymes and cellular specializations. The mechanism of C4 carbon concentration mostly involves coordinated function of mesophyll and bundle sheath cells. Important adaptations of the C4 syndrome include increased vein density and the development of photosynthetic bundle sheath cells with low gas conductance. In addition, the enzymes and transporters of the C4 pathway evolved via the co-option of multiple genes, each derived from a specific lineage of isoforms present in nonC4-ancestors. In particular, the adaptation of C4 enzymes resulted in a variety of structural and biochemical modifications, generally leading to increased catalytic efficiency and regulation by metabolites and post-translational modifications. Differences in these adaptations are particularly evident in the C4-acid decarboxylation step, which can be catalyzed by three decarboxylases that define the C4 subtypes. Associated with the biochemical subtypes, there are also differences in the extend of grana staking and localization of bundle sheath cells chloroplasts. The presence of a suberin layer and symplastic connections also likely vary among the different C4-subtypes. This review examines the current understanding of the diversity of structural and functional changes in key components of the C4 carbon concentration mechanism. This knowledge is necessary not only to identify divergent solutions for convergent optimization of C4 components in different C4 lineages, but also to guide their creation for rational synthetic biology approaches.

Characterization of a novel thermophilic cyanobacterium within Trichocoleusaceae, Trichothermofontia sichuanensis gen. et sp. nov., and its CO2-concentrating mechanism.

Thermophiles from extreme thermal environments have shown tremendous potential regarding ecological and biotechnological applications. Nevertheless, thermophilic cyanobacteria remain largely untapped and are rarely characterized. Herein, a polyphasic approach was used to characterize a thermophilic strain, PKUAC-SCTB231 (hereafter B231), isolated from a hot spring (pH 6.62, 55.5°C) in Zhonggu village, China. The analyses of 16S rRNA phylogeny, secondary structures of 16S-23S ITS and morphology strongly supported strain B231 as a novel genus within Trichocoleusaceae. Phylogenomic inference and three genome-based indices further verified the genus delineation. Based on the botanical code, the isolate is herein delineated as Trichothermofontia sichuanensis gen. et sp. nov., a genus closely related to a validly described genus Trichocoleus. In addition, our results suggest that Pinocchia currently classified to belong to the family Leptolyngbyaceae may require revision and assignment to the family Trichocoleusaceae. Furthermore, the complete genome of Trichothermofontia B231 facilitated the elucidation of the genetic basis regarding genes related to its carbon-concentrating mechanism (CCM). The strain belongs to β-cyanobacteria according to its β-carboxysome shell protein and 1B form of Ribulose bisphosphate Carboxylase-Oxygenase (RubisCO). Compared to other thermophilic strains, strain B231contains a relatively low diversity of bicarbonate transporters (only BicA for HCO3- transport) but a higher abundance of different types of carbonic anhydrase (CA), β-CA (ccaA) and γ-CA (ccmM). The BCT1 transporter consistently possessed by freshwater cyanobacteria was absent in strain B231. Similar situation was occasionally observed in freshwater thermal Thermoleptolyngbya and Thermosynechococcus strains. Moreover, strain B231 shows a similar composition of carboxysome shell proteins (ccmK1-4, ccmL, -M, -N, -O, and -P) to mesophilic cyanobacteria, the diversity of which was higher than many thermophilic strains lacking at least one of the four ccmK genes. The genomic distribution of CCM-related genes suggests that the expression of some components is regulated as an operon and others in an independently controlled satellite locus. The current study also offers fundamental information for future taxogenomics, ecogenomics and geogenomic studies on distribution and significance of thermophilic cyanobacteria in the global ecosystem.

Enhancing overall carbon fixation and energy conversion with formate in green microalga Chlamydomonas reinhardtii

The natural carbon-concentration mechanism (CCM) is the solution to achieve high photosynthetic efficiency. Mimicking the natural CCM, exogenous formate which can be produced by renewable energy and CO2 artificially, was introduced into a model photosynthetic organism, Chlamydomonas reinhardtii, to improve the overall photosynthesis. To release the adverse effects of formate, the combination of different strategies, including co-substrate, evolution, and conditions optimization, the formate tolerance to over 40 mM, which was higher than previous irreversible inhibition limitation of formate on C. reinhardtii, was achieved, and formate-mediated enhancement of growth and photosynthesis activity was obtained with a 27 % biomass improvement and a 60 % increase in photosynthetic oxygen evolution. Interestingly, 82 % of formate was converted with the illumination, indicating the existence of energy interaction between photosynthesis and formate assimilation. Meanwhile, formate suppressed respiration and reduced a 29 % biomass decline in the night. Moreover, supported by the enzyme inhibitor, formate consumption, and biochemical compositions, it was postulated that formate was assimilated via the pyruvate formate-lyase (PFL) enzyme to stimulate the increase of photosynthetic components to maintain photosynthesis activity. With the overexpressed PFL and conditions optimization, formate could be further used as the sole organic carbon source for higher biomass in C. reinhardtii. This work presents the potentiality of introducing formate as artificial carbon intermediates to natural metabolic pathways for higher-than-nature photosynthetic efficiency.