Β-carotene Accumulation Research Articles

Quantifying the Impact of Nitrogen Concentration Variations on Quality Characteristics of Soilless Cultivated Cherry Tomato

Within the domain of macronutrients, nitrogen has garnered significant recognition as a pivotal element due to its profound influence on both the vegetative growth and quality attributes of a crop. An abundance of scientific research has consistently demonstrated that attaining optimal productivity necessitates the meticulous management of soil moisture availability, complemented by the timely provision of nitrogen during critical growth stage intervals. The study aimed to optimize the nitrogen concentration in a hydroponic solution by testing a range of concentrations. The objective was to systematically examine the effects of different nitrogen levels and determine the optimal concentration for the desired outcomes. The assessment was carried out in a meticulously controlled greenhouse environment from October 2019 to April 2021, with the primary objective of systematically investigating the impact on quality characteristics specific to the rosa variety of cherry tomato. To investigate the effects of different nitrogen concentrations, four distinct levels of 4 mM, 6 mM, 8 mM, and 10 mM per litre of nutrient solution were employed. The nitrogen source consisted of a combination of NO3- and NH4+, with a ratio of 40:60, respectively. The nutrient solution with an 8 mM nitrogen concentration exhibited a significantly augmented effect on increasing the equatorial and polar diameter of fruits, average fruit weight, pericarp thickness, as well as the accumulation of total chlorophyll, ascorbic acid, lycopene, β-carotene, and total soluble solids (TSS) content. These findings demonstrate the pronounced and positive influence of the 8 mM nitrogen concentration on the aforementioned quality characteristics.

Unbalanced Expression of Structural Genes in Carotenoid Pathway Contributes to the Flower Color Formation of the Osmanthus Cultivar 'Yanzhi Hong'.

Carotenoids are important natural pigments that are responsible for the fruit and flower colors of many plants. The composition and content of carotenoid can greatly influence the color phenotype of plants. However, the regulatory mechanism underling the divergent behaviors of carotenoid accumulation, especially in flower, remains unclear. In this study, a new cultivar Osmanthus fragrans 'Yanzhi Hong' was used to study the regulation of carotenoid pigmentation in flower. Liquid chromatograph-mass spectrometer (LC-MS) analysis showed that β-carotene, phytoene, lycopene, γ-carotene, and lutein were the top five pigments enriched in the petals of 'Yanzhi Hong'. Through transcriptome analysis, we found that the expression of the structural genes in carotenoid pathway was imbalanced: most of the structural genes responsible for lycopene biosynthesis were highly expressed throughout the flower developmental stages, while those for lycopene metabolism kept at a relatively lower level. The downregulation of LYCE, especially at the late developmental stages, suppressed the conversion from lycopene to α-carotene but promoted the accumulation of β-carotene, which had great effect on the carotenoid composition of 'Yanzhi Hong'. Ethylene response factor (ERF), WRKY, basic helix-loop-helix (bHLH), v-myb avian myeloblastosis viral oncogene homolog (MYB), N-Acetylcysteine (NAC), auxin response factor (ARF), and other transcription factors (TFs) have participated in the flower color regulation of 'Yanzhi Hong', which formed co-expression networks with the structural genes and functioned in multiple links of the carotenoid pathway. The results suggested that the cyclization of lycopene is a key link in determining flower color. The modification of the related TFs will break the expression balance between the upstream and downstream genes and greatly influence the carotenoid profile in flowers, which can be further used for creating colorful plant germplasms.

Retargeting of heterologous enzymes results in improved β-carotene synthesis in Saccharomyces cerevisiae.

