Chromosome-level genome assembly and transcriptome of the green alga Chromochloris zofingiensis illuminates astaxanthin production.

Microalgae have potential to help meet energy and food demands without exacerbating environmental problems. There is interest in the unicellular green alga Chromochloris zofingiensis, because it produces lipids for biofuels and a highly valuable carotenoid nutraceutical, astaxanthin. To advance understanding of its biology and facilitate commercial development, we present a C. zofingiensis chromosome-level nuclear genome, organelle genomes, and transcriptome from diverse growth conditions. The assembly, derived from a combination of short- and long-read sequencing in conjunction with optical mapping, revealed a compact genome of ∼58 Mbp distributed over 19 chromosomes containing 15,274 predicted protein-coding genes. The genome has uniform gene density over chromosomes, low repetitive sequence content (∼6%), and a high fraction of protein-coding sequence (∼39%) with relatively long coding exons and few coding introns. Functional annotation of gene models identified orthologous families for the majority (∼73%) of genes. Synteny analysis uncovered localized but scrambled blocks of genes in putative orthologous relationships with other green algae. Two genes encoding beta-ketolase (BKT), the key enzyme synthesizing astaxanthin, were found in the genome, and both were up-regulated by high light. Isolation and molecular analysis of astaxanthin-deficient mutants showed that BKT1 is required for the production of astaxanthin. Moreover, the transcriptome under high light exposure revealed candidate genes that could be involved in critical yet missing steps of astaxanthin biosynthesis, including ABC transporters, cytochrome P450 enzymes, and an acyltransferase. The high-quality genome and transcriptome provide insight into the green algal lineage and carotenoid production.

Construction of a fusion enzyme for astaxanthin formation and its characterisation in microbial and plant hosts: A new tool for engineering ketocarotenoids.

The high-value ketocarotenoid astaxanthin, a natural red colorant with powerful antioxidant activity, is synthesised from ß-carotene by a hydroxylase and an oxygenase enzyme, which perform the addition of two hydroxyl and keto moieties, respectively. Several routes of intermediates, depending on the sequence of action of these enzymes, lead to the formation of astaxanthin. In the present study, the enzyme activities of 3, 3' ß-carotene hydroxylase (CRTZ) and 4, 4' ß-carotene oxygenase (CRTW) have been combined through the creation of "new to nature" enzyme fusions in order to overcome leakage of non-endogenous intermediates and pleotropic effects associated with their high levels in plants. The utility of flexible linker sequences of varying size has been assessed in the construction of pZ-W enzyme fusions. Frist, in vivo color complementation assays in Escherichia coli have been used to evaluate the potential of the fusion enzymes. Analysis of the carotenoid pigments present in strains generated indicated that the enzyme fusions only possess both catalytic activities when CRTZ is attached as the N-terminal module. Astaxanthin levels in E. coli cells were increased by 1.4-fold when the CRTZ and CRTW enzymes were fused compared to the individual enzymes. Transient expression in Nicotiana benthamiana was then performed in order to assess the potential of the fusions in a plant system. The production of valuable ketocarotenoids was achieved using this plant-based transient expression system. This revealed that CRTZ and CRTW, transiently expressed as a fusion, accumulated similar levels of astaxanthin compared to the expression of the individual enzymes whilst being associated with reduced ketocarotenoid intermediate levels (e.g. phoenicoxanthin, canthaxanthin and 3-OH-echinenone) and a reduced rate of leaf senescence after transformation. Therefore, the quality of the plant material producing the ketocarotenoids was enhanced due to a reduction in the stress induced by the accumulation of high levels of heterologous ketocarotenoid intermediates. The size of the linkers appeared to have no effect upon activity. The potential of the approach to production of valuable plant derived products is discussed.

Stepwise pathway engineering to the biosynthesis of zeaxanthin, astaxanthin and capsanthin in rice endosperm.

