Lycopene β-cyclase expression influences plant physiology, development, and metabolism in tobacco plants.
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ABSTRACT:
SUBMITTER: Kossler S
PROVIDER: S-EPMC8006556 | biostudies-literature |
REPOSITORIES: biostudies-literature
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Project description:Malnutrition affects growth and development in humans and causes socio-economic losses. Normal maize is deficient in essential amino acids, lysine and tryptophan; and vitamin-A. Crop biofortification is a sustainable and economical approach to alleviate micronutrient malnutrition. We combined favorable alleles of crtRB1 and lcyE genes into opaque2 (o2)-based four inbreds viz. QLM11, QLM12, QLM13, and QLM14 using marker-assisted backcross breeding. These are parents of quality protein maize versions of two elite hybrids viz. Buland and PMH1, grown in India. Gene-based SSRs for o2 and InDel markers for crtRB1 and lcyE were successfully employed for foreground selection in BC1F1, BC2F1, and BC2F2 generations. The recurrent parent genome recovery ranged from 88.9 to 96.0% among introgressed progenies. Kernels of pyramided lines possessed a high concentration of proA (7.14-9.63 ppm), compared to 1.05 to 1.41 ppm in the recurrent parents, while lysine and tryptophan ranged from 0.28-0.44% and 0.07-0.09%, respectively. The reconstituted hybrids (RBuland and RPMH1) showed significant enhancement of endosperm proA (6.97-9.82 ppm), tryptophan (0.07-0.09%), and lysine (0.29-0.43%), while grain yield was at par with their original versions. The dissemination of reconstituted hybrids holds significant promise to alleviate vitamin-A deficiency and protein-energy malnutrition in developing countries.
| S-EPMC8209105 | biostudies-literature
Project description:The ancestor of tomato underwent whole genome triplication ca. 71 Myr ago followed by widespread gene loss. However, few of the triplicated genes are retained in modern day tomato including lycopene beta cyclase that mediates conversion of lycopene to ?-carotene. The fruit specific ?-carotene formation is mediated by a chromoplast-specific paralog of lycopene beta cyclase (CYC-B) gene. Presently limited information is available about how the variations in CYC-B gene contributed to its neofunctionalization. CYC-B gene in tomato clade contained several SNPs and In-Dels in the coding sequence (33 haplotypes) and promoter region (44 haplotypes). The CYC-B gene coding sequence in tomato appeared to undergo purifying selection. The transit peptide sequence of CYC-B protein was predicted to have a stronger plastid targeting signal than its chloroplast specific paralog indicating a possible neofunctionalization. In promoter of two Bog (Beta old gold) mutants, a NUPT (nuclear plastid) DNA fragment of 256 bp, likely derived from a S. chilense accession, was present. In transient expression assay, this promoter was more efficient than the "Beta type" promoter. CARGATCONSENSUS box sequences are required for the binding of the MADS-box regulatory protein RIPENING INHIBITOR (RIN). The loss of CARGATCONSENSUS box sequence from CYC-B promoter in tomato may be related to attenuation of its efficiency to promote higher accumulation of ?-carotene than lycopene during fruit ripening.
| S-EPMC4829152 | biostudies-literature
Project description:Lycopene β-cyclases are key enzymes located at the branch point of the carotenoid biosynthesis pathway. However, the transcriptional regulatory mechanisms of LCYb1 in citrus with abundant carotenoid accumulation are still unclear. To understand the molecular basis of CsLCYb1 expression, we isolated and functionally characterized the 5' upstream sequences of CsLCYb1 from citrus. The full-length CsLCYb1 promoter and a series of its 5' deletions were fused to the β-glucuronidase (GUS) reporter gene and transferred into different plants (tomato, Arabidopsis and citrus callus) to test the promoter activities. The results of all transgenic species showed that the 1584 bp upstream region from the translational start site displayed maximal promoter activity, and the minimal promoter containing 746 bp upstream sequences was sufficient for strong basal promoter activity. Furthermore, the CsLCYb1 promoter activity was developmentally and tissue-specially regulated in transgenic Arabidopsis, and it was affected by multiple hormones and environmental cues in transgenic citrus callus under various treatments. Finer deletion analysis identified an enhancer element existing as a tandem repeat in the promoter region between -574 to -513 bp and conferring strong promoter activity. The copy numbers of the enhancer element differed among various citrus species, leading to the development of a derived simple sequence repeat marker to distinguish different species. In conclusion, this study elucidates the expression characteristics of the LCYb1 promoter from citrus and further identifies a novel enhancer element required for the promoter activity. The characterized promoter fragment would be an ideal candidate for genetic engineering and seeking of upstream trans-acting elements.