Carotenoids are a class of hydrophobic substances that are important as food and feed colorants and as antioxidants. The pathway for β-carotene synthesis has been expressed in various yeast species, albeit with rather low yields and titers. The inefficient conversion of phytoene to lycopene is often regarded as a bottleneck in the pathway. In this study, we aimed at the improvement of β-carotene production in Saccharomyces cerevisiae by specifically engineering the enzymatic reactions producing and converting phytoene. We show that phytoene is stored in intracellular lipid droplets, whereas the enzyme responsible for its conversion, phytoene dehydrogenase, CrtI, is located at the endoplasmic reticulum, like the bifunctional enzyme CrtYB that catalyses the reaction before and after CrtI. To improve the accessibility of phytoene for CrtI and to delay its storage in lipid droplets, we tested the relocation of CrtI and CrtYB to mitochondria. However, only the retargeting of CrtYB resulted in an improvement of the β-carotene content, whereas the mitochondrial variant of CrtI was not functional. Surprisingly, a cytosolic variant of this enzyme, which we obtained through the elimination of its carboxy-terminal membrane anchor, caused an increase in β-carotene accumulation. Overexpression of this CrtI variant in an optimized medium resulted in a strain with a β-carotene content of 79 mg g-1 cell dry weight, corresponding to a 76-fold improvement over the starting strain. The retargeting of heterologously expressed pathway enzymes improves β-carotene production in S. cerevisiae, implicating extensive inter-organellar transport phenomena of carotenoid precursors. In addition, strong overexpression of carotenoid biosynthetic enzymes and the optimization of cultivation conditions are required for high contents.

Boosting pro-vitamin A content and bioaccessibility in leaves by combining engineered biosynthesis and storage pathways with high-light treatments.

Biofortification of green leafy vegetables with pro-vitamin A carotenoids, such as β-carotene, has remained challenging to date. Here, we combined two strategies to achieve this goal. One of them involves producing β-carotene in the cytosol of leaf cells to avoid the negative impacts on photosynthesis derived from changing the balance of carotenoids and chlorophylls in chloroplasts. The second approach involves the conversion of chloroplasts into non-photosynthetic, carotenoid-overaccumulating chromoplasts in leaves agroinfiltrated or infected with constructs encoding the bacterial phytoene synthase crtB, leaving other non-engineered leaves of the plant to sustain normal growth. A combination of these two strategies, referred to as strategy C (for cytosolic production) and strategy P (for plastid conversion mediated by crtB), resulted in a 5-fold increase in the amount of β-carotene in Nicotiana benthamiana leaves. Following several attempts to further improve β-carotene leaf contents by metabolic engineering, hormone treatments and genetic screenings, it was found that promoting the proliferation of plastoglobules with increased light-intensity treatments not only improved β-carotene accumulation but it also resulted in a much higher bioaccessibility. The combination of strategies C and P together with a more intense light treatment increased the levels of accessible β-carotene 30-fold compared to controls. We further demonstrated that stimulating plastoglobule proliferation with strategy P, but also with a higher-light treatment alone, also improved β-carotene contents and bioaccessibility in edible lettuce (Lactuca sativa) leaves.

Genomics and transcriptomics reveal β-carotene synthesis mechanism in Dunaliella salina.

Dunaliella salina is by far the most salt-tolerant organism and contains many active substances, including β-carotene, glycerol, proteins, and vitamins, using in the production of dried biomass or cell extracts for the biofuels, pharmaceutical formulations, food additives, and fine chemicals, especially β-carotene. We report a high-quality genome sequence of D. Salina FACHB435, which has a 472 Mb genome size, with a contig N50 of 458 Kb. A total of 30,752 protein-coding genes were predicted. The annotation results evaluated by BUSCO was shown that completeness was 91.0% and replication was 53.1%. The fragments were 6.3% and the deletions were 2.6%. Phylogenomic and comparative genomic analyses revealed that A. thaliana diverged from Volvocales about 448 million years ago, then Volvocales C. eustigma, D. salina, and other species diverged about 250 million years ago. High light could promote the accumulation of β-carotene in D. salina at a 13 d stage of culture. The enrichment of DEGs in KEGG, it notes that the predicted up-regulated genes of carotenoid metabolic pathway include DsCrtB, DsPDS, DsZ-ISO, DsZDS, DsCRTISO, DsLUT5, DsCrtL-B, and DsCCD8, while the predicted down-regulated genes include DsCrtF, and DsLUT1. The four genes that were both up-regulated and down-regulated were DsZEP, DsCrtR-b, DsCruA/P and DsCrtZ 4. The research results can provide scientific basis for the industrialization practice of D. salina.