Carotenoid pigments are valuable components of the human diet. A notable example is ß-carotene, or provitamin A, which is converted into the derivatives astaxanthin and capsanthin, via the common intermediate zeaxanthin. To generate rice varieties producing diverse carotenoids beyond ß-carotene, we specifically used a Capsicum ß-carotene hydroxylase gene, B (CaBch) and a codon optimized version of the same gene, stB (stBch) to increase zeaxanthin synthesis. We also used a recombinant BAK gene (CaBch-2A-HpBkt), consisting of the CaBch sequence and a Haematococcus ß-carotene ketolase gene (HpBkt) linked by a bicistronic 2 A sequence, as well as a codon optimized recombinant stBAK gene (stBch-2A-stBkt) to create astaxanthin synthesis. The four cassettes to seed-specifically express the B, stB, BAK and stBAK genes were individually combined with a PAC gene (CaPsy-2A-PaCrtI) cassette to previously impart ß-carotene-enriched trait in rice endosperm. The single T-DNA vectors of B-PAC, stB-PAC, BAK-PAC and stBAK-PAC resulted in the accumulation of zeaxanthin and astaxanthin in the endosperm of the transgenic rice seeds. In addition, an extended version on the carotenoid pathway was introduced into rice to allow the production of capsanthin, by intercrossing a B-PAC rice line with a Ccs rice line, which harbors a Capsicum capsanthin-capsorubin synthase gene. Ultimately, we developed three functional rice varieties: B-PAC (0.8 µg/g zeaxanthin, deep yellow), stBAK-PAC (1.4 µg/g ketocarotenoids, including astaxanthin, pinkish red) and B-PAC x Ccs (0.4 µg/g of ketoxanthophylls, including capsanthin, orange-red) with the similar levels of total carotenoids to PAC rice, suggesting the capacity was dependent on ß-carotene levels. Collectively, a combination of genetic engineering and conventional breeding is effective for multi-step metabolic engineering and biochemical pathway extension.

Illustrating and Enhancing the Biosynthesis of Astaxanthin and Docosahexaenoic Acid in Aurantiochytrium sp. SK4.

The marine thraustochytrids are a promising source of docosahexaenoic acid (DHA) and the ketocarotenoid astaxanthin. In this study, the biosynthetic pathways of these two important metabolites in Aurantiochytrium sp. SK4 was illustrated by the analyses of the genome, transcriptome, key enzymes, and pathway products. Two sets of genes were involved in two pathways for the biosynthesis of fatty acids. The absence of Δ-15 desaturase genes and the presence of docosapentaenoic acid (DPA), up to 12% of total fatty acids suggest that Aurantiochytrium sp. SK4 may synthesize DHA mainly via a polyketide synthase (PKS) pathway. Three enzymes, namely geranyl diphosphate synthase (GPPS), farnysyl diphosphate synthase (FPPS), and geranylgeranyle diphosphate synthase (GGPPS) were found to be involved in the formation of GGPP that was subsequently catalyzed to ß-carotene by a trifunctional CrtIBY enzyme. ß-Carotene might be ketolated and then hydroxylated into astaxanthin based on the carotenoid profiles. The formation of GGPP was proposed to be the limiting steps for carotenoid production. Overexpression of the Archaeoglobus GPS together with the Escherichia coli isopentenyl pyrophosphate isomerase, and Vitreoscilla hemoglobin resulted in not only 1.85- and 5.02-fold increases of total carotenoids and astaxanthin, but also 2.40- and 2.74-fold increases of total fatty acids and DHA. This study provides insights into the biosynthesis of carotenoids and fatty acids in Aurantiochytrium.

Role of media composition in biomass and astaxanthin production of Haematococcus pluvialis under two-stage cultivation.

In the present study, the effects of four different culture media on the growth, astaxanthin production and morphology of Haematococcus pluvialis LUGU were studied under two-step cultivation. The interactions between astaxanthin synthesis and secondary messengers, reactive oxygen species (ROS) and mitogen-activated protein kinases (MAPK) were also investigated. In the first green vegetative cell stage, maximal biomass productivity (86.54 mg L-1 day-1) was obtained in BBM medium. In the induction stage, the highest astaxanthin content (21.5 mg g-1) occurred in BG-11 medium, which was higher than in any other media. The expressions of MAPK and astaxanthin biosynthetic genes in BG-11 were higher than in any other media, whereas the ROS content was lower. Biochemical and physiological analyses suggested that the ROS, MAPK and astaxanthin biosynthetic gene expression was involved in astaxanthin biosynthesis in H. pluvialis under different culture media conditions. This study proposes a two-step cultivation strategy to efficiently produce astaxanthin using microalgae.

Metabolic engineering of Escherichia coli for high-level astaxanthin production with high productivity.

