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Highlights • Karenia brevis is not affected by changes in environmentally relevant CO 2 concentrations. • Karenia brevis maintains an efficient and regulated CO 2 concentrating mechanism (CCM). • The CCM can sustain dense blooms Karenia brevis. • Rising CO 2 can potentially elevate the negative socioeconomic effects of Karenia brevis. Abstract Karenia brevis is a marine dinoflagellate commonly found in the Gulf of Mexico and important both ecologically and economically due to its production of the neurotoxin brevetoxin, which can cause respiratory illness in humans and widespread death of marine animals. K. brevis strains have previously shown to be sensitive to changes in CO 2 , both in terms of growth as well as toxin production. Our study aimed to understand this sensitivity by measuring underlying mechanisms, such as photosynthesis, carbon acquisition, and photophysiology. K. brevis (CCFWC-126) did not show a significant response in growth, cellular composition of carbon and nitrogen, nor in photosynthetic rates between p CO 2 concentrations of 150, 400, or 780 µatm. However, a strong response in its acquisition of inorganic carbon was found. Half saturation values for CO 2 increased from 1.5 to 3.3 µM, inorganic carbon preference switched from HCO 3 − to CO 2 (14–56% CO 2 usage), and external carbonic anhydrase activity was downregulated by 23% when comparing low and high p CO 2. We conclude that K. brevis must employ an efficient and regulated CO 2 concentrating mechanism (CCM) to maintain constant carbon fixation and growth across p CO 2 levels. No statistically significant correlation between CO 2 and brevetoxin content was found, yet a positive trend with enhanced p CO 2 was detected. This study is the first explaining how this socioeconomically important species is able to efficiently supply inorganic carbon for photosynthesis, which can potentially prolong bloom situations. This study also highlights that elevated CO 2 concentrations, as projected for a future ocean, can affect underlying physiological processes of K. brevis , some of which could lead to increases in cellular brevetoxin production and therefore increased impacts on coastal ecosystems and economies. [ABSTRACT FROM AUTHOR]

Heterosigma akashiwo is a raphidophyte known for forming ichthyotoxic blooms. In order to predict the potential impacts of rising CO 2 on H. akashiwo it is necessary to understand the factors influencing growth rates over a range of CO 2 concentrations. Here we examined the physiology and gene expression response of H. akashiwo to concentrations from 200 to 1000 ppm CO 2. Growth rate data were combined from this and previous studies and fit with a CO 2 limitation-inhibition model that revealed an apparent growth optimum around 600–800 ppm CO 2. Physiological changes included a significant increase in C:N ratio at ∼800 ppm CO 2 and a significant decrease in hydrogen peroxide concentration at ∼1000 ppm. Whole transcriptome sequencing of H. akashiwo revealed sharp distinctions in metabolic pathway gene expression between ∼600 and ∼800 ppm CO 2. Hierarchical clustering by co-expression identified groups of genes with significant correlations to CO 2 and growth rate. Genes with significant differential expression with CO 2 included carbon concentrating mechanism genes such as beta-carbonic anhydrases and a bicarbonate transporter, which may underpin shifts in physiology. Genes involved in cell motility were significantly changed by both elevated CO 2 and growth rate, suggesting that future ocean conditions could modify swimming behavior in this species. [ABSTRACT FROM AUTHOR]

Microalgae can tolerate different CO 2 concentrations and its mechanism of rapid CO 2 fixation is key to understanding the relationship between intracellular substance and energy conversion, as well as to optimize the production of microalgae bioproducts. The present study conducted a transcriptome-based analysis of gene expression of an industrial Chlorella pyrenoidosa strain, FACHB-9, with high oil production ability under different CO 2 concentrations. Transcriptome-based gene expression analysis indicated the redirection of central metabolism under CO 2 deprivation. The C3 pathway served as the main metabolic pathway for C. pyrenoidosa , which was subjected to a high CO 2 environment in the present study. Similar to C4 plants wherein limited CO 2 activates CO 2 -concentrating mechanism to compensate for the low activity of RuBisCO in the Calvin cycle, C. pyrenoidosa undergoes CO 2 compensation by active transport to ensure sufficient amounts of Ci for its growth and metabolism. Comparative analysis has allowed the identification of several candidate genes for further strain improvement. These genes encode proteins that might be CAs, as indicated by its localization to the cell membrane or chloroplast membrane, or act as Ci transporters that assist Ci transmembrane transportation. Certain types of ABC transport proteins and CCCH-type zinc finger proteins also showed significant changes in gene expression. These findings indicate that these may be promising targets for functional and genetic modification studies. The results not only reveal the significance of the mechanism of carbon sequestration to eukaryotic green algae, but also provide a basis for further construction of the energy microalgae cell factory with high efficiency carbon biofixation capacity. [ABSTRACT FROM AUTHOR]