Inhibitory effects of chlorophyll pigments on the bioaccessibility of β-carotene: Influence of chlorophyll structure and oil matrix

Chlorophylls and β-carotene are fat-soluble phytochemicals in daily diets, while their bioaccessibility interaction remains unknown. Eight dietary chlorophylls and their derivatives (chlorophyll a, chlorophyll b, pheophytin a, pheophytin b, chlorophyllide a, chlorophyllide b, pheophorbide a, pheophorbide b) were combined with β-carotene in six different oil matrices (corn oil, coconut oil, medium-chain triglycerides, peanut oil, olive oil and fish oil) and were subjected to in vitro digestion. Generally, chlorophylls significantly decreased β-carotene bioaccessibility by competitive incorporation into micelles. Dephytylated chlorophylls had a greater inhibitory effect on the micellarization and bioaccessibility of β-carotene compared to phytylated chlorophylls. In their co-digestion system, olive oil group exhibited the smallest particle size and biggest zeta potential in both digesta and micelles. For chlorophylls, the phytol group and their levels are key factors, which was also buttressed by the mice model where additional supplementation of pheophorbide a significantly hindered the accumulation of β-carotene and retinoids compounds.

High β-carotene accumulation in transgenic eggplant fruits grown under artificial light.

Eggplant (Solanum melongena L.) fruits are known to contain few carotenoids such as β-carotene, which are abundant in congener tomato fruits. In a previous study, we introduced a fruit-specific EEF48 gene promoter-driven crtB gene encoding phytoene synthase (PSY) of Erwinia uredovora into eggplant 'Senryo No. 2'. The transgenic plants grown in a greenhouse set fruits that accumulated β-carotene (∼1.67 µg g-1FW) in the T0 and T1 generations. In the present study, we grew T1 and T2 generations of the transgenic eggplant plants in artificial climate chambers to investigate their fruit set and β-carotene accumulation. No clear difference in β-carotene accumulation was observed in the fruit of transgenic plants grown under either HID (high-intensity discharge) or LED (light-emitting diode) light, or between T1 and T2 generations. The β-carotene accumulation (8.83 µg g-1FW on average) was approximately 5 times higher than the previous results obtained from greenhouse-grown plants. However, the fruit weight and size of the T-DNA (+) plants were significantly smaller than that of their null-segregant T-DNA (-) plants derived from the same line, suggesting that β-carotene accumulation may inhibit fruit development. Considering that a part of plants grown under LED irradiation failed to set fruits or set smaller fruits than those grown under HID irradiation, the light condition in the LED chamber may not be sufficient to promote fruit development. The present results are expected to provide valuable information for the selection of transgenic eggplants with high β-carotene content in fruit under artificial lighting.

Multi-omics provide insights into the regulation of DNA methylation in pear fruit metabolism

BackgroundExtensive research has been conducted on fruit development in crops, but the metabolic regulatory networks underlying perennial fruit trees remain poorly understood. To address this knowledge gap, we conduct a comprehensive analysis of the metabolome, proteome, transcriptome, DNA methylome, and small RNAome profiles of pear fruit flesh at 11 developing stages, spanning from fruitlet to ripening. Here, we systematically investigate the metabolic landscape and regulatory network involved.ResultsWe generate an association database consisting of 439 metabolites and 14,399 genes to elucidate the gene regulatory network of pear flesh metabolism. Interestingly, we detect increased DNA methylation in the promoters of most genes within the database during pear flesh development. Application of a DNA methylation inhibitor to the developing fruit represses chlorophyll degradation in the pericarp and promotes xanthophyll, β-carotene, and abscisic acid (ABA) accumulation in the flesh. We find the gradual increase in ABA production during pear flesh development is correlated with the expression of several carotenoid pathway genes and multiple transcription factors. Of these transcription factors, the zinc finger protein PbZFP1 is identified as a positive mediator of ABA biosynthesis in pear flesh. Most ABA pathway genes and transcription factors are modified by DNA methylation in the promoters, although some are induced by the DNA methylation inhibitor. These results suggest that DNA methylation inhibits ABA accumulation, which may delay fruit ripening.ConclusionOur findings provide insights into epigenetic regulation of metabolic regulatory networks during pear flesh development, particularly with regard to DNA methylation.

Development of a Gene-Based Marker Set for Orange-Colored Watermelon Flesh with a High β-Carotene Content.