Astaxanthin is a reddish keto-carotenoid classified as a xanthophyll found in various microbes and marine organisms. As a powerful antioxidant having up to 100 times more potency than other carotenoids such as ß-carotene, lutein, and lycopene, astaxanthin is a versatile compound utilized in animal feed, food pigment, health promotion and cosmetic industry. Here, we report development of metabolically engineered Escherichia coli capable of producing astaxanthin to a high concentration with high productivity. First, the heterologous crt genes (crtE, crtY, crtI, crtB, and crtZ) from Pantoea ananatis and the truncated BKT gene (trCrBKT) from Chlamydomonas reinhardtii were introduced to construct the astaxanthin biosynthetic pathway. Then, eight different fusion tags were examined by attaching them to the N- or C-terminus of the trCrBKT membrane protein to allow stable expression and to efficiently guide trCrBKT to the E. coli membrane. When the signal peptide of OmpF and TrxA were tagged to the N-terminus and C-terminus of trCrBKT, respectively, astaxanthin production reached 12.90 mg/L (equivalent to 3.84 mg/gDCW), which was 2.08-fold higher than that obtained without tagging. Upon optimization of culture conditions, this engineered strain WLGB-RPP harboring pAX15 produced 332.23 mg/L (5.38 mg/gDCW) of astaxanthin with the productivity of 3.79 mg/L/h by fed-batch fermentation. In order to further increase astaxanthin production, in silico flux variability scanning based on enforced objective flux (FVSEOF) was performed to identify gene overexpression targets. The engineered strain WLGB-RPP (pAX15, pTrc-ispDF) which simultaneously overexpressing the ispD and ispF genes identified by FVSEOF produced astaxanthin to a higher concentration of 377.10 mg/L (6.26 mg/gDCW) with a productivity of 9.20 mg/L/h upon induction with 1 mM IPTG. When cells were induced with 0.5 mM IPTG to reduce the metabolic burden, astaxanthin concentration further increased to 432.82 mg/L (7.12 mg/gDCW) with a productivity of 9.62 mg/L/h. To more stably maintain plasmid during the fed-batch fermentation of WLGB-RPP (pAX15, pTrc-ispDF), the post-segregational killing hok/sok system was introduced. This strain produced 385.04 mg/L (6.98 mg/gDCW) of astaxanthin with a productivity of 7.86 mg/L/h upon induction with 0.5 mM IPTG. The strategies reported here will be useful for the enhanced production of astaxanthin and related carotenoid products by engineered E. coli strains.

Essential oils from unexplored aromatic plants quench biofilm formation and virulence of Methicillin resistant Staphylococcus aureus.

In the current study we have evaluated the antibiofilm and antivirulent properties of unexplored essential oils (EOs) obtained from Pogostemon heyneanus and Cinnamomum tamala against Methicillin Resistant Staphylococcus aureus (MRSA) strains. The EOs from both the aromatic plants was screened for their ability to prevent biofilm formation and to disrupt preformed biofilms. The efficacy of both the EOs to disrupt the preformed biofilms of various MRSA strains was determined by Confocal Laser Scanning Microscopy (CLSM) and Scanning Electron Microscopy (SEM).The EOs were further able to reduce the Extracellular polymeric substance (EPS) and slime synthesis the two factors of the biofilm assemblage. The EOs was also found to be effective in reducing virulence factors like staphyloxanthin and hemolysin. In silico docking studies were performed for the major components of essential oils and dehydroxysqualene synthase of MRSA which is responsible for the synthesis of staphyloxanthin. The results suggest that (E)-nerolidol showed better binding affinity towards the enzyme. Other compounds have similar binding strengths with the enzyme. Furthermore, the synergistic effect EOs along with the commercially available DNaseI and Marine Bacterial DNase (MBD) showed that the synergistic effect had enhanced biofilm disruption ability. The results show that EOs from P. heyneanus and C. tamala has potential antivirulent and biofilm inhibitory properties against clinical and drug resistant S. aureus strains. The present study highlights the importance of bioprospecting plant based natural products as an alternative for antibiotics owing to the emergence of multi-drug resistant strains.

Network analytics approach towards identifying potential antivirulence drug targets within the Staphylococcus aureus staphyloxanthin biosynthetic network.