| S-EPMC5020073 | biostudies-literature
Project description:Carotenoids are the main pigments responsible of the colouration of Citrus fruits. The beta-cyclization of lycopene, catalysed by the lycopene beta-cyclases (beta-LCY), seems to be a key regulatory step of the carotenoid pathway. In the present study, two beta-LCYs from orange fruits (Citrus sinensis), named Csbeta-LCY1 and Csbeta-LCY2 have been isolated and the activity of the encoded proteins was demonstrated by functional analysis. Csbeta-LCY1 was expressed at low levels and remained relatively constant during fruit ripening while Csbeta-LCY2 showed a chromoplast-specific expression and a marked induction in both peel and pulp of orange fruits in parallel with the accumulation of beta,beta-xanthophylls. The potential involvement of Csbeta-LCY2 in the accumulation of lycopene, characteristic of some Citrus species such as red grapefruits, was investigated. Expression of Csbeta-LCY2 and another seven carotenoid biosynthetic genes were studied in the peel and pulp of the high lycopene-accumulating grapefruit, Star Ruby, and compared with those of ordinary Navel orange. In Star Ruby, the accumulation of lycopene during fruit maturation was associated with a substantial reduction in the expression of both beta-LCY2 and beta-CHX genes with respect to Navel orange. Moreover, two different alleles of beta-LCY2: beta-LCY2a and beta-LCY2b were isolated from both genotypes, and functional assays demonstrated that the lycopene beta-cyclase activity of the allele b was almost null. Interestingly, Star Ruby grapefruit predominantly expressed the unfunctional beta-LCY2b allele during fruit ripening whereas Navel oranges preferably expressed the functional allele. It is suggested that the presence of diverse alleles of the beta-LCY2 gene, encoding enzymes with altered activity, with different transcript accumulation may be an additional regulatory mechanism of carotenoid synthesis involved in the accumulation of lycopene in red grapefruits.
| S-EPMC2671624 | biostudies-literature
Project description:Lycopene ?-cyclase (LYCB) is the key enzyme for the synthesis of ?-carotene, a valuable component of the human diet. In this study, tomato constitutively express Lycb-1 was engineered. The ?-carotene level of transformant increased 4.1 fold, and the total carotenoid content increased by 30% in the fruits. In the transgenic line, the downstream ?-branch metabolic fluxes were repressed during the three developmental stages while ?-carotene content increased in the ripe stage. Microarray analysis in the ripe stage revealed that the constitutive expression of Lycb-1 affected a number of pathways including the synthesis of fatty acids, flavonoids and phenylpropanoids, the degradation of limonene and pinene, starch and sucrose metabolism and photosynthesis. This study provided insight into the regulatory effect of Lycb-1 gene on plant carotenoid metabolism and fruit transcriptome.
| S-EPMC3286453 | biostudies-literature
Project description:BackgroundCarotenoids are a group of C40 isoprenoid molecules that play diverse biological and ecological roles in plants. Tomato is an important vegetable in human diet and provides the vitamin A precursor beta-carotene. Genes encoding enzymes involved in carotenoid biosynthetic pathway have been cloned. However, regulation of genes involved in carotenoid biosynthetic pathway and accumulation of specific carotenoid in chromoplasts are not well understood. One of the approaches to understand regulation of carotenoid metabolism is to characterize the promoters of genes encoding proteins involved in carotenoid metabolism. Lycopene beta-cyclase is one of the crucial enzymes in carotenoid biosynthesis pathway in plants. Its activity is required for synthesis of both alpha-and beta-carotenes that are further converted into other carotenoids such as lutein, zeaxanthin, etc. This study describes the isolation and characterization of chromoplast-specific Lycopene beta-cyclase (CYC-B) promoter from a green fruited S. habrochaites genotype EC520061.ResultsA 908 bp region upstream to the initiation codon of the Lycopene beta-cyclase gene was cloned and identified as full-length promoter. To identify promoter region necessary for regulating developmental expression of the ShCYC-B gene, the full-length promoter and its three different 5' truncated fragments were cloned upstream to the initiation codon of GUS reporter cDNA in binary vectors. These four plant transformation vectors were separately transformed in to Agrobacterium. Agrobacterium-mediated transient and stable expression systems were used to study the GUS expression driven by the full-length promoter and its 5' deletion fragments in tomato. The full-length promoter showed a basal level activity in leaves, and its expression was upregulated > 5-fold in flowers and fruits in transgenic tomato plants. Deletion of -908 to -577 bp 5' to ATG decreases the ShCYC-B promoter strength, while deletion of -908 to -437 bp 5' to ATG led to significant increase in the activity of GUS in the transgenic plants. Promoter deletion analysis led to the identification of a short promoter region (-436 bp to ATG) that exhibited a higher promoter strength but similar developmental expression pattern as compared with the full-length ShCYC-B promoter.ConclusionFunctional characterization of the full-length ShCYC-B promoter and its deletion fragments in transient expression system in fruto as well as in stable transgenic tomato revealed that the promoter is developmentally regulated and its expression is upregulated in chromoplast-rich flowers and fruits. Our study identified a short promoter region with functional activity and developmental expression pattern similar to that of the full-length ShCYC-B promoter. This 436 bp promoter region can be used in promoter::reporter fusion molecular genetic screens to identify mutants impaired in CYC-B expression, and thus can be a valuable tool in understanding carotenoid metabolism in tomato. Moreover, this short promoter region of ShCYC-B may be useful in genetic engineering of carotenoid content and other agronomic traits in tomato fruits.