The fruit flesh of watermelons differs depending on the distinct carotenoid composition. Orange-colored flesh relates to the accumulation of β-carotene, which is beneficial to human health. Canary-yellow-fleshed OTO-DAH and orange-β-fleshed (orange-fleshed with high β-carotene) NB-DAH near-isogenic lines (NILs) were used to determine the genetic mechanism attributed to orange watermelon flesh. For genetic mapping, an F2 population was developed by crossing the two NILs. The segregation ratio of flesh color in the F2 population indicated that the orange-β flesh of the NB-DAH NIL was controlled by a single incompletely dominant gene. Through a comparative analysis of the whole-genome sequences of the parent lines and NILs, a major introgression region unique to the NB-DAH NIL was detected on Chr. 1; this was considered a candidate region for harboring genes that distinguish orange from canary-yellow and red flesh. Among the 13 genes involved in the carotenoid metabolic pathway in watermelons, only ClPSY1 (ClCG01G008470), which encodes phytoene synthase 1, was located within the introgression region. The genotyping of F2 plants using a cleaved amplified polymorphic sequence marker developed from a non-synonymous SNP in ClPSY1 revealed its relationship with orange-β flesh. The insights gained in this study can be applied to marker-assisted breeding for this desirable trait.

Reducing PHYTOENE SYNTHASE activity fine-tunes the abundance of a cis-carotene-derived signal that regulates the PIF3/HY5 module and plastid biogenesis.

PHYTOENE SYNTHASE (PSY) is a rate-limiting enzyme catalysing the first committed step of carotenoid biosynthesis, and changes in PSY gene expression and/or protein activity alter carotenoid composition and plastid differentiation in plants. Four genetic variants of PSY (psy-4, psy-90, psy-130, and psy-145) were identified using a forward genetics approach that rescued leaf virescence phenotypes and plastid abnormalities displayed by the Arabidopsis CAROTENOID ISOMERASE (CRTISO) mutant ccr2 (carotenoid and chloroplast regulation 2) when grown under a shorter photoperiod. The four non-lethal mutations affected alternative splicing, enzyme-substrate interactions, and PSY:ORANGE multi-enzyme complex binding, constituting the dynamic post-transcriptional fine-tuning of PSY levels and activity without changing localization to the stroma and protothylakoid membranes. psy genetic variants did not alter total xanthophyll or β-carotene accumulation in ccr2, yet they reduced specific acyclic linear cis-carotenes linked to the biosynthesis of a currently unidentified apocarotenoid signal regulating plastid biogenesis, chlorophyll biosynthesis, and photomorphogenic regulation. ccr2 psy variants modulated the PHYTOCHROME-INTERACTING FACTOR 3/ELONGATED HYPOCOTYL 5 (PIF3/HY5) ratio, and displayed a normal prolamellar body formation in etioplasts and chlorophyll accumulation during seedling photomorphogenesis. Thus, suppressing PSY activity and impairing PSY:ORANGE protein interactions revealed how cis-carotene abundance can be fine-tuned through holoenzyme-metabolon interactions to control plastid development.

Population genomics identifies genetic signatures of carrot domestication and improvement and uncovers the origin of high-carotenoid orange carrots.

Here an improved carrot reference genome and resequencing of 630 carrot accessions were used to investigate carrot domestication and improvement. The study demonstrated that carrot was domesticated during the Early Middle Ages in the region spanning western Asia to central Asia, and orange carrot was selected during the Renaissance period, probably in western Europe. A progressive reduction of genetic diversity accompanied this process. Genes controlling circadian clock/flowering and carotenoid accumulation were under selection during domestication and improvement. Three recessive genes, at the REC, Or and Y2 quantitative trait loci, were essential to select for the high α- and β-carotene orange phenotype. All three genes control high α- and β-carotene accumulation through molecular mechanisms that regulate the interactions between the carotenoid biosynthetic pathway, the photosynthetic system and chloroplast biogenesis. Overall, this study elucidated carrot domestication and breeding history and carotenoid genetics at a molecular level.

Discovery of candidate genes related to carotenoid accumulation based on yellow sarcocarp bud mutation peach resource