Staphylococcus aureus is associated with several clinically significant infections among humans and infections associated with antibiotic-resistant strains are growing in frequency. Antivirulence strategies shift the target of drugs from bacterial growth to the infection process resulting to milder evolutionary pressure for the development of bacterial resistant strains. Staphyloxanthin (STX) is a yellowish-orange carotenoid pigment synthesized by S. aureus and this carotenoid functions as an important virulence factor for the bacteria. In this study, we elucidated whether network analytics can be used as a viable tool to identify significant components in the STX biosynthetic network which in-turn could serve as possible antivirulence drug targets. For confirmation, we correlated our results to known drugs that were able to inhibit STX biosynthesis. Throughout this study, we established that crtN(1) activity and 4,4'-diaponeurosporene amounts are significant components in the STX biosynthetic network and, moreover, network analytics can aid in identifying antivirulence drug targets within the STX biosynthetic network. Similarly, we found that network analytics is capable of identifying multiple potential targets simultaneously. Taken together, we propose that an effective antivirulence drug against S. aureus STX biosynthesis would involve targeting crtN(1) activity, 4,4'-diaponeurosporene levels, or both components.

Small-molecule targeting of a diapophytoene desaturase inhibits S. aureus virulence.

The surge of antibiotic resistance in Staphylococcus aureus has created a dire need for innovative anti-infective agents that attack new targets, to overcome resistance. In S. aureus, carotenoid pigment is an important virulence factor because it shields the bacterium from host oxidant killing. Here we show that naftifine, a US Food and Drug Administration (FDA)-approved antifungal drug, blocks biosynthesis of carotenoid pigment at nanomolar concentrations. This effect is mediated by competitive inhibition of S. aureus diapophytoene desaturase (CrtN), an essential enzyme for carotenoid pigment synthesis. We found that naftifine attenuated the virulence of a variety of clinical S. aureus isolates, including methicillin-resistant S. aureus (MRSA) strains, in mouse infection models. Specifically, we determined that naftifine is a lead compound for potent CrtN inhibitors. In sum, these findings reveal that naftifine could serve as a chemical probe to manipulate CrtN activity, providing proof of concept that CrtN is a druggable target against S. aureus infections.

Changes in the fucoxanthin production and protein profiles in Cylindrotheca closterium in response to blue light-emitting diode light.

BACKGROUND: Marine diatoms have a higher fucoxanthin content in comparison to macroalgae. Fucoxanthin features many potent bioactive properties, particularly anti-obesity properties. Despite the great potential for harvesting larger amounts of fucoxanthin, the impacts of light quality (light source, intensity, and photoperiod) on fucoxanthin production and the essential proteins involved in fucoxanthin biosynthesis in marine diatoms remain unclear. RESULTS: In the present study, Cylindrotheca closterium was selected from four different species of diatoms based on its high fucoxanthin content and productivity. Optimal light conditions (light source, intensity, and regime) were determined by a "Design of Experiment" approach (software MODDE Pro 11 was used). The model indicated that an 18/6 light/darkness regime increased fucoxanthin productivity remarkably as opposed to a 12/12 or 24/0 regime. Eventually, blue light-emitting diode light, as an alternative to fluorescent light, at 100 µmol/m2/s and 18/6 light/darkness regime yielded maximum fucoxanthin productivity and minimal energy consumption. The fucoxanthin production of C. closterium under the predicted optimal light conditions was assessed both in bottle and bag photobioreactors (PBRs). The high fucoxanthin content (25.5 mg/g) obtained from bag PBRs demonstrated the feasibility of large-scale production. The proteomes of C. closterium under the most favorable and unfavorable fucoxanthin biosynthesis light/darkness regimes (18/6 and 24/0, respectively) were compared to identify the essential proteins associated with fucoxanthin accumulation by matrix-assisted laser desorption/ionization-time of flight-mass spectrometry. Six proteins that were up-regulated in the 18/6 regime but down-regulated in the 24/0 were identified as important chloroplastic proteins involved in photosynthesis, energy metabolism, and cellular processes. CONCLUSIONS: Blue light-emitting diode light at 100 µmol/m2/s and 18/6 light/darkness regime induced maximum fucoxanthin productivity in C. closterium and minimized energy consumption. The high fucoxanthin production in the bag photobioreactor under optimal light conditions demonstrated the possibility of commercialization. Proteomics suggests that fucoxanthin biosynthesis is intimately associated with the photosynthetic efficiency of the diatom, providing another technical and bioengineering outlook on fucoxanthin enhancement.

