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The circadian dynamics of the expression of key genes of carotenoid metabolism (PSY2, LCYE, CrtRB1, and NCED1) in the photosynthetic tissue of tomato Solanum lycopersicum L. (cultivar Korneevsky) plants was characterized. An in silico analysis of the gene expression pattern was carried out and a high level of their transcripts was detected in the leaf tissue. qRT-PCR analysis of gene expression was performed at six time points during the day and showed the highest levels of PSY2, LCYE, and NCED1 transcripts in the second half of the light phase and CrtRB1 at the end of the dark phase. The content and composition of carotenoids in leaf tissue in the middle of the day was determined; it was shown that the leaf accumulates 1.5 times more compounds of the ɛ/β-branch of carotenoid biosynthesis pathway than compounds of the β/β-branch. [ABSTRACT FROM AUTHOR]

The NAC transcription factors play an important regulatory role in plant growth and development. In order to explore the impact of NAC6 on carotenoid metabolism in postharvest mango fruit, this study investigated changes in the carotenoid contents as well as the expression of NAC6 and key enzymes involved in carotenoid synthesis and metabolism in the mature peel and pulp of 2 mango varieties, “Tainong” and “Jinhuang”, at commercial maturity, during postharvest storage under constant temperature (20 ℃), constant humidity (85%), and dark conditions. The results showed that both mango varieties contained α-carotene, lutein, β-carotene, β-cryptoxanthin, and zeaxanthin. Throughout the storage process, the expression of NAC6 in both mango varieties showed a continuous upward trend, reaching a peak in the late stage of storage, followed by a slight decline. Correlation analysis showed that NAC6 might induce the gene expression of 1-deoxy-D-xylulose-5-phosphate reductoisomerase, farnesyl pyrophosphate synthase, geranylgeranyl pyrophosphate synthase, phytoene synthase, phytoene desaturase, ζ-carotene desaturase, carotenoid isomerase, β-carotene hydroxylase, and violaxanthin de-epoxidase in mango fruit, while inhibiting the gene expression of lycopene β-cyclase, neoxanthin synthase, and 9-cis-epoxycarotenoid dioxygenase. The expression of NAC6 in the peel and pulp of “Tainong” mango was significantly negatively correlated with the gene expression of carotenoid cleavage dioxygenase (CCD), with correlation coefficients of −0.841 0 and −0.838 3, respectively. NAC6 might delay the degradation of carotenoids by inhibiting the expression of CCD. The results from this research provide an experimental basis for the postharvest storage and preservation of mango fruit as well as its quality control.

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Carotenoids play crucial physiological roles in animals. A comprehensive investigation into the mechanism of carotenoid metabolism in oysters will establish a theoretical foundation for further development of its carotenoid-rich traits. However, the information on the function of miRNA in β-carotene metabolism in oysters is limited. To elucidate the mechanisms underlying miRNA regulation of carotenoid metabolism in oysters, we compared the expressions of miRNA in digestive gland tissues of Pacific oyster (Crassostrea gigas) fed with a β-carotene supplemented diet and a normal diet, respectively. A total of 690 candidate miRNAs in the Pacific oyster digestive gland tissues were identified, including 590 known miRNAs and 111 unknown miRNAs. Three differentially expressed miRNAs were obtained in the carotenoid-fed and normal groups, associated to 137 differentially expressed target genes. Moreover, the GO enrichment analysis revealed that the differentially expressed target genes were mainly involved in transmembrane transport activity. KEGG enrichment showed that the differentially expressed target genes were involved in ABC transport. Analysis of the mRNA-miRNA network revealed that novel0025 played a central role in carotenoid metabolism, and it was negatively correlated with the expression of 46 mRNAs. In addition, down-regulated expression of novel0025 upregulated the expression of the lipoprotein gene LOC105342186, suggesting a potential regulatory role in carotenoid metabolism. Our results provide useful information for elucidating the miRNA regulation mechanism during carotenoids metabolism in the Pacific oyster. [ABSTRACT FROM AUTHOR]

Carotenoid cleavage oxygenases can cleave carotenoids into a range of biologically important products. Carotenoid isomerooxygenase (NinaB) and β, β-carotene 15, 15′-monooxygenase (BCO1) are two important oxygenases. In order to understand the roles that both oxygenases exert in crustaceans, we first investigated NinaB-like (EsNinaBl) and BCO1-like (EsBCO1l) within the genome of Chinese mitten crab (Eriocheir sinensis). Their functions were then deciphered through an analysis of their expression patterns, an in vitro β-carotene degradation assay, and RNA interference. The results showed that both EsNinaBl and EsBCO1l contain an RPE65 domain and exhibit high levels of expression in the hepatopancreas. During the molting stage, EsNinaBl exhibited significant upregulation in stage C, whereas EsBCO1l showed significantly higher expression levels at stage AB. Moreover, dietary supplementation with β-carotene resulted in a notable increase in the expression of EsNinaBl and EsBCO1l in the hepatopancreas. Further functional assays showed that the EsNinaBl expressed in E. coli underwent significant changes in its color, from orange to light; in addition, its β-carotene cleavage was higher than that of EsBCO1l. After the knockdown of EsNinaBl or EsBCO1l in juvenile E. sinensis, the expression levels of both genes were significantly decreased in the hepatopancreas, accompanied by a notable increase in the redness (a*) values. Furthermore, a significant increase in the β-carotene content was observed in the hepatopancreas when EsNinaBl-mRNA was suppressed, which suggests that EsNinaBl plays an important role in carotenoid cleavage, specifically β-carotene. In conclusion, our findings suggest that EsNinaBl and EsBCO1l may exhibit functional co-expression and play a crucial role in carotenoid cleavage in crabs. [ABSTRACT FROM AUTHOR]

Flower development is one of the most vital pathways in plant development, during which the epigenetic regulation of gene expression is essential. DNA methylation, the most conserved epigenetic modification, participates in gene expression regulation and transposable element silencing. Honeysuckle (Lonicera japonica) is an important medicinal plant renowned for its colorful and fragrant flowers. Honeysuckle flowers change color from white to gold as a result of carotenoid accumulation during development. However, the role of DNA methylation in flower color changes is not well understood in L. japonica. Here, we performed whole-genome bisulfite sequencing and transcriptome sequencing during flowering development in honeysuckle. The results showed that a decrease in the levels of genome-wide average DNA methylation during flower development and changes in DNA methylation were associated with the expression of demethylase genes. Moreover, many genes involved in carotenoid biosynthesis and degradation, such as LjPSY1, LjPDS1, LjLCYE, and LjCCD4, have altered expression levels because of hypomethylation, indicating that DNA methylation plays an important role in flower color changes in honeysuckle. Taken together, our data provide epigenetic insights into flower development and color change in honeysuckles.

The evolution of teleost coloration is strongly linked with the fish-specific genome duplication (FSGD). Sepiapterin reductase (Spr) is commonly required in pteridine pigment synthesis and two paralogs are found in teleosts. However, the functions of spra and sprb in pigmentation have not been completely elucidated. Here we found that spra was intensively expressed in skin, fins and scales of koi carp, while sprb was ubiquitously expressed in all tissues. Then, the Spr enzyme activity inhibitor, sulfapyridine (SFD), was used to explore the phenotype and genes variation after spr inhibition. Following the administration of SFD, spra was significantly suppressed but sprb was up-regulated in skin. Additionally, the color phenotypes shifted from red to yellow due to a reduction in carotenoid concentration and the number of xanthophores/erythrophores in scales. Furthermore, the genes involved in carotenoid metabolism were influenced. However, pteridine metabolism-related genes rarely exhibited obvious changes. The results indicate that spra inhibition resulted in the dysregulation of carotenoid metabolism and pigment cells reduction, whereas sprb was likely to maintain the conserved functions in pteridine synthesis for metabolic homoeostasis. This study provided evidences for the different functions of spr paralogs in carotenoid and pteridine metabolism and for better understanding of the relationship between genome duplication and color pattern diversification.