The carotenoid composition and content in the yellow sarcocarp peach are a direct indicator of quality. Therefore, discoveries regarding the identification of candidate genes related to carotenoid accumulation and their regulatory mechanisms would be beneficial for yellow sarcocarp peach breeding. In this study, the white sarcocarp peach cultivar ‘21shiji’ (21SJ) and its unique yellow sarcocarp bud mutation individual (21BM) were used for carotenoid determination and candidate gene discovery based on UPLC-APCI-MS/MS and RNA-seq. In total, 11 carotenoid compositions showed significant differences between ‘21SJ’ and ‘21BM’, including four carotenes and seven xanthophylls. Furthermore, the accumulation of β-carotene, zeaxanthin, β-cryptoxanthin, antheraxanthin, violaxanthin, and capsanthin showed different tendencies between ‘21SJ’ and ‘21BM’ during the ripening stage and influenced ‘21BM’ yellow sarcocarp formation. Using transcriptome analysis, we found that the differential expression of candidate genes PpZEP, PpVDE1, PpCCD4, and PpCCS were major influencing factors. Two transfactors, PpbHLH14 and PpMYB73, were significantly related to PpZEP, PpVDE1, PpCCD4, and PpCCS expression and carotenoid accumulation after weighted gene co-expression network analysis. The identified structural genes and transcription factors in this study will further enrich carotenoid accumulation regulatory mechanism in peach and will provide an important theoretical reference for yellow sarcocarp peach breeding.

Concomitant Production of Erythritol and β-Carotene by Engineered Yarrowia lipolytica.

While the expansion of the erythritol production industry has resulted in unprecedented production of yeast cells, it also suffers from a lack of effective utilization. β-Carotene is a value-added compound that can be synthesized by engineered Yarrowia lipolytica. Here, we first evaluated the production performance of erythritol-producing yeast strains under two different morphologies and then successfully constructed a chassis with yeast-like morphology by deleting Mhy1 and Cla4 genes. Subsequently, β-carotene synthesis pathway genes, CarRA and CarB from Blakeslea trispora, were introduced to construct the β-carotene and erythritol coproducing Y. lipolytica strain ylmcc. The rate-limiting genes GGS1 and tHMG1 were overexpressed to increase the β-carotene yield by 45.32-fold compared with the strain ylmcc. However, increased β-carotene accumulation led to prolonged fermentation time; therefore, transporter engineering through overexpression of YTH1 and YTH3 genes was used to alleviate fermentation delays. Using batch fermentation in a 3 L bioreactor, this engineered Y. lipolytica strain produced erythritol with production, yield, and productivity values of 171 g/L, 0.56 g/g glucose, and 2.38 g/(L·h), respectively, with a concomitant β-carotene yield of 47.36 ± 0.06 mg/g DCW. The approach presented here improves the value of erythritol-producing cells and offers a low-cost technique to obtain hydrophobic terpenoids.

Overexpression of S-R enhances the accumulation of biomass, fatty acids, and β-carotene in Schizochytrium

Strategies for enhancing biomass accumulation and increasing the production of fatty acids and β-carotene in Schizochytrium are hindered by the lack of suitable targets. In this study, S-R, a RING (really interesting new gene) finger domain-containing protein, was identified in Schizochytrium, with homologs found in the family Thraustochytriaceae. Transgenic strains overexpressing S-R showed a minor improvement in cell growth but a significant increase in total fatty acids content by 1.29- to 1.36-fold. Almost all individual saturated fatty acids exhibited significant increases, with the greatest increase observed in the C14:0 content, by 1.52- to 1.78-fold. Additionally, the β-carotene content of S-R strains was significantly upregulated. Overexpression of s-r conferred hypersaline tolerance in Schizochytrium, with a significant increase in dry cell weight, total fatty acids and β-carotene, likely due to the upregulation of glycerol and proline. This study provides a feasible strategy to engineer Thraustochytriaceae for efficient biomass and biochemical production.

Metabolic engineering of Escherichia coli for high-level production of violaxanthin

BackgroundXanthophylls are a large class of carotenoids that are found in a variety of organisms and play particularly important roles in the light-harvesting and photoprotection processes of plants and algae. Violaxanthin is an important plant-derived xanthophyll with wide potential applications in medicines, foods, and cosmetics because of its antioxidant activity and bright yellow color. To date, however, violaxanthins have not been produced using metabolically engineered microbes on a commercial scale. Metabolic engineering for microbial production of violaxanthin is hindered by inefficient synthesis pathway in the heterologous host. We systematically optimized the carotenoid chassis and improved the functional expression of key enzymes of violaxanthin biosynthesis in Escherichia coli.ResultsCo-overexpression of crtY (encoding lycopene β-cyclase), crtZ (encoding β-carotene 3-hydroxylase), and ZEP (encoding zeaxanthin epoxidase) had a notable impact on their functions, resulting in the accumulation of intermediate products, specifically lycopene and β-carotene. A chassis strain that did not accumulate the intermediate was optimized by several approaches. A promoter library was used to optimize the expression of crtY and crtZ. The resulting strain DZ12 produced zeaxanthin without intermediates. The expression of ZEP was further systematically optimized by using DZ12 as the chassis host. By using a low copy number plasmid and a modified dithiol/disulfide system, and by co-expressing a full electron transport chain, we generated a strain producing violaxanthin at about 25.28 ± 3.94 mg/g dry cell weight with decreased byproduct accumulation.ConclusionWe developed an efficient metabolically engineered Escherichia coli strain capable of producing a large amount of violaxanthin. This is the first report of a metabolically engineered microbial platform that could be used for the commercial production of violaxanthin.