A common mechanism explains the induction of aerobic fermentation and adaptive antioxidant response in Phaffia rhodozyma.

BACKGROUND: Growth conditions that bring about stress on Phaffia rhodozyma cells encourage the synthesis of astaxanthin, an antioxidant carotenoid, which protects cells against oxidative damage. Using P. rhodozyma cultures performed with and without copper limitation, we examined the kinetics of astaxanthin synthesis along with the expression of asy, the key astaxanthin synthesis gene, as well as aox, which encodes an alternative oxidase protein. RESULTS: Copper deficiency had a detrimental effect on the rates of oxygen consumption and ethanol reassimilation at the diauxic shift. In contrast, copper deficiency prompted alcoholic fermentation under aerobic conditions and had a favorable effect on the astaxanthin content of cells, as well as on aox expression. Both cultures exhibited strong aox expression while consuming ethanol, but particularly when copper was absent. CONCLUSION: We show that the induction of either astaxanthin production, aox expression, or aerobic fermentation exemplifies the crucial role that redox imbalance plays in triggering any of these phenomena. Based on our own results and data from others, we propose a mechanism that rationalizes the central role played by changes of respiratory activity, which lead to redox imbalances, in triggering both the short-term antioxidant response as well as fermentation in yeasts and other cell types.

Food Modulation Controls Astaxanthin Accumulation in Eggs of the Sea Urchin Arbacia lixula.

The carotenoid astaxanthin has strong antioxidant properties with beneficial effects for various degenerative diseases. This carotenoid is produced by some microalgae species when cultivated in particular conditions, and, interestingly, it is a predominant carotenoid in aquatic animals throughout a broad range of taxa. Recently, astaxanthin was detected in the eggs of the sea urchin Arbacia lixula in relevant concentrations when this organism was maintained in culture. These results have paved the way for deeper research into astaxanthin production by this species, particularly in regards to how astaxanthin production can be modulated by diet. Results showed that the highest content of astaxanthin in eggs was observed in sea urchins fed on a diet enriched with Spirulina platensis. This result was confirmed by the high antioxidant activity recorded in the egg extracts of these animals. Our results suggest that (i) the sea urchin A. lixula is able to synthesize astaxanthin from precursors obtained from food, and (ii) it is possible to modulate the astaxanthin accumulation in sea urchin eggs by modifying the proportions of different food ingredients provided in their diet. This study demonstrates the large potential of sea urchin cultivation for the eco-sustainable production of healthy supplements for nutraceutical applications.

A Hetero-Photoautotrophic Two-Stage Cultivation Process for Production of Fucoxanthin by the Marine Diatom Nitzschia laevis.

There is currently much interest in fucoxanthin due to its broad beneficial health effects. The major commercial source of fucoxanthin is marine seaweed, which has many shortcomings, and has thus restricted its large-scale production and more diversified applications. In this study, growth characteristics and fucoxanthin accumulation were evaluated to explore the potential of the marine diatom Nitzschia laevis in fucoxanthin production. The results suggested that heterotrophic culture was more effective for cell growth, while the mixotrophic culture was favorable for fucoxanthin accumulation. A two-stage culture strategy was consequently established. A model of exponential fed-batch culture led to a biomass concentration of 17.25 g/L. A mix of white and blue light significantly increased fucoxanthin content. These outcomes were translated into a superior fucoxanthin productivity of 16.5 mg/(L·d), which was more than 2-fold of the best value reported thus far. The culture method established herein therefore represents a promising strategy to boost fucoxanthin production in N. laevis, which might prove to be a valuable natural source of commercial fucoxanthin.

Optimum Production Conditions, Purification, Identification, and Antioxidant Activity of Violaxanthin from Microalga Eustigmatos cf. polyphem (Eustigmatophyceae).