Carbon dioxide (CO 2) and light are essential for high photosynthetic rates of microalgal cultures. Microalgal photosynthetic behavior at low CO 2 concentrations has not been revealed yet at the same level of detail as leaf photosynthesis. In the present study, we investigated the short-term photosynthetic response of suspended Chlorella sorokiniana to limiting light intensity and CO 2 concentration. We used a novel CO 2 -based photosynthesis monitor originating from leaf research but equipped with an aquatic chamber sparged with CO 2 enriched air. Photosynthesis was measured by employing a steady-state CO 2 mass balance over the chamber. Light and carbon response curves were determined under constant pH, temperature, dissolved oxygen, light intensity or dissolved carbon dioxide. We determined the volumetric mass transfer coefficient of the aquatic chamber to accurately convert gaseous CO 2 partial pressure into aqueous CO 2 concentration to evaluate the CO 2 response measurements. Light response measurements on dilute algal cultures revealed a high photosynthetic capacity on a time scale of minutes which by far exceeded the average CO 2 uptake on a time scale of hours (cell growth). Light response measurements with dense and fully absorbing cultures provided accurate insight into the response of microalgal mass culture to changing incident irradiance, including the efficiency of photosynthesis of the algal culture as a whole. CO 2 response measurements demonstrated severe CO 2 limitation at dissolved CO 2 levels of <20 μM giving a concrete target for optimization of CO2 supply in large-scale cultivation systems. Measurements at a background of either 21 % O 2 (air) or 2 % O 2 indicated the existence of photorespiration in Chlorella. Moreover, the magnitude of photorespiration first increased and then decreased again while reduced dissolved CO 2 levels, indicating the activation of a carbon concentrating mechanism at low CO 2. Simultaneous measurement of the quantum yield of PSII linear electron transport (ϕ PSII) and CO 2 uptake revealed the transition of CO 2 limited photosynthesis towards light-limited photosynthesis. • Algal photosynthetic response to limiting light intensity and CO 2 concentration (81 characters) • Photosynthetic response measurement requires apt choice of biomass concentration (82 characters). • CO 2 response at different O 2 concentrations suggested photorespiration in Chlorella (85 characters). • Activation of carbon concentrating mechanism amid measurements was indicated (78 characters). • Monod equation is too simple to describe algal photosynthetic responses precisely (83 characters). [ABSTRACT FROM AUTHOR]

Inorganic carbon (C i ) is the major sink for photosynthetic reductant in organisms capable of oxygenic photosynthesis. In the absence of abundant C i , the cyanobacterium Synechocystis sp. strain PCC6803 expresses a high affinity C i acquisition system, the CO 2 -concentrating mechanisms (CCM), controlled by the transcriptional regulator CcmR and the metabolites NADP + and α-ketoglutarate, which act as co-repressors of CcmR by modulating its DNA binding. The CCM thus responds to internal cellular redox changes during the transition from C i -replete to C i -limited conditions. However, the actual changes in the metabolic state of the NADPH/NADP + system that occur during the transition to C i -limited conditions remain ill-defined. Analysis of changes in the redox state of cells experiencing C i limitation reveals systematic changes associated with physiological adjustments and a trend towards the quinone and NADP pools becoming highly reduced. A rapid and persistent increase in F 0 was observed in cells reaching the C i -limited state, as was the induction of photoprotective fluorescence quenching. Systematic changes in the fluorescence induction transients were also observed. As with Chl fluorescence, a transient reduction of the NADPH pool (‘M’ peak), is assigned to State 2→ State 1 transition associated with increased electron flow to NADP + . This was followed by a characteristic decline, which was abolished by C i limitation or inhibition of the Calvin –Benson–Bassham (CBB) cycle and is thus assigned to the activation of the CBB cycle. The results are consistent with the proposed regulation of the CCM and provide new information on the nature of the Chl and NADPH fluorescence induction curves. [ABSTRACT FROM AUTHOR]

Microalgae are a diverse group of photosynthetic organisms that can be exploited for the production of different compounds, ranging from crude biomass and biofuels to high value-added biochemicals and synthetic proteins. Traditionally, algal biotechnology relies on bioprospecting to identify new highly productive strains and more recently, on forward genetics to further enhance productivity. However, it has become clear that further improvements in algal productivity for biotechnology is impossible without combining traditional tools with the arising molecular genetics toolkit. We review recent advantages in developing high throughput screening methods, preparing genome-wide mutant libraries, and establishing genome editing techniques. We discuss how algae can be improved in terms of photosynthetic efficiency, biofuel and high value-added compound production. Finally, we critically evaluate developments over recent years and explore future potential in the field. [ABSTRACT FROM AUTHOR]