Overexpression of potato ORANGE (StOR) and StOR mutant in Arabidopsis confers increased carotenoid accumulation and tolerance to abiotic stress

ORANGE (OR) plays essential roles in regulating carotenoid homeostasis and enhancing the ability of plants to adapt to environmental stress. However, OR proteins have been functionally characterized in only a few plant species, and little is known about the role of potato OR (StOR). In this study, we characterized the StOR gene in potato (Solanum tuberosum L. cv. Atlantic). StOR is predominantly localized to the chloroplast, and its transcripts are tissue-specifically expressed and significantly induced in response to abiotic stress. Compared with wild type, overexpression of StOR increased β-carotene levels up to 4.8-fold, whereas overexpression of StORHis with a conserved arginine to histidine substitution promoted β-carotene accumulation up to 17.6-fold in Arabidopsis thaliana calli. Neither StOR nor StORHis overexpression dramatically affected the transcript levels of carotenoid biosynthetic genes. Furthermore, overexpression of either StOR or StORHis increased abiotic stress tolerance in Arabidopsis, which was associated with higher photosynthetic capacity and antioxidative activity. Taken together, these results indicate that StOR could be exploited as a potential new genetic tool for the improvement of crop nutritional quality and environmental stress tolerance.

Carotenoid sequestration protein FIBRILLIN participates in CmOR-regulated β-carotene accumulation in melon.

Chromoplasts are plant organelles with a unique ability to sequester and store massive carotenoids. Chromoplasts have been hypothesized to enable high levels of carotenoid accumulation due to enhanced sequestration ability or sequestration substructure formation. However, the regulators that control the substructure component accumulation and substructure formation in chromoplasts remain unknown. In melon (Cucumis melo) fruit, β-carotene accumulation in chromoplasts is governed by ORANGE (OR), a key regulator for carotenoid accumulation in chromoplasts. By using comparative proteomic analysis of a high β-carotene melon variety and its isogenic line low-β mutant that is defective in CmOr with impaired chromoplast formation, we identified carotenoid sequestration protein FIBRILLIN1 (CmFBN1) as differentially expressed. CmFBN1 expresses highly in melon fruit tissue. Overexpression of CmFBN1 in transgenic Arabidopsis (Arabidopsis thaliana) containing ORHis that genetically mimics CmOr significantly enhances carotenoid accumulation, demonstrating its involvement in CmOR-induced carotenoid accumulation. Both in vitro and in vivo evidence showed that CmOR physically interacts with CmFBN1. Such an interaction occurs in plastoglobules and results in promoting CmFBN1 accumulation. CmOR greatly stabilizes CmFBN1, which stimulates plastoglobule proliferation and subsequently carotenoid accumulation in chromoplasts. Our findings show that CmOR directly regulates CmFBN1 protein levels and suggest a fundamental role of CmFBN1 in facilitating plastoglobule proliferation for carotenoid sequestration. This study also reveals an important genetic tool to further enhance OR-induced carotenoid accumulation in chromoplasts in crops.