Violaxanthin is a major xanthophyll pigment in the microalga Eustigmatos cf. polyphem, but the amount produced after propagation can vary depending upon culture conditions. In this study, the effects of cultivation time, nitrogen concentration, light intensity, and culture mode on violaxanthin production were investigated. The results showed that this microalga vigorously grew and maintained a high level of violaxanthin in the fed-batch culture, and the highest violaxanthin productivity of 1.10 ± 0.03 mg L-1 d-1 was obtained under low light illumination with 18 mM of initial nitrogen supply for ten days. Additionally, violaxanthin was purified from E. cf. polyphem by silica gel chromatography and preparative high-performance liquid chromatography (PHPLC), and identified with high-resolution mass spectrometry (HRMS). The antioxidant activity of the purified violaxanthin was evaluated by three tests in vitro: reducing power assay, 2,2-diphenyl-1-picrylhydrazyl (DPPH), and 2,2-azobis-3-ethylbenzthiazoline-6-sulphonic acid (ABTS) radical assays. The strongest inhibition of purified violaxanthin occurred during the scavenging of ABTS⁺ radicals, with EC50 of 15.25 µg mL-1. In conclusion, this is the first report to investigate the effects of different culture conditions on violaxanthin accumulation in E. cf. polyphem and provide a novel source for the production of violaxanthin that can be used for food and pharmaceutical applications.

Combined production of fucoxanthin and EPA from two diatom strains Phaeodactylum tricornutum and Cylindrotheca fusiformis cultures.

Fucoxanthin and eicosapentaenoic acid (EPA) provide significant health benefits for human population. Diatom is a potential natural livestock for the combined production of EPA and fucoxanthin. In this study, first, the effects of three important parameters including light intensity, nitrogen concentration and salinity were evaluated for the production of EPA and fucoxanthin in two diatom strains Phaeodactylum tricornutum and Cylindrotheca fusiformis. And then, two steps method based on light intensity were applied to produce EPA and fucoxanthin in large scale. Higher light intensity was first adopted for the high growth rate and lipid content of diatom, and after a period of time, light intensity was lowered to enhance the accumulation of fucoxanthin and EPA. In final, the highest EPA yields were 62.55 and 27.32 mg L-1 for P. tricornutum and C. fusiformis, and the fucoxanthin yield reached 8.32 and 6.05 mg L-1, respectively.

Transcriptome Analysis in Haematococcus pluvialis: Astaxanthin Induction by High Light with Acetate and Fe2.

Haematococcus pluvialis is a commercial microalga, that produces abundant levels of astaxanthin under stress conditions. Acetate and Fe2+ are reported to be important for astaxanthin accumulation in H. pluvialis. In order to study the synergistic effects of high light stress and these two factors, we obtained transcriptomes for four groups: high light irradiation (HL), addition of 25 mM acetate under high light (HA), addition of 20 µM Fe2+ under high light (HF) and normal green growing cells (HG). Among the total clean reads of the four groups, 156,992 unigenes were found, of which 48.88% were annotated in at least one database (Nr, Nt, Pfam, KOG/COG, SwissProt, KEGG, GO). The statistics for DEGs (differentially expressed genes) showed that there were more than 10 thousand DEGs caused by high light and 1800-1900 DEGs caused by acetate or Fe2+. The results of DEG analysis by GO and KEGG enrichments showed that, under the high light condition, the expression of genes related to many pathways had changed, such as the pathway for carotenoid biosynthesis, fatty acid elongation, photosynthesis-antenna proteins, carbon fixation in photosynthetic organisms and so on. Addition of acetate under high light significantly promoted the expression of key genes related to the pathways for carotenoid biosynthesis and fatty acid elongation. Furthermore, acetate could obviously inhibit the expression of genes related to the pathway for photosynthesis-antenna proteins. For addition of Fe2+, the genes related to photosynthesis-antenna proteins were promoted significantly and there was no obvious change in the gene expressions related to carotenoid and fatty acid synthesis.

Enhancement of astaxanthin production from Haematococcus pluvialis under autotrophic growth conditions by a sequential stress strategy.

The study of microalgal culture has been growing in recent decades, because the cellular structure of microalgae has diverse highly valuable metabolites that have attract attention of numerous companies and research groups. The pigment astaxanthin is considered one of the most powerful antioxidants in nature. The microalga Haematococcus pluvialis was proposed as one of the best natural astaxanthin sources, because it can accumulate high amount of the pigment. In this work, we studied different stress treatments on H. pluvialis growth cultures as well as astaxanthin production under autotrophic growth conditions. The results showed that extending nitrogen starvation before increasing radiation intensity up to 110 µmol photons m-2 s-1 during late the palmella cell phase incremented the astaxanthin concentration up to 2.7% of dry biomass with an efficient light energy utilization during the stress stage.