MicroRNAs (miRNAs) are endogenous small non-coding RNAs that play important roles in several biological processes, including the regulation of body color. Leopard coral grouper (Plectropomus leopardus) is a valuable marine aquaculture fish; however, at present, there are no published reports on its early body color development. The skin color of P. leopardus undergoes a transition from transparent to red from 26 days post-hatching (dph) to 30 dph. In this study, we performed miRNA sequencing on 26 dph (Transparent, PT) and 30 dph (Red, PR) skin samples of P. leopardus to explore the molecular mechanism underlying red color formation. A total of 44.31 M and 37.55 M clean tags were obtained from PT and PR group, respectively. Among these tags, 981 miRNAs were identified, including 493 known and 488 novel miRNAs. A total of 106 differentially expressed miRNAs (DEMs) were identified in PT vs. PR, with 43 up-regulated and 63 down-regulated miRNAs in the PR group, compared to the PT group (|fold change| > 2 and p-value < 0.05). A miRNA-mRNA network based on 18 candidate miRNAs and 53 target genes related to pigmentation, and KEGG enrichment analysis of the target genes of all DEMs, revealed that miRNAs involved in the formation of red skin color were mainly related to: 1) the inhibition of melanin synthesis (miR-141-z, miR-206-z, miR-206-y, miR-27-z, miR-137-y, miR-204-x, miR-204-y, miR-211-x, miR-211-z); 2) chromatophore development (miR-206-z, miR-206-y, miR-499-y, miR-1-z, miR-2188-x, miR-423-x); and 3) carotenoid metabolism (miR-204-x, miR-204-y, miR-499-y). This study demonstrates the potential role of miRNAs in red color formation and lays the foundation for the molecular mechanism of body color polymorphism in P. leopardus.

SUMMARY: The accumulation of carotenoids, such as xanthophylls, lycopene, and carotenes, is responsible for the color of carrot (Daucus carota subsp. sativus) fleshy roots. The potential role of DcLCYE, encoding a lycopene ε‐cyclase associated with carrot root color, was investigated using cultivars with orange and red roots. The expression of DcLCYE in red carrot varieties was significantly lower than that in orange carrots at the mature stage. Furthermore, red carrots accumulated larger amounts of lycopene and lower levels of α‐carotene. Sequence comparison and prokaryotic expression analysis revealed that amino acid differences in red carrots did not affect the cyclization function of DcLCYE. Analysis of the catalytic activity of DcLCYE revealed that it mainly formed ε‐carotene, while a side activity on α‐carotene and γ‐carotene was also observed. Comparative analysis of the promoter region sequences indicated that differences in the promoter region may affect the transcription of DcLCYE. DcLCYE was overexpressed in the red carrot 'Benhongjinshi' under the control of the CaMV35S promoter. Lycopene in transgenic carrot roots was cyclized, resulting in the accumulation of higher levels of α‐carotene and xanthophylls, while the β‐carotene content was significantly decreased. The expression levels of other genes in the carotenoid pathway were simultaneously upregulated. Knockout of DcLCYE in the orange carrot 'Kurodagosun' by CRISPR/Cas9 technology resulted in a decrease in the α‐carotene and xanthophyll contents. The relative expression levels of DcPSY1, DcPSY2, and DcCHXE were sharply increased in DcLCYE knockout mutants. The results of this study provide insights into the function of DcLCYE in carrots, which could serve as a basis for creating colorful carrot germplasms. Significance Statement: In this study, we found that the accumulation of lycopene and carotenes in carrot (Daucus carota subsp. sativus) root was related to the expression of the lycopene ε‐cyclase gene DcLCYE. Catalytic activity analysis in Escherichia coli and gene function verification in transgenic carrot demonstrated that DcLCYE could alter the metabolic flow of carotenes. [ABSTRACT FROM AUTHOR]

Objective: Low plasma levels of carotenoids are associated with mortality and chronic disease states. Genetic studies in animals revealed that the tissue accumulation of these dietary pigments is associated with the genes encoding β-carotene oxygenase 2 (BCO2) and the scavenger receptor class B type 1 (SR-B1). Here we examined in mice how BCO2 and SR-B1 affect the metabolism of the model carotenoid zeaxanthin that serves as a macular pigment in the human retina. Methods: We used mice with a lacZ reporter gene knock-in to determine Bco2 expression patterns in the small intestine. By genetic dissection, we studied the contribution of BCO2 and SR-B1 to zeaxanthin uptake homeostasis and tissue accumulation under different supply conditions (50 mg/kg and 250 mg/kg). We determined the metabolic profiles of zeaxanthin and its metabolites in different tissues by LC-MS using standard and chiral columns. An albino Isx−/−/Bco2−/− mouse homozygous for Tyrc−2J was generated to study the effect of light on ocular zeaxanthin metabolites. Results: We demonstrate that BCO2 is highly expressed in enterocytes of the small intestine. Genetic deletion of Bco2 led to enhanced accumulation of zeaxanthin, indicating that the enzyme serves as a gatekeeper of zeaxanthin bioavailability. Relaxing the regulation of SR-B1 expression in enterocytes by genetic deletion of the transcription factor ISX further enhanced zeaxanthin accumulation in tissues. We observed that the absorption of zeaxanthin was dose-dependent and identified the jejunum as the major zeaxanthin-absorbing intestinal region. We further showed that zeaxanthin underwent oxidation to ε,ε-3,3′-carotene-dione in mouse tissues. We detected all three enantiomers of the zeaxanthin oxidation product whereas the parent zeaxanthin only existed as (3R, 3′R)-enantiomer in the diet. The ratio of oxidized to parent zeaxanthin varied between tissues and was dependent on the supplementation dose. We further showed in an albino Isx−/−/Bco2−/− mouse that supra-physiological supplementation doses (250 mg/kg) with zeaxanthin rapidly induced hypercarotenemia with a golden skin phenotype and that light stress increased the concentration of oxidized zeaxanthin in the eyes. Conclusions: We established the biochemical basis of zeaxanthin metabolism in mice and showed that tissue factors and abiotic stress affect the metabolism and homeostasis of this dietary lipid.

This article is a Commentary on Li et al. (2023), 240: 285–301. [ABSTRACT FROM AUTHOR]

Carotenoid biosynthesis is closely associated with abscisic acid (ABA) during the ripening process of non-climacteric fruits, but the regulatory mechanism that links ABA signaling to carotenoid metabolism remains largely unclear. Here, we identified two master regulators of ABA-mediated citrus fruit coloration, CsERF110 and CsERF53, which activate the expression of carotenoid metabolism genes (CsGGPPS, CsPSY, CsPDS, CsCRTISO, CsLCYB2, CsLCYE, CsHYD, CsZEP, and CsNCED2) to facilitate carotenoid accumulation. Further investigations showed that CsERF110 not only activates the expression of CsERF53 by binding to its promoter but also interacts with CsERF53 to form the transcriptional regulatory module CsERF110-CsERF53. We also discovered a positive feedback regulatory loop between the ABA signal and carotenoid metabolism regulated by the transcriptional regulatory module CsERF110-CsERF53. Our results reveal that the CsERF110-CsERF53 module responds to ABA signaling, thereby orchestrating citrus fruit coloration. Considering the importance of carotenoid content for citrus and many other carotenoid-rich crops, the revelation of molecular mechanisms that underlie ABA-mediated carotenoid biosynthesis in plants will facilitate the development of transgenic/gene-editing approaches, further contributing to improving the quality of citrus and other carotenoid-rich crops., (Copyright © 2024 Huazhong Agricultural University. Published by Elsevier Inc. All rights reserved.)