As all plastids that have been investigated so far can be traced back to endosymbiotic uptake of cyanobacteria by heterotrophic host cells, they accordingly show a high similarity regarding photosynthesis, which includes both the photosystems and the biochemical reactions around the CO 2 fixation via the Calvin–Bassham cycle. Major differences between the different algal and plant groups may include the presence or absence of carbon concentrating mechanisms, pyrenoids, Rubisco activases, carbonic anhydrases as well as differences in the regulation of the Calvin–Bassham cycle. This review describes the diversity of primary carbon fixation steps in algae and plants and the respective regulatory mechanisms. [ABSTRACT FROM AUTHOR]

Abstract: We investigated the catalytic activity and inhibition of the β-class carbonic anhydrase (CA, EC 4.2.1.1) CahB1, from the relict cyanobacterium Coleofasciculus chthonoplastes (previously denominated Microcoleus chthonoplastes). The enzyme showed good activity as a catalyst for the CO2 hydration, with a k cat of 2.4×105 s−1 and a k cat/K m of 6.3×107 M−1 s−1. A range of inorganic anions and small molecules were investigated as inhibitors of CahB1. Perchlorate and tetrafluoroborate did not inhibit the enzyme (K Is >200mM) whereas selenate and selenocyanide were ineffective inhibitors too, with K Is of 29.9–48.61mM. The halides, pseudohalides, carbonate, bicarbonate, trithiocarbonate and a range of heavy metal ions-containing anions were submillimolar–millimolar inhibitors (K Is in the range of 0.15–0.90mM). The best CahB1 inhibitors were N,N-diethyldithiocarbamate, sulfamate, sulfamide, phenylboronic acid and phenylarsonic acid, with K Is in the range of 8–75μM, whereas acetazolamide inhibited the enzyme with a K I of 76nM. This is the first kinetic and inhibition study of a cyanobacterial CA. As these enzymes are widespread in many cyanobacteria, being crucial for the carbon concentrating mechanism which assures substrate to RubisCO for the CO2 fixation by these organisms, a detailed kinetic/inhibition study may be essential for a better understanding of this superfamily of metalloenzymes and for potential biotechnological applications in biomimetic CO2 capture processes. [Copyright &y& Elsevier]

Abstract: Coccolithophorid algae (Haptophycea) are mainly marine unicellular phytoplankton. The coccolithophorids are of global interest as they can fix carbon by photosynthesis as well as in calcium carbonate (coccoliths). They are the largest carbon sinks and one of the largest primary producers on the planet. They can also produce high amounts of lipids which have a high potential application as a renewable fuel and alternative food source. This paper reviews current knowledge on coccolithophorid algae photosynthesis and calcification and their potential industrial applications. [Copyright &y& Elsevier]

Abstract: Many photosynthetic organisms have developed inorganic carbon (Ci) concentrating mechanisms (CCMs) that increase the CO2 concentration within the vicinity of ribulose-1,5-bisphosphate carboxylase/oxygenase (RubisCO). Several CCMs, such as four carbon (C4) and crassulacean acid metabolism (CAM), bicarbonate accumulation systems and capsular structures around RubisCO have been described in great detail. These systems are believed to have evolved several times as mechanisms that acclimate organisms to unfavourable growth conditions. Based on recent experimental evidence we propose the occurrence of another more general CCM system present in all plants. This basal CCM (bCCM) is supposed to be composed of mitochondrial carbonic anhydrases (a β-type carbonic anhydrase and the γ-type carbonic anhydrase domain of the mitochondrial NADH dehydrogenase complex) and probably further unknown components. The bCCM is proposed to reduce leakage of CO2 from plant cells and allow efficient recycling of mitochondrial CO2 for carbon fixation in chloroplasts. [Copyright &y& Elsevier]