Comparative transcriptome analysis of Haematococcus pluvialis on astaxanthin biosynthesis in response to irradiation with red or blue LED wavelength.

The unicellular green microalga Haematococcus pluvialis has the highest content of the natural antioxidant, astaxanthin. Previously, it was determined that astaxanthin accumulation in H. pluvialis could be induced by blue-wavelength irradiation; however, the molecular mechanism remains unknown. The present study aimed to compare the transcriptome of H. pluvialis, with respect to astaxanthin biosynthesis, under the monochromatic red (660 nm) or blue (450 nm) light-emitting diode (LED) irradiation. Among a total of 165,372 transcripts, we identified 67,703 unigenes, of which 2245 and 171 were identified as differentially expressed genes (DEGs) in response to blue and red irradiation, respectively. Interestingly, expressional changes of blue light receptor cryptochromes were detected in response to blue and/or red LED irradiation in H. pluvialis, which may directly and indirectly regulate astaxanthin biosynthesis. In accordance with this observation, expression of the BKT and CHY genes, which are part of the downstream section of the astaxanthin biosynthetic pathway, was significantly upregulated by blue LED irradiation compared with their expression under control white irradiation. Contrastingly, they were downregulated by red LED irradiation. Our transcriptome study provided molecular insights that highlighted the different of responses of H. pluvialis to red and blue irradiation, especially for astaxanthin biosynthesis.

The Staphylococcus aureus AirSR Two-Component System Mediates Reactive Oxygen Species Resistance via Transcriptional Regulation of Staphyloxanthin Production.

Staphylococcus aureus is an important opportunistic pathogen and is the etiological agent of many hospital- and community-acquired infections. The golden pigment, staphyloxanthin, of S. aureus colonies distinguishes it from other staphylococci and related Gram-positive cocci. Staphyloxanthin is the product of a series of biosynthetic steps that produce a unique membrane-embedded C30 golden carotenoid and is an important antioxidant. We observed that a strain with an inducible airR overexpression cassette had noticeably increased staphyloxanthin production compared to the wild-type strain under aerobic culturing conditions. Further analysis revealed that depletion or overproduction of the AirR response regulator resulted in a corresponding decrease or increase in staphyloxanthin production and susceptibility to killing by hydrogen peroxide, respectively. Furthermore, the genetic elimination of staphyloxanthin during AirR overproduction abolished the protective phenotype of increased staphyloxanthin production in a whole-blood survival assay. Promoter reporter and gel shift assays determined that the AirR response regulator is a direct positive regulator of the staphyloxanthin-biosynthetic operon, crtOPQMN, but is epistatic to alternative sigma factor B. Taken together, these data indicate that AirSR positively regulates the staphyloxanthin-biosynthetic operon crtOPQMN, promoting survival of S. aureus in the presence of oxidants.

The diatom Phaeodactylum tricornutum adjusts nonphotochemical fluorescence quenching capacity in response to dynamic light via fine-tuned Lhcx and xanthophyll cycle pigment synthesis.

Diatoms contain a highly flexible capacity to dissipate excessively absorbed light by nonphotochemical fluorescence quenching (NPQ) based on the light-induced conversion of diadinoxanthin (Dd) into diatoxanthin (Dt) and the presence of Lhcx proteins. Their NPQ fine regulation on the molecular level upon a shift to dynamic light conditions is unknown. We investigated the regulation of Dd + Dt amount, Lhcx gene and protein synthesis and NPQ capacity in the diatom Phaeodactylum tricornutum after a change from continuous low light to 3 d of sine (SL) or fluctuating (FL) light conditions. Four P. tricornutum strains with different NPQ capacities due to different expression of Lhcx1 were included. All strains responded to dynamic light comparably, independently of initial NPQ capacity. During SL, NPQ capacity was strongly enhanced due to a gradual increase of Lhcx2 and Dd + Dt amount. During FL, cells enhanced their NPQ capacity on the first day due to increased Dd + Dt, Lhcx2 and Lhcx3; already by the second day light acclimation was accomplished. While quenching efficiency of Dt was strongly lowered during SL conditions, it remained high throughout the whole FL exposure. Our results highlight a more balanced and cost-effective photoacclimation strategy of P. tricornutum under FL than under SL conditions.