Abstract Background Chlorophyll (Chl) is a vital photosynthetic pigment involved in capturing light energy and energy conversion. In this study, the color conversion of inner-leaves from green to yellow in the new wucai (Brassica campestris L.) cultivar W7–2 was detected under low temperature. The W7–2 displayed a normal green leaf phenotype at the seedling stage, but the inner leaves gradually turned yellow when the temperature was decreased to 10 °C/2 °C (day/night), This study facilitates us to understand the physiological and molecular mechanisms underlying leaf color changes in response to low temperature. Results A comparative leaf transcriptome analysis of W7–2 under low temperature treatment was performed on three stages (before, during and after leaf color change) with leaves that did not change color under normal temperature at the same period as a control. A total of 67,826 differentially expressed genes (DEGs) were identified. Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway and Gene Ontology (GO) analysis revealed that the DEGs were mainly enriched in porphyrin and Chl metabolism, carotenoids metabolism, photosynthesis, and circadian rhythm. In the porphyrin and chlorophyll metabolic pathways, the expression of several genes was reduced [i.e. magnesium chelatase subunit H (CHLH)] under low temperature. Almost all genes [i.e. phytoene synthase (PSY)] in the carotenoids (Car) biosynthesis pathway were downregulated under low temperature. The genes associated with photosynthesis [i.e. photosystem II oxygen-evolving enhancer protein 1 (PsbO)] were also downregulated under LT. Our study also showed that elongated hypocotyl5 (HY5), which participates in circadian rhythm, and the metabolism of Chl and Car, is responsible for the regulation of leaf color change and cold tolerance in W7–2. Conclusions The color of inner-leaves was changed from green to yellow under low temperature in temperature-sensitive mutant W7–2. Physiological, biochemical and transcriptomic studies showed that HY5 transcription factor and the downstream genes such as CHLH and PSY, which regulate the accumulation of different pigments, are required for the modulation of leaf color change in wucai under low temperature.

Drought stress is one of the abiotic stresses that limits crop production and greatly affects crop yield. Enhancement of plant stress resistance by NaCl pretreatment has been reported, but the mechanism by which NaCl pretreatment activates cotton stress resistance remains unclear. In this study, upland cotton (Gossypium hirsutum cv H177) was used as the material to conducted the treatments with three replications: 0 Mm NaCl + 0% PEG6000 (Polyethylene glycol), 0 mM NaCl + 15% PEG6000, 50 mM NaCl + 15% PEG6000 to explore the molecular mechanism by which NaCl improves the drought tolerance of cotton. The results showed that pretreatment with 50 mM NaCl could alleviate the adverse effects of PEG on cotton seeds while promoting the elongation of root length. RNA-seq showed that NaCl specifically induced the expression of carotenoid-related genes. By silencing the upstream gene GHLUT2 of lutein synthesis, it was found that the chlorophyll of silenced plants decreased, and leaf wilting was more sensitive to drought. We found that NaCl enhanced the drought resistance of cotton by regulating genes related to the carotenoid and abscisic acid downstream synthesis pathways. This study provides a new reference for the study of drought resistance in cotton and a theoretical basis for the molecular breeding of cotton.

Plectropomus leopardus is a valuable marine aquaculture fish, prized by consumers for its high nutritional value and gorgeous appearance. The economic significance of this species is largely influenced by its body color, prompting researchers to become increasingly interested in understanding the mechanisms behind its color formation. However, previous studies have mainly focused on the molecular mechanisms of body color formation during the adult stage of this species. In this study, it was observed that the body color of P. leopardus undergoes four distinct stages during early morphogenesis: transparent (PT), red (PR), red with spots (PRS), and brown with spots (PBS). The L*a*b* values of skin at each stage differ significantly. The number and distribution of erythrophores and melanophores play a crucial role in determining the body colors of P. leopardus. Transcriptome analysis of the skin at these stages revealed multiple differentially expressed genes (DEGs) and signaling pathways related to pigmentation. These pigmentation-related DEGs can be categorized into carotenoid metabolism-related genes (e.g., scarb1, ldlr, plin2, apod, ttc39b , etc), melanin synthesis-related genes (e.g., tyr, tyrp1, dct, pmel, slc7a11 , etc), and genes related to pigment cell development (pnp, ednrb, csf1r1, sox10, bnc2). GO and KEGG enrichment analysis indicated that processes like melanin synthesis (e.g., wnt signal pathway, melanogenesis, tyrosine metabolism, etc.), lipid metabolism (e.g., cholesterol metabolism, linoleic acid metabolism, arachidonic metabolism, fatty acid biosynthesis, etc.), and pigment cell development (e.g., pigment cell differentiation, cell development, etc.) are likely involved in the body color formation in P. leopardus. Finally, we proposed a hypothesis regarding carotenoid metabolism in erythrophores and melanin synthesis in melanophores of P. leopardus skin, drawing from existing research reports and transcriptome data. Overall, this study provides a new perspective on the molecular regulatory mechanisms of body color formation in P. leopardus , laying a theoretical foundation for molecular-assisted breeding of body color traits in this species. • P. leopardus undergoes four distinct body color stages during early morphogenesis. • Genes and pathways related to carotenoid metabolism, melanin synthesis and pigment cell development were identified. • Proposed a hypothesis about carotenoid metabolism in erythrophores and melanogenesis in melanophores of P. leopardus skin. [ABSTRACT FROM AUTHOR]

Intense red colours in birds are often owing to ketocarotenoids (KCs). In many land birds, KCs are oxidized from dietary yellow precursors, presumably by the avian carotenoid ketolase CYP2J19, the regulation and constraints of which have important implications for condition-dependence and honest signalling of carotenoid colour displays. We investigated hepatic CYP2J19 gene expression in the seasonally and sexually dichromatic southern red bishop (Euplectes orix) in relation to season, sex, progression of the prenuptial moult, testis size, body condition, redness and circulating sex steroids. A coloration function of CYP2J19 is supported by a seasonal upregulation prior to and during the carotenoid-depositing stage of the male prenuptial moult. However, CYP2J19 expression was similarly high in females (which do not moult prenuptially), and remained high in males after moult, suggesting additional or alternative roles of hepatic CYP2J19 or its products, such as detoxification or antioxidant functions. In males, the CYP2J19 upregulation preceded and was unrelated to the rise in plasma testosterone, but was correlated with androstenedione, probably of adrenal origin and compatible with luteinizing hormone-induced and (in females) oestrogen-suppressed moult. Finally, contrary to ideas that carotenoid ketolation rate mediates honest signalling of male quality, CYP2J19 expression was not related to plumage redness or male body condition.

Low temperature combined with low light (LL) is an adverse factor seriously affecting pepper productivity and quality. However, little is known about the molecular mechanisms related to LL stress responses. Therefore, transcriptome and physiological changes in Hangjiao No. 2 (H2) and Xiangtela No. 2 (X2) pepper were studied under normal conditions and LL. We found 8,392 and 8,028 differentially expressed genes in H2 and X2, respectively, significantly associated with photosynthesis, photosynthesis antenna proteins, and carotenoids, were enriched in 27 and 40 gene ontology terms in H2 and X2, respectively, and 14 and 16 Kyoto Encyclopedia of Gene and Genomes pathways. The accuracy and reliability of the RNA-Seq results were confirmed by qRT-PCR. Furthermore, carotenoid-related genes ZDS, CA1, CA2, NCED, LOC107840293, and LOC107850059 functioned in response to LL. Additionally, LL significantly decreased photosynthesis capacity, photosynthetic pigment contents, as well as maximum quantum efficiency, and changed carotenoid-related compounds, revealing photosynthesis and carotenoids were involved in LL stress response. Our findings provide insight into LL stress-induced transcriptional expression patterns.