Transcriptome and metabolome profiling identify factors potentially involved in pro-vitamin A accumulation in cassava landraces

Cassava (Manihot esculenta Crantz) is a predominant food security crop in several developing countries. Its storage roots, rich in carbohydrate, are deficient in essential micronutrients, including provitamin A carotenoids.Increasing carotenoid content in cassava storage roots is important to reduce the incidence of vitamin A deficiency, a public health problem in sub-Saharan Africa. However, cassava improvement advances slowly, mainly due to limited information on the molecular factors influencing β-carotene accumulation in cassava.To address this problem, we performed comparative transcriptomic and untargeted metabolic analyses of roots and leaves of eleven African cassava landraces ranging from white to deep yellow colour, to uncover regulators of carotenoid biosynthesis and accumulation with conserved function in yellow cassava roots.Sequence analysis confirmed the presence of a mutation, known to influence β-carotene content, in PSY transcripts of deep yellow but not of pale yellow genotypes. We identified genes and metabolites with expression and accumulation levels significantly associated with β-carotene content. Particularly an increased activity of the abscisic acid catabolism pathway together with a reduced amount of L-carnitine, may be related to the carotenoid pathway flux, higher in yellow than in white storage roots. In fact, NCED_3.1 was specifically expressed at a lower level in all yellow genotypes suggesting that it could be a potential target for increasing carotenoid accumulation in cassava.These results expand the knowledge on metabolite compositions and molecular mechanisms influencing carotenoid biosynthesis and accumulation in cassava and provide novel information for biotechnological applications and genetic improvement of cassava with high nutritional values.

Effects of Light on the Growth and β-carotene Accumulation in the Green Algae Dunaliella salina

The marine microalgae Dunaliella salina holds significant economic value due to its rich content of β-carotene, a natural pigment with high antioxidant capacity, immune-stimulating properties, and a crucial role in antioxidant reactions with vitamins C and E. This study evaluated the effects of light factors on the growth and biosynthesis of β-carotene in the Dunaliella salina strain isolated from coastal waters in Central Vietnam. The microalgae strain was cultured in the static f/2 medium at 25°C, under different light spectra (blue, red, and white) and light intensities (13.5, 27.0, and 40.5 µmol photon/m2/s), with a similar light/dark cycle of 16/8. The results indicated that the microalgae strain showed the highest growth rate and production yield of β-carotene under the red LED light, whereas the highest β-carotene accumulation in each microalgal cell was obtained under the blue LED light. Regarding light intensity, the best growth and β-carotene production yield was observed under the highest experimental light intensity of 40.5 µmol photon/m2/s, but the highest β-carotene content per algal cell was reached under the lowest intensity (13.5 µmol photon/m2/s). These findings provide important scientific data for further research to identify the optimal conditions for increasing biomass and stimulating the accumulation of valuable secondary compounds of the microalgae Dunaliella salina in Vietnam.

Roles of Two Phytoene Synthases and Orange Protein in Carotenoid Metabolism of the β-Carotene-Accumulating Dunaliella salina.

Phytoene synthase (PSY) is a key enzyme in carotenoid metabolism and often regulated by orange protein. However, few studies have focused on the functional differentiation of the two PSYs and their regulation by protein interaction in the β-carotene-accumulating Dunaliella salina CCAP 19/18. In this study, we confirmed that DsPSY1 from D. salina possessed high PSY catalytic activity, whereas DsPSY2 almost had no activity. Two amino acid residues at positions 144 and 285 responsible for substrate binding were associated with the functional variance between DsPSY1 and DsPSY2. Moreover, orange protein from D. salina (DsOR) could interact with DsPSY1/2. DbPSY from Dunaliella sp. FACHB-847 also had high PSY activity, but DbOR could not interact with DbPSY, which might be one reason why it could not highly accumulate β-carotene. Overexpression of DsOR, especially the mutant DsORHis, could significantly improve the single-cell carotenoid content and change cell morphology (with larger cell size, bigger plastoglobuli, and fragmented starch granules) of D. salina. Overall, DsPSY1 played a dominant role in carotenoid biosynthesis in D. salina, and DsOR promoted carotenoid accumulation, especially β-carotene via interacting with DsPSY1/2 and regulating the plastid development. Our study provides a new clue for the regulatory mechanism of carotenoid metabolism in Dunaliella. IMPORTANCE Phytoene synthase (PSY) as the key rate-limiting enzyme in carotenoid metabolism can be regulated by various regulators and factors. We found that DsPSY1 played a dominant role in carotenogenesis in the β-carotene-accumulating Dunaliella salina, and two amino acid residues critical in the substrate binding were associated with the functional variance between DsPSY1 and DsPSY2. Orange protein from D. salina (DsOR) can promote carotenoid accumulation via interacting with DsPSY1/2 and regulating the plastid development, which provides new insights into the molecular mechanism of massive accumulation of β-carotene in D. salina.