The availability of inorganic carbon (C i) as the source for photosynthesis is fluctuating in aquatic environments. Despite the involvement of transcriptional regulators CmpR and NdhR in regulating genes encoding C i transporters at limiting CO 2 , the C i -sensing mechanism is largely unknown among cyanobacteria. Here we report that a cAMP-dependent transcription factor SyCRP1 mediates C i response in Synechocystis. The mutant ∆ sycrp1 exhibited a slow-growth phenotype and reduced maximum rate of bicarbonate-dependent photosynthetic electron transport (V max) compared to wild-type at the scarcity of CO 2. The number of carboxysomes was decreased significantly in the ∆ sycrp1 at low CO 2 consistent with its reduced V max. The DNA microarray analysis revealed the upregulation of genes encoding C i transporters in ∆sycrp1. The membrane-localized SyCRP1 was released into the cytosol in wild-type cells shifted from low to high CO 2 or upon cAMP treatment. Soluble His-tagged SyCRP1 was shown to target DNA-binding sites upstream of the C i -regulated genes sbtA and ccmK 3. In addition, cAMP enhanced the binding of SyCRP1 to its target sites. Our data collectively suggest that the C i is sensed through the second messenger cAMP releasing membrane-bound SyCRP1 into cytoplasm under sufficient CO 2 conditions. Hence, SyCRP1 is a possible regulator of carbon concentrating mechanism, and such a regulation might be mediated via sensing C i levels through cAMP in Synechocystis. • It has been known that cAMP receptor proteins (CRP) play a crucial role in regulation of gene expression in bacteria. • Transcription regulators of CCM genes were decribed previously, however C i -sensing mechanism in cyanobacteria is unclear. • A membrane associated Synechocystis CRP (SyCRP1), releases to cytosol when bound to CAMP to regulate CCM. • Thus, SyCRP1 mediates C i response through the signalling molecule cAMP in Synechocytis. [ABSTRACT FROM AUTHOR]

Microalgae are promising alternative feedstocks that can be used to produce biofuels and replace conventional fossil fuels. In order to increase the lipid production of microalgae, strain development via genetic engineering is required. To date, metabolic engineering studies have mainly focused on lipid synthesis pathways. However, the complicated metabolic pathways that exist in many organelles along with limitations of genetic engineering tools have made it difficult to obtain desirable microalgal strains. As an alternative strategy, we tried to increase carbon flux into fatty acid synthesis by overexpressing the NADP-dependent malic enzyme, NsME1, in Nannochloropsis salina. We found that the biomass and fatty acid methyl ester (FAME) contents were increased in NsME1 overexpressing transformants, such that the FAME yield of the top-producing NsME1 transformant was 53% higher than the wild type. To understand the effects of NsME1 on lipid production, we analyzed the total carbon concentration and NADPH/NADP ratio, which were found to be enhanced in the transformants. We also investigated mRNA expression levels of genes involved in C4-like carbon concentrating mechanism and fatty acid synthesis by quantitative real-time PCR, and confirmed their positive contribution to fatty acid production. Taken together, our results demonstrate that overexpression of NsME1 could simultaneously improve the carbon concentration and reducing power in cells, thereby increasing the lipid and FAME yields of Nannochloropsis salina. We suggest that NsME1 can serve as a promising genetic target for enhancing lipid-based bio-products in Nannochloropsis and potentially other industrial microalgae. • NsME1 overexpression affected total carbon concentration in N. salina culture. • NsME1 overexpression increased NADPH/NADP ratio. • NsME1 overexpression increased dry cell weight and fatty acid contents. [ABSTRACT FROM AUTHOR]

• Responses of R. raciborskii to different pH/CO 2 scenarios were tested;. • Bicarbonate availability governed growth and saxitoxin production in the cyanobacterium;. • R. raciborskii ITEP-A1 is a bicarbonate-user (potentially expresses the BicA transporter);. • Dissolved inorganic carbon and pH influenced saxitoxins production and toxicity in R. raciborskii ITEP-A1. Raphidiopsis raciborskii is a widely distributed, potentially toxic cyanobacterium described as a tropical-subtropical species. However, its occurrence in temperate regions has been expanding. Understanding the environmental factors underlying the expansion and colonization success of Raphidiopsis has been the object of numerous studies. However, less is known regarding its responses to pH and inorganic carbon in water. Thus, the aim of the present study was to investigate the effects of changes in pH and dissolved inorganic carbon on growth and saxitoxins production in the strain R. raciborskii ITEP-A1. We incubated batch cultures with different unbuffered and buffered pH (neutral-acid and alkaline) and inorganic carbon availability (CO 2 -rich air bubbling and the addition of NaHCO 3) to assess the effect of these factors on the growth, toxin production as well as saxitoxins composition of the cyanobacterium. The carbon concentrating mechanism (CCM) system of ITEP-A1 was also characterized by an in silico analysis of its previously sequenced genome. The growth and saxitoxins production of R. raciborskii were affected. The addition of sodium bicarbonate and air bubbling enhanced the growth of the cyanobacterium in alkaline pH. In contrast, saxitoxins production and relative toxicity were decreased. Moreover, significant changes in the cellular composition of saxitoxins were strongly related to pH changes. ITEP-A1 potentially expresses the low-flux bicarbonate transporter BicA, an efficient CCM which uptakes most of its carbon from HCO 3 −. Hence, increasing the diffusion of CO 2 in alkaline eutrophic lakes is likely to increase R. raciborskii dominance , but produce less toxic blooms. Image, graphical abstract [ABSTRACT FROM AUTHOR]