Carotenoids play vital roles in the coloration of plant tissues and organs, particularly fruits; however, the regulation of carotenoid metabolism in fruits during ripening is largely unknown. Here, we show that red light promotes fruit coloration by inducing accelerated degreening and carotenoid accumulation in kumquat fruits. Transcriptome profiling revealed that a NAC (NAM/ATAF/CUC2) family transcription factor, FcrNAC22, is specifically induced in red light-irradiated fruits. FcrNAC22 localizes to the nucleus, and its gene expression is up-regulated as fruits change color. Results from dual luciferase, yeast one-hybrid assays and electrophoretic mobility shift assays indicate that FcrNAC22 directly binds to, and activates the promoters of three genes encoding key enzymes in the carotenoid metabolic pathway. Moreover, FcrNAC22 overexpression in citrus and tomato fruits as well as in citrus callus enhances expression of most carotenoid biosynthetic genes, accelerates plastid conversion into chromoplasts, and promotes color change. Knock down of FcrNAC22 expression in transiently transformed citrus fruits attenuates fruit coloration induced by red light. Taken together, our results demonstrate that FcrNAC22 is an important transcription factor that mediates red light-induced fruit coloration via up-regulation of carotenoid metabolism. [ABSTRACT FROM AUTHOR]

Background: Chlorophyll (Chl) is a vital photosynthetic pigment involved in capturing light energy and energy conversion. In this study, the color conversion of inner-leaves from green to yellow in the new wucai (Brassica campestris L.) cultivar W7–2 was detected under low temperature. The W7–2 displayed a normal green leaf phenotype at the seedling stage, but the inner leaves gradually turned yellow when the temperature was decreased to 10 °C/2 °C (day/night), This study facilitates us to understand the physiological and molecular mechanisms underlying leaf color changes in response to low temperature. Results: A comparative leaf transcriptome analysis of W7–2 under low temperature treatment was performed on three stages (before, during and after leaf color change) with leaves that did not change color under normal temperature at the same period as a control. A total of 67,826 differentially expressed genes (DEGs) were identified. Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway and Gene Ontology (GO) analysis revealed that the DEGs were mainly enriched in porphyrin and Chl metabolism, carotenoids metabolism, photosynthesis, and circadian rhythm. In the porphyrin and chlorophyll metabolic pathways, the expression of several genes was reduced [i.e. magnesium chelatase subunit H (CHLH)] under low temperature. Almost all genes [i.e. phytoene synthase (PSY)] in the carotenoids (Car) biosynthesis pathway were downregulated under low temperature. The genes associated with photosynthesis [i.e. photosystem II oxygen-evolving enhancer protein 1 (PsbO)] were also downregulated under LT. Our study also showed that elongated hypocotyl5 (HY5), which participates in circadian rhythm, and the metabolism of Chl and Car, is responsible for the regulation of leaf color change and cold tolerance in W7–2. Conclusions: The color of inner-leaves was changed from green to yellow under low temperature in temperature-sensitive mutant W7–2. Physiological, biochemical and transcriptomic studies showed that HY5 transcription factor and the downstream genes such as CHLH and PSY, which regulate the accumulation of different pigments, are required for the modulation of leaf color change in wucai under low temperature. [ABSTRACT FROM AUTHOR]

Granulation is a serious physiological disorder in citrus fruit, which impairs the fruit quality and causes economic losses to the citrus industry. 'Harumi' ((Citrus unshiu × C. sinensis) × C. reticulate), a mandarin-type variety, is prone to granulation, especially after long-term storage. In the present study, to elucidate the molecular mechanism of granulation, physiochemical changes were investigated in the juice sacs of 'Harumi' during the granulation process. The results showed that large size fruit (L fruit, transverse diameter of 8.5 - 10.0 cm) was prone to granulation, while small size fruit (S fruit, transverse diameter of 5.0 - 6.5 cm) was not. In L fruit, lignin content increased rapidly, which was accompanied by the reduction of the content of sugars (fructose and glucose) and organic acid (citric acid) during the granulation process. Moreover, the occurrence of granulation inhibited carotenoid biosynthesis in the 'Harumi' fruit. In L fruit, the content of the major carotenoids, β-cryptoxanthin, all- trans -violaxanthin, and 9- cis -violaxanthin, were decreased during the granulation process, and as a result, total carotenoid content in L fruit was 75.8% lower than that in S fruit in the fifth month after harvest. The real-time PCR results showed that the expression of carotenoid biosynthetic genes (CitPSY , CitPDS , CitZDS , CitLCYb2 , and CitHYb) was down-regulated, while the expression of carotenoid catabolic genes (CitNCED2 and CitNCED3) was up-regulated in L fruit during the granulation process. This study is the first to investigate the carotenoid metabolism during the granulation process, which will provide new insights into exploring the causes of granulation in citrus fruit. [Display omitted] • The large fruit of 'Harumi' is prone to granulation. • Lignin content increased rapidly during the granulation process. • Sugars and organic acids were consumed during the granulation process. • Carotenoid biosynthesis was inhibited in the granulated fruit of 'Harumi'. [ABSTRACT FROM AUTHOR]

β-Apocarotenoids are eccentric cleavage products of carotenoids formed by chemical and enzymatic oxidations. They occur in foods containing carotenoids and thus might be directly absorbed from the diet. However, there is limited information about their intestinal absorption. The present research examined the kinetics of uptake and metabolism of β-apocarotenoids. Caco-2 cells were grown on 6-well plastic plates until a differentiated cell monolayer was achieved. β-Apocarotenoids were prepared in Tween 40 micelles, delivered to differentiated cells in serum-free medium, and incubated at 37°C for up to 8 h. There was rapid uptake of β-apo-8′-carotenal into cells, and β-apo-8′-carotenal was largely converted to β-apo-8′-carotenoic acid and a minor metabolite that we identified as 5,6-epoxy-β-apo-8′-carotenol. There was also rapid uptake of β-apo-10′-carotenal into cells, and β-apo-10′-carotenal was converted into a major metabolite identified as 5,6-epoxy-β-apo-10′-carotenol and a minor metabolite that is likely a dihydro-β-apo-10′-carotenol. Finally, there was rapid cellular uptake of β-apo-13-carotenone, and this compound was extensively degraded. These results suggest that dietary β-apocarotenals are extensively metabolized in intestinal cells via pathways similar to the metabolism of retinal. Thus, they are likely not absorbed directly from the diet.

Low temperature combined with low light (LL) is an adverse factor seriously affecting pepper productivity and quality. However, little is known about the molecular mechanisms related to LL stress responses. Therefore, transcriptome and physiological changes in Hangjiao No. 2 (H2) and Xiangtela No. 2 (X2) pepper were studied under normal conditions and LL. We found 8,392 and 8,028 differentially expressed genes in H2 and X2, respectively, significantly associated with photosynthesis, photosynthesis antenna proteins, and carotenoids, were enriched in 27 and 40 gene ontology terms in H2 and X2, respectively, and 14 and 16 Kyoto Encyclopedia of Gene and Genomes pathways. The accuracy and reliability of the RNA-Seq results were confirmed by qRT-PCR. Furthermore, carotenoid-related genes ZDS, CA1, CA2, NCED, LOC107840293, and LOC107850059 functioned in response to LL. Additionally, LL significantly decreased photosynthesis capacity, photosynthetic pigment contents, as well as maximum quantum efficiency, and changed carotenoid-related compounds, revealing photosynthesis and carotenoids were involved in LL stress response. Our findings provide insight into LL stress-induced transcriptional expression patterns. [ABSTRACT FROM AUTHOR]

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Due to the ongoing prevalence of vitamin A deficiency (VAD) in developing countries there has been a large effort towards increasing the carotenoid content of staple foods via biofortification. Common strategies used for carotenoid biofortification include altering flux through the biosynthesis pathway to direct synthesis to a specific product, generally β-carotene, or via increasing the expression of genes early in the carotenoid biosynthesis pathway. Recently, carotenoid biofortification strategies are turning towards increasing the retention of carotenoids in plant tissues either via altering sequestration within the cell or via downregulating enzymes known to cause degradation of carotenoids. To date, little attention has focused on increasing the stability of carotenoids, which may be a promising method of increasing carotenoid content in staple foods. Combining strategies targeting multiple metabolic levels including synthesis, storage, and turnover will be necessary to achieve optimal outcomes for biofortification projects. The plastidial proteome is a key to understanding the sequestration and storage of carotenoids. Attenuating the activity of carotenoid cleavage dioxygenases and lipoxygenases, enzymes that drive carotenoid turnover, can improve carotenoid retention. Increasing the vitamin E content of crops may have a similar effect. Carotenoids are often found esterified to fatty acids. This process increases carotenoid sequestration, accumulation, and stability and may be a novel target for future biofortification projects. [ABSTRACT FROM AUTHOR]

Magnesium (Mg) has been found to promote the color change citrus peel, but the underlying mechanism remains unclear. In this study, Mg application treatment significantly increased the Mg content in the peel of Satsuma mandarin fruit during the expansion stage (90–150 days after flowering). Mg fertilizer treatment was also found to significantly reduce chlorophyll (Chl) a content and increase β, β-xanthophyll (β-cryptoxanthin, zeaxanthin, neoxanthin), and abscisic acid (ABA) content in the flavedo during the key period of color change in Satsuma mandarin (165–195 days after flowering). Under Mg fertilizer treatment, Chl biosynthesis genes (CitGGDR , CitCHLH , CitCHLM , CitCHL27 , and CitPORA) was down-regulated and that of Chl degradation genes (CitNYC , CitPPH , CitPAO , and CitRCCR) was up-regulated, which resulted in a lower Chl content in the flavedo than the control. Moreover, Mg fertilizer treatment up-regulated carotenoid biosynthesis genes (CitPSY , CitPDS , CitZDS , CitLCYb1 , CitLCYb2 , CitZEP , CitNCED2 , and CitNCED3), leading to the accumulation of β, β-xanthophyll and ABA in the flavedo. Mg fertilizer treatment also promoted sugar (mainly fructose, glucose, and sucrose) accumulation in fruit flavedo, which showed significant negative correlations with Chl, α-carotene, lutein, and β-carotene, but positive correlations with β, β-xanthophyll compositions, and ABA in the flavedo. An analysis integrating transcriptome and metabolome data revealed that Mg may promote the color change of Satsuma mandarin fruit flavedo by regulating various metabolic pathways related to peel coloration, such as sugar accumulation, Chl degradation, carotenoid and ABA biosynthesis, and carbon fixation. The results can explain the regulatory pathways and networks of Mg-induced citrus peel coloration. [Display omitted] • Mg application promoted chlorophyll degradation in Satsuma mandarin flavedo. • Mg application increased β-cryptoxanthin, zeaxanthin, neoxanthin, and ABA contents in Satsuma mandarin flavedo. • Mg application increased sugar accumulation in Satsuma mandarin flavedo. • The impacts may be ascribed to the expression of genes associated with pigment-sugar metabolism. [ABSTRACT FROM AUTHOR]

The approach of providing an oxygenic photosynthetic organism with a cyclic electron transfer system, i.e., a far-red light-driven proton pump, is widely proposed to maximize photosynthetic efficiency via expanding the absorption spectrum of photosynthetically active radiation. As a first step in this approach, Gloeobacter rhodopsin was expressed in a PSI-deletion strain of Synechocystis sp. PCC6803. Functional expression of Gloeobacter rhodopsin, in contrast to Proteorhodopsin, did not stimulate the rate of photoheterotrophic growth of this Synechocystis strain, analyzed with growth rate measurements and competition experiments. Nevertheless, analysis of oxygen uptake and—production rates of the Gloeobacter rhodopsin-expressing strains, relative to the ΔPSI control strain, confirm that the proton-pumping Gloeobacter rhodopsin provides the cells with additional capacity to generate proton motive force. Significantly, expression of the Gloeobacter rhodopsin did modulate levels of pigment formation in the transgenic strain.

Carotenoids are essential for photosynthesis and photoprotection in photosynthetic organisms, which are widely used in food coloring, feed additives, nutraceuticals, cosmetics, and pharmaceuticals. Carotenoid biofortification in crop plants or algae has been considered as a sustainable strategy to improve human nutrition and health. However, the regulatory mechanisms of carotenoid accumulation are still not systematic and particularly scarce in algae. This article focuses on the regulatory mechanisms of carotenoid accumulation in plants and algae through regulatory factors (transcription factors and regulatory proteins), demonstrating the complexity of homeostasis regulation of carotenoids, mainly including transcriptional regulation as the primary mechanism, subsequent post-translational regulation, and cross-linking with other metabolic processes. Different organs of plants and different plant/algal species usually have specific regulatory mechanisms for the biosynthesis, storage, and degradation of carotenoids in response to the environmental and developmental signals. In plants and algae, regulators such as MYB, bHLH, MADS, bZIP, AP2/ERF, WRKY, and orange proteins can be involved in the regulation of carotenoid metabolism. And many more regulators, regulatory networks, and mechanisms need to be explored. Our paper will provide a basis for multitarget or multipathway engineering for carotenoid biofortification in plants and algae.

Carotenoid composition has been studied in mesophilic, nitrogen-fixing cyanobacterium Anabaena sp. PCC7120 grown photoautotrophically, under diazotrophic conditions at four different temperatures (15 °C, 23 °C, 30 °C and 37 °C). The relative accumulation of chlorophyll, carotenoids and proteins was the highest at temperature of 23 °C. At a suboptimal temperature (15 °C) β-carotene was the dominant carotenoid compound, whereas the increase in temperature caused ketocarotenoids (echinenone, canthaxanthin, keto-myxoxanthophyll) to accumulate. A significant increase in the accumulation of phytoene synthase (CrtB) transcript was observed at both extreme growth temperatures (15 °C and 37 °C). The relative amount of β-carotene ketolase (CrtW) transcript directly corresponded to the accumulation of its product (keto-myxoxanthophyll) with a maximum at 30 °C and a profound decrease at 37 °C, whereas the transcription level of β-carotene ketolase (CrtO) was significantly decreased only at a suboptimal temperature (15 °C). These results show that temperature affects the functioning of the carotenoid biosynthesis pathway in Anabaena cells under photoautotrophic growth. Specifically, the balance between β-carotene and ketocarotenoids is altered according to temperature conditions. The transcriptional regulation of genes encoding enzymes active both at the early (CrtB) and the final steps (CrtO, CrtW) of the carotenoid biosynthetic pathway may participate in the acclimation mechanism of cyanobacteria to low and high temperatures. [ABSTRACT FROM AUTHOR]

Background: In peach fruit, carotenoid accumulation in the mesocarp causes the difference between yellow and white genotypes. The latter are generally characterized by a peculiar and more intense aroma, because of higher release of volatiles deriving from dioxygenase‐catalysed breakdown of the tetraterpene skeleton. The rate of carotenoid oxidation was investigated in peach (Prunus persica L.) fruits harvested at various stages of development. Two couples of white and yellow‐fleshed isogenic varieties and an ancestral white‐fleshed genotype were analysed, which had previously shown to differ in Carotenoid Cleavage Dioxygenase 4 allelic composition resulting in various combinations of putatively active/inactive proteins. Results: Carotenoid bleaching activity was localized in the insoluble fraction of fruit flesh chromoplasts. Higher rates of trans‐β‐apo‐8′‐carotenal than β‐carotene bleaching suggest that the first cleavage reaction is the rate‐limiting step. Consistently, HPLC analysis did not show the appearance of coloured intermediates in reaction mixtures. High levels of substrate breakdown were found during the initial phases of fruit development in all genotypes examined, whereas significant differences were evident during the second exponential growth phase and ripening onset. Also, the ratio of carotene versus carotenale utilization varied significantly. Conclusion: Pattern comparison among activity levels measured in vitro on chromoplast enriched fractions suggests that cleavage enzyme(s) other than Carotenoid Cleavage Dioxygenase 4 play a significant role in carotenoid breakdown during fruit development and ripening. © 2018 Society of Chemical Industry [ABSTRACT FROM AUTHOR]

This study aims to further examine the effect of Magnesium (Mg) application on fruit quality and carotenoid metabolism in Satsuma mandarin pulp. For this, a field experiment was using 20-year-old Satsuma mandarin (C. unshiu Marc.) for two treatment; (1) CK treatment (without Mg), (2) Mg fertilizer treatment (200 g MgO plant−1). Compared with CK, Mg treatment substantially raised the Mg content in pulp at 90 to 150 DAF (the fruit expansion period), increasing by 15.69%–21.74%. Mg treatment also increased fruit TSS content by 15.84% and 9.88%, decreased fruit TA content in by 34.25% and 33.26% at 195 DAF and 210 DAF (the fruit ripening period). Moreover, at 120 to 195 DAF, Mg treatment significantly increased the levels of lutein, β-cryptoxanthin, zeaxanthin and violaxanthin in the pulp. This can be explained by the increased expression of important biosynthetic genes, including CitPSY , CitPDS , CitLCYb1 , CitLCYb2 , CitLCYe , CitHYb , and CitZEP , that played a role in altering the carotenoid composition. The findings of this research offer a novel approach for augmenting both the economic and nutritional worth of citrus fruits. • Mg increases the Mg content in Satsuma mandarin pulp at fruit expansion period. • Improving the flavor of satsuma mandarin fruit by applying Mg. • Mg increases lutein, β-cryptoxanthin, zeaxanthin and violaxanthin in pulp. • Mg accelerates carotenoid accumulation via carotenoid metabolism gene expression. [ABSTRACT FROM AUTHOR]

Osmanthus fragrans ‘Yinbi Shuanghui’ not only has a beautiful shape and fresh floral fragrance, but also rich leaf colors that change, making the tree useful for landscaping. In order to study the mechanisms of color formation in O. fragrans ‘Yinbi Shuanghui’ leaves, we analyzed the colored and green leaves at different developmental stages in terms of leaf pigment content, cell structure, and transcriptome data. We found that the chlorophyll content in the colored leaves was lower than that of green leaves throughout development. By analyzing the structure of chloroplasts, the colored leaves demonstrated more stromal lamellae and low numbers of granum thylakoid. However, there was a large number of plastoglobuli. Using transcriptome sequencing, we demonstrated that the expression of differentially expressed genes (DEGs) involved in chlorophyll degradation was upregulated, i.e., heme oxygennase-1 (HO1), pheophorbide a oxidase (PAO), and chlorophyllase-2 (CLH2), affecting the synthesis of chlorophyll in colored leaves. The stay-green gene (SGR) was upregulated in colored leaves. Genes involved in carotenoid synthesis, i.e., phytoene synthase 1 (PSY1) and 1-Deoxyxylulose-5-phosphate synthase (DXS), were downregulated in colored leaves, impeding the synthesis of carotenoids. In the later stage of leaf development, the downregulated expression of Golden2-Like (GLK) inhibited chloroplast development in colored leaves. Using weighted gene co-expression network analysis (WGCNA) to investigate the correlation between physiological indicators and DEGs, we chose the modules with the highest degree of relevance to chlorophyll degradation and carotenoid metabolism. A total of five genes (HSFA2, NFYC9, TCP20, WRKY3, and WRKY4) were identified as hub genes. These analyses provide new insights into color formation mechanisms in O. fragrans ‘Yinbi Shuanghui’ leaves at the transcriptional level.

The dried powder of Paracoccus carotinifaciens (PANAFERD-AX®) contains (3S,3'S)-astaxanthin as a major carotenoid. Administration of PANAFERD-AX® for pigmentation of the prawn Penaeus japonicus was examined. Total carotenoid contents in the carapace, muscle, and head of the prawn were dose-dependently increased depending on the amount of PANAFERD-AX® administered. Furthermore, not only the amounts of astaxanthins (astaxanthin diester, astaxanthin monoester, and free astaxanthin) but also the amounts of yellow xanthophylls, isoastaxanthin (1), 5,6-dihydropenaeusxanthin (2), penaeusxanthin (3), tetrahydroxypirardixanthin (4), and curstaxanthin (5), were dose-dependently increased with the administration of PANAFERD-AX®. 5,6-Dihydropenaeusxanthin (2) and penaeusxanthin (3) are new carotenoids isolated from the prawn P. japonicus. These structures were determined to be (3R,4S,5R,6R,6'S)- 5,6-dihydro-3,4,4'- trihydroxy-β,ε-caroten-3'-one (2) and (3R,4S,6'S)-3,4,4'-trihydroxy-β,ε-caroten-3'-one (3) by UV/vis, ESI TOF MS, 1H NMR, and CD spectral data. The metabolism of astaxanthin to these yellow xanthophylls in the prawn was discussed. [ABSTRACT FROM AUTHOR]

This study aimed to investigate the key coloured compounds (anthocyanins, carotenoids…etc) and the key proteins associated to their synthesis or degradation of peach fruit peel ( Prunus persica ‘Hujingmilu’) using iTRAQ techniques at different peach developmental stages and during the postharvest period. Peach fruits were collected 50 d, 90 d, 95 d, 100 d, and 105 day after flowering, as well as fruits being stored for 3 days after picking at 105 days after flowering. The key coloured compounds of ‘Hujingmilu ‘honey peach peel is composed of, among others, chlorophyll, anthocyanin, carotenoids, zeaxanthin, and lycopene. Peel color was regulated by chlorophyll degradation pathways, carotenoid metabolism pathways, and anthocyanin metabolism pathways, the iTRAQ technique identified 1848 unique proteins, of which 842 were related to biological processes, and 25 were related to color and 33 were related to energy transfer during fruit development and postharvest. During the development of peach fruit, light-harvesting complex chlorophyll a/b binding protein (CAB) and protochlorophyllide reductase(POR) are the key functional proteins for regulating chlorophyll metabolism, anthocyanidin 3-O-glucosyltransferase (3 G T) and UFGT are the key functional proteins for regulating anthocyanidin metabolism, PSY, zeta-carotene desaturase (ZCD) and carotenoid cleavage dioxygenase (CCD) are the key functional proteins for regulating carotenoid metabolism during the development and the postharvest period of peach fruit. Mg-protoporphyrin IX chelatase and pheophorbide a oxygenase (PAO) are the key functional proteins for regulating chlorophyll metabolism during postharvest. The key proteins were regulated a little earlier or accompany with the increase or decrease of peach peel coloured compounds. These results provide further understanding of the key coloured compounds of peach peel color, key proteins associated to coloured compounds, and reveal how the differential proteins take part in coloured compound metabolism pathway during fruit development and postharvest. [ABSTRACT FROM AUTHOR]

Abstract: Red carotenoid colours in birds are widely assumed to be sexually selected quality indicators, but this rests on a very incomplete understanding of genetic mechanisms and honesty‐mediating costs. Recent progress was made by the implication of the gene CYP2J19 as an avian carotenoid ketolase, catalysing the synthesis of red C4‐ketocarotenoids from yellow dietary precursors, and potentially a major mechanism behind red coloration in birds. Here, we investigate the role of CYP2J19 in the spectacular colour diversification of African weaverbirds (Ploceidae), represented by five genera and 16 species: eight red, seven yellow and one without carotenoid coloration. All species had a single copy of CYP2J19, unlike the duplication found in the zebra finch, with high expression in the retina, confirming its function in colouring red oil droplets. Expression was weak or undetected in skin and follicles of pigment‐depositing feather buds, as well as in beaks and tarsi, including those of the red‐billed quelea. In contrast, the hepatic (liver) expression of CYP2J19 was consistently higher (>14‐fold) in seven species with C4‐ketocarotenoid coloration than in species without (including one red species), an association strongly supported by a phylogenetic comparative analysis. The results suggest a critical role of the candidate ketolase, CYP2J19, in the evolution of red C4‐ketocarotenoid colour variation in ploceids. As ancestral state reconstruction suggests that ketocarotenoid coloration has evolved twice in this group (once in Euplectes and once in the Quelea/Foudia clade), we argue that while CYP2J19 has retained its ancestral role in the retina, it has likely been co‐opted for red coloration independently in the two lineages, via increased hepatic expression. [ABSTRACT FROM AUTHOR]

• The whole genome AP2/ERF transcription factor family of pummelo was analyzed by bioinformatics. • Different color parameters, CCI values, Chlorophyll a, chlorophyll b, carotenoid content were observed in fruit exocarp between 'Sanhong' with and without bagging. • The expression levels of several carotenoid metabolism structure genes in pummelo fruit exocarp were measured. • CitERF23, CitERF27 and CitERF32 were proposed as candidate genes related to carotenoid metabolism in pummelo fruit exocarp. Pummelo (Citrus maxima) belong to subfamily Aurantioideae of Rutaceae. In production, 'Sanhongmiyou' exocarp turned out red after bagging but remained yellow green without bagging. Carotenoids were the main color in pummelo exocarp. The AP2/ERF family has been confirmed to be associated with fruit coloring by regulating genes related to carotenoid metabolism in multiple species. To better understand the molecular mechanisms of exocarp coloration and carotenoid accumulation, a total of 126 AP2/ERF transcription factor family members distributed on 9 chromosomes were screened from pummelo genome database and divided into 3 subfamilies including 110 ERF subfamilies, 13 AP2 subfamilies and 3 RAV subfamilies. 26 pairs of genes from the AP2/ERF transcription factor family were colinear. 18 pairs of genes were evolutionarily conserved and functionally different after tandem repetition events. Multi-species collinearity analysis showed close evolutionary relationship between pummelo, Actinidia chinensis and Eriobotrya japonica. Cis-acting elements prediction in the promoter region revealed that members of AP2/ERF family are rich in cis-acting elements which respond to light and plant hormones. Color parameters L*, a*, b* and CCI values were measured. Chlorophyll a,chlorophyll b, carotenoid content in fruit exocarp were determined. Eight ERF genes related to the carotenoid metabolism were screened out and tested by qRT-PCR. The expression pattern of structure genes in carotenoid metabolism were also measured with qRT-PCR. The expression patterns of CitERF23, CitERF27 and CitERF32 appeared correlation with pigment content and color indexes. Meanwhile, CitERF23 showed significant positively correlation with ZDS and CDD4. CitERF27 showed significant positively correlation with CDD1. CitERF32 showed significant positively correlation with PDS. CitERF120 showed significant positively correlation with CDD1. CitERF23, CitERF27 and CitERF32 were proposed as a candidate gene regulating carotenoid metabolism in the AP2/ERF family. [ABSTRACT FROM AUTHOR]

Low plasma levels of carotenoids are associated with mortality and chronic disease states. Genetic studies in animals revealed that the tissue accumulation of these dietary pigments is associated with the genes encoding β-carotene oxygenase 2 (BCO2) and the scavenger receptor class B type 1 (SR-B1). Here we examined in mice how BCO2 and SR-B1 affect the metabolism of the model carotenoid zeaxanthin that serves as a macular pigment in the human retina. We used mice with a lacZ reporter gene knock-in to determine Bco2 expression patterns in the small intestine. By genetic dissection, we studied the contribution of BCO2 and SR-B1 to zeaxanthin uptake homeostasis and tissue accumulation under different supply conditions (50 mg/kg and 250 mg/kg). We determined the metabolic profiles of zeaxanthin and its metabolites in different tissues by LC-MS using standard and chiral columns. An albino Isx −/− /Bco2 −/− mouse homozygous for Tyrc−2J was generated to study the effect of light on ocular zeaxanthin metabolites. We demonstrate that BCO2 is highly expressed in enterocytes of the small intestine. Genetic deletion of Bco2 led to enhanced accumulation of zeaxanthin, indicating that the enzyme serves as a gatekeeper of zeaxanthin bioavailability. Relaxing the regulation of SR-B1 expression in enterocytes by genetic deletion of the transcription factor ISX further enhanced zeaxanthin accumulation in tissues. We observed that the absorption of zeaxanthin was dose-dependent and identified the jejunum as the major zeaxanthin-absorbing intestinal region. We further showed that zeaxanthin underwent oxidation to ε,ε-3,3′-carotene-dione in mouse tissues. We detected all three enantiomers of the zeaxanthin oxidation product whereas the parent zeaxanthin only existed as (3R, 3′R)-enantiomer in the diet. The ratio of oxidized to parent zeaxanthin varied between tissues and was dependent on the supplementation dose. We further showed in an albino Isx −/− / Bco2 −/− mouse that supra-physiological supplementation doses (250 mg/kg) with zeaxanthin rapidly induced hypercarotenemia with a golden skin phenotype and that light stress increased the concentration of oxidized zeaxanthin in the eyes. We established the biochemical basis of zeaxanthin metabolism in mice and showed that tissue factors and abiotic stress affect the metabolism and homeostasis of this dietary lipid. [Display omitted] • Intestinal BCO2 expression serves as a gatekeeper of carotenoid accumulation in mice. • Interaction between ISX and SR-B1 modulates carotenoid absorption and accumulation. • Abiotic stress and tissue factors affect carotenoid homeostasis. • Albino Isx −/− /Bco2 −/− mouse serves as a multifaceted model for carotenoid biology. [ABSTRACT FROM AUTHOR]

• Male C57BL/6J mice were fed a 0.2 % fucoxanthin-containing diet for one week. • Total lipids in the tissues were analyzed to identify carotenoid fatty acid esters. • Fucoxanthinol-3-fatty acid esters abundantly accumulated in the liver. • Epididymal white adipose tissue was rich in amarouciaxanthin A-3-fatty acid esters. • Fucoxanthinol and amaroucixanthin A can be acylated in fucoxanthin-fed mice tissues. Administered carotenoid fatty acid esters are thought to be hydrolyzed to their free forms and absorbed into the body, and information on the tissue distribution of carotenoid fatty acid esters has been limited. Fucoxanthin, a marine carotenoid, exhibits various health benefits, including anti-diabetic and anti-obesity effects. However, fucoxanthin metabolism in mammals remains unclear. Herein, we investigated the fatty acid esters of fucoxanthin metabolites, fucoxanthinol and amarouciaxanthin A, in the tissues of male C57BL/6J mice fed a fucoxanthin-containing diet for one week. Fucoxanthinol and amarouciaxanthin A-3-esters accumulated abundantly in the liver and epididymal white adipose tissue, respectively. These esters were less detectable in the serum and other tissues. Therefore, it is suggested that fucoxanthinol and amarouciaxanthin A are partially acylated in the liver and epididymal white adipose tissue after being transported through the body as their free forms. This study presents a novel carotenoid metabolic pathway in mammals. [ABSTRACT FROM AUTHOR]

Isotope dilution is currently the most accurate technique in humans to determine vitamin A status and bioavailability/bioconversion of provitamin A carotenoids such as β-carotene. However, limits of MS detection, coupled with extensive isolation procedures, have hindered investigations of physiologically-relevant doses of stable isotopes in large intervention trials. Here, a sensitive liquid chromatography-tandem mass spectrometry (LC/MS/MS) analytical method was developed to study the plasma response from coadministered oral doses of 2 mg [13C10]β-carotene and 1 mg [13C10]retinyl acetate in human subjects over a 2 week period. A reverse phase C18 column and binary mobile phase solvent system separated β-carotene, retinol, retinyl acetate, retinyl linoleate, retinyl palmitate/retinyl oleate, and retinyl stearate within a 7 min run time. Selected reaction monitoring of analytes was performed under atmospheric pressure chemical ionization in positive mode at m/z 537→321 and m/z 269→93 for respective [12C]β-carotene and [12C] retinoids; m/z 547→330 and m/z 274→98 for [13C10]β-carotene and [13C5] cleavage products; and m/z 279→100 for metabolites of [13C10]retinyl acetate. A single one-phase solvent extraction, with no saponification or purification steps, left retinyl esters intact for determination of intestinally-derived retinol in chylomicrons versus retinol from the liver bound to retinol binding protein. Coadministration of [13C10]retinyl acetate with [13C10]β-carotene not only acts as a reference dose for inter-individual variations in absorption and chylomicron clearance rates, but also allows for simultaneous determination of an individual's vitamin A status.

Although the functions of carotenogenic genes are well documented, little is known about the mechanisms that regulate their expression, especially those genes involved in α - and β-branch carotenoid metabolism., In this study, an R2R3-MYB transcriptional factor (CrMYB68) that directly regulates the transformation of α- and β-branch carotenoids was identified using Green Ougan (MT), a stay-green mutant of Citrus reticulata cv Suavissima. A comprehensive analysis of developing and harvested fruits indicated that reduced expression of β-carotene hydroxylases 2 ( CrBCH2) and 9-cis-epoxycarotenoid dioxygenase 5 ( CrNCED5) was responsible for the delay in the transformation of α- and β-carotene and the biosynthesis of ABA. Additionally, the expression of these genes was negatively correlated with the expression of CrMYB68 in MT., Further, electrophoretic mobility shift assays (EMSAs) and dual luciferase assays indicated that CrMYB68 can directly and negatively regulate CrBCH2 and CrNCED5. Moreover, transient overexpression experiments using leaves of Nicotiana benthamiana indicated that CrMYB68 can also negatively regulate NbBCH2 and NbNCED5., To overcome the difficulty of transgenic validation, we quantified the concentrations of carotenoids and ABA, and gene expression in a revertant of MT. The results of these experiments provide more evidence that CrMYB68 is an important regulator of carotenoid metabolism. [ABSTRACT FROM AUTHOR]

Interactions between the structure of a metabolic network and its functional properties underlie its evolutionary diversification, but the mechanism by which such interactions arise remains elusive. Particularly unclear is whether metabolic fluxes that determine the concentrations of compounds produced by a metabolic network, are causally linked to a network's structure or emerge independently of it. A direct empirical study of populations where both structural and functional properties vary among individuals' metabolic networks is required to establish whether changes in structure affect the distribution of metabolic flux. In a population of house finches ( Haemorhous mexicanus), we reconstructed full carotenoid metabolic networks for 442 individuals and uncovered 11 structural variants of this network with different compounds and reactions. We examined the consequences of this structural diversity for the concentrations of plumage-bound carotenoids produced by flux in these networks. We found that concentrations of metabolically derived, but not dietary carotenoids, depended on network structure. Flux was partitioned similarly among compounds in individuals of the same network structure: within each network, compound concentrations were closely correlated. The highest among-individual variation in flux occurred in networks with the strongest among-compound correlations, suggesting that changes in the magnitude, but not the distribution of flux, underlie individual differences in compound concentrations on a static network structure. These findings indicate that the distribution of flux in carotenoid metabolism closely follows network structure. Thus, evolutionary diversification and local adaptations in carotenoid metabolism may depend more on the gain or loss of enzymatic reactions than on changes in flux within a network structure. [ABSTRACT FROM AUTHOR]

DNA methylation is known to play an important role in various developmental processes in plants. However, there is a general lack of understanding about the possible functions of DNA methylation in fruit trees. Using callus as a model, methylome, transcriptome and metabolite changes were assessed after treatment with the DNA methyltransferase inhibitor 5-azacytidine (5azaC). Genome-wide methylome analysis revealed the demethylation of a diverse of genes, including many genes encoding transcription factors (TFs), genes involved in biological processes, and the up-regulation of a wide range of transposable elements (TEs). Combined with the RNAseq data, we observed no obvious genome-wide correlation between the changes in methylation status and expression levels. Furthermore, 5azaC treatment induced carotenoid degradation along with strong activation of carotenoid cleavage dioxygenases 1 (CpCCD1). Functional complementation analysis in bacterial system showed that CpCCD1 exhibited strong catalytic activities toward zeaxanthin, b-carotene and lycopene. In summary, 5azaC treatments induced carotenoid degradation by CpCCD1 activation and led to a genome-wide demethylation effect. [ABSTRACT FROM AUTHOR]

Carotenoid supplementation can prevent and reduce the risk of age-related macular degeneration (AMD) and other ocular disease, but until now, there has been no validated and well-characterized mouse model which can be employed to investigate the protective mechanism and relevant metabolism of retinal carotenoids. β-Carotene oxygenases 1 and 2 (BCO1 and BCO2) are the only two carotenoid cleavage enzymes found in animals. Mutations of the bco2 gene may cause accumulation of xanthophyll carotenoids in animal tissues, and BCO1 is involved in regulation of the intestinal absorption of carotenoids. To determine whether or not mice deficient in BCO1 and/or BCO2 can serve as a macular pigment mouse model, we investigated the retinal accumulation of carotenoids in these mice when fed with zeaxanthin, lutein, or β-carotene using an optimized carotenoid feeding method. HPLC analysis revealed that all three carotenoids were detected in sera, livers, retinal pigment epithelium (RPE)/choroids, and retinas of all of the mice, except that no carotenoid was detectable in the retinas of wild type (WT) mice. Significantly higher amounts of zeaxanthin and lutein accumulated in the retinas of BCO2 knockout ( bco2 -/- ) mice and BCO1/BCO2 double knockout ( bco1 -/- /bco2 -/- ) mice relative to BCO1 knockout ( bco1 -/- ) mice, while bco1 -/- mice preferred to take up β-carotene. The levels of zeaxanthin and lutein were higher than β-carotene levels in the bco1 -/- /bco2 -/- retina, consistent with preferential uptake of xanthophyll carotenoids by retina. Oxidative metabolites were detected in mice fed with lutein or zeaxanthin but not in mice fed with β-carotene. These results indicate that bco2 -/- and bco1 -/- /bco2 -/- mice could serve as reasonable non-primate models for macular pigment function in the vertebrate eye, while bco1 -/- mice may be more useful for studies related to β-carotene. [ABSTRACT FROM AUTHOR]

[Display omitted] • Melatonin treatment maintained the sensory quality of postharvest broccoli florets. • Melatonin induced the enrichment of endogenous melatonin in broccoli florets. • Melatonin down-regulated expression of carotenoid metabolism-related genes. • Melatonin inhibited contents of β-carotene, β-cryptoxanthin, zeaxanthin and lutein. The effect of melatonin treatment on the carotenoid metabolism in broccoli florets during storage was explored. The results indicated that 100 µmol/L of melatonin maintained the sensory quality of broccoli florets, which retarded the increase of the L* value and the decrease of the H value. Melatonin treatment increased the activities of tryptophan decarboxylase (TDC), tryptamine 5-hydroxylase (T5H), serotonin N -acetyltransferase (SNAT) and N -acetylserotonin methyltransferase (ASMT), leading to the enrichment of endogenous melatonin content in broccoli florets. Meanwhile, the treatment inhibited the concentrations of β-carotene, β-cryptoxanthin, zeaxanthin and lutein, which was beneficial in delaying the yellowing of broccoli. In addition, a series of carotenoid biosynthetic genes such as BoPSY, BoPDS, BoZDS, BoLCYβ and BoZEP was also suppressed by melatonin. Further analysis revealed that the lower carotenoid content and the down-regulated BoNCED expression in treated broccoli resulted in less accumulation of abscisic acid precursors, inhibiting abscisic acid production during the yellowing process. [ABSTRACT FROM AUTHOR]

Carotenoids play a critical role in animal and human health. Animals and humans are unable to synthesize carotenoids de novo, and therefore rely upon diet as sources of these compounds. However, major staple cereals often contain only small amounts of carotenoids in their grain. Consequently, there is considerable interest in genetic manipulation of carotenoid content in cereal grain. In this review, we focus on carotenoid metabolism and regulation in non-green plant tissues, as well as genetic manipulation in staple cereals such as rice, maize, and wheat. Significant progress has been made in three aspects: (1) seven carotenogenes play vital roles in carotenoid regulation in non-green plant tissues, including DXS (1-deoxyxylulose-5-phosphate synthase) influencing isoprenoid precursor supply, PSY (phytoene synthase), LCYB (β-cyclase) and LCYE (ε-cyclase) controlling biosynthesis, HYDB (1-hydroxy-2-methyl-2-(E)-butenyl 4-diphosphate reductase) and CCDs (carotenoid cleavage dioxygenases) responsible for degradation, and OR (orange) conditioning sequestration sink; (2) pro-vitamin A-biofortified crops, such as rice and maize, were developed by either metabolic engineering or marker-assisted breeding; (3) QTLs for carotenoid content on chromosomes 3B, 7A, and 7B were consistently identified, eight carotenogenes including 23 loci were detected, and ten gene-specific markers for carotenoid accumulation were developed and applied in wheat improvement. A comprehensive and deeper understanding of the regulatory mechanisms of carotenoid metabolism in crops will be benefitical in improving our precision in improving carotenoid contents. Genomic selection and gene editing are emerging as transformative technologies for vitamin A biofortification.