| S-EPMC2923535 | biostudies-literature
Project description:The composition of carotenoids, along with anthocyanins and chlorophyll, accounts for the distinctive range of colour found in the Actinidia (kiwifruit) species. Lutein and beta-carotene are the most abundant carotenoids found during fruit development, with beta-carotene concentration increasing rapidly during fruit maturation and ripening. In addition, the accumulation of beta-carotene and lutein is influenced by the temperature at which harvested fruit are stored. Expression analysis of carotenoid biosynthetic genes among different genotypes and fruit developmental stages identified Actinidia lycopene beta-cyclase (LCY-beta) as the gene whose expression pattern appeared to be associated with both total carotenoid and beta-carotene accumulation. Phytoene desaturase (PDS) expression was the least variable among the different genotypes, while zeta carotene desaturase (ZDS), beta-carotene hydroxylase (CRH-beta), and epsilon carotene hydroxylase (CRH-epsilon) showed some variation in gene expression. The LCY-beta gene was functionally tested in bacteria and shown to convert lycopene and delta-carotene to beta-carotene and alpha-carotene respectively. This indicates that the accumulation of beta-carotene, the major carotenoid in these kiwifruit species, appears to be controlled by the level of expression of LCY-beta gene.
| S-EPMC2736891 | biostudies-literature
Project description:BackgroundChemical and biological processes dictate an individual organism's ability to recognize and respond to other organisms. A small but growing body of evidence suggests that plants may be capable of recognizing and responding to neighboring plants in a species specific fashion. Here we tested whether or not individuals of the invasive exotic weed, Centaurea maculosa, would modulate their defensive strategy in response to different plant neighbors.ResultsIn the greenhouse, C. maculosa individuals were paired with either conspecific (C. maculosa) or heterospecific (Festuca idahoensis) plant neighbors and elicited with the plant defense signaling molecule methyl jasmonate to mimic insect herbivory. We found that elicited C. maculosa plants grown with conspecific neighbors exhibited increased levels of total phenolics, whereas those grown with heterospecific neighbors allocated more resources towards growth. To further investigate these results in the field, we conducted a metabolomics analysis to explore chemical differences between individuals of C. maculosa growing in naturally occurring conspecific and heterospecific field stands. Similar to the greenhouse results, C. maculosa individuals accumulated higher levels of defense-related secondary metabolites and lower levels of primary metabolites when growing in conspecific versus heterospecific field stands. Leaf herbivory was similar in both stand types; however, a separate field study positively correlated specialist herbivore load with higher densities of C. maculosa conspecifics.ConclusionsOur results suggest that an individual C. maculosa plant can change its defensive strategy based on the identity of its plant neighbors. This is likely to have important consequences for individual and community success.
| S-EPMC3095278 | biostudies-literature
Project description:Ripening is an important stage of fruit development. To screen the genes associated with pigment formation in tomato fruit, a suppression subtractive hybridization (SSH) cDNA library was constructed by using tomato fruit in the green ripe and break ripe stages, and 129 differential genes were obtained. Using redness as a screening marker, virus-induced gene silencing (VIGS) of the differential genes was performed with a sprout vacuum-infiltration system (SVI). The results showed that silencing the SlNAP7 gene affected the chloroplast development of tomato leaves, manifesting as a photo-bleaching phenotype, and silenced fruit significantly affected the accumulation of lycopene, manifested as a yellow phenotype. In our study, we found that silencing the SlNAP7 gene downregulates the expression of the POR and PORA genes and destroys the normal development of the chloroplast. The expression of related genes included in the lycopene biosynthesis pathway was not significantly changed, but lycopene accumulation was significantly reduced in tomato fruit. Perhaps it was caused by the destruction of the chromoplast, which leads to the oxidation of lycopene. The results show that the SlNAP7 gene influences chloroplast development and lycopene accumulation in tomato.
| S-EPMC5144135 | biostudies-literature
Project description:Traditional yellow maize though contains high kernel carotenoids, the concentration of provitamin A (proA) is quite low (<2 ?g/g), compared to recommended level (15 ?g/g). It also possesses poor endosperm protein quality due to low concentration of lysine and tryptophan. Natural variant of crtRB1 (?-carotene hydroxylase) and lcyE (lycopene-?-cyclase) cause significant enhancement of proA concentration, while recessive allele, opaque2 (o2) enhances the level of these amino acids. Development of biofortified maize enriched in proA, lysine and tryptophan thus holds significance in alleviation of micronutrient malnutrition. In the present study, marker-assisted stacking of crtRB1, lcyE and o2 was undertaken in the genetic background of four maize hybrids (HQPM1, HQPM4, HQPM5, and HQPM7) popularly grown in India. HP704-22 and HP704-23 were used as donors, while four elite QPM parents viz., HKI161, HKI163, HKI193-1, and HKI193-2 were used as recipients. CrtRB1 showed severe segregation distortion, while lcyE segregated as per the expectation. Recovery of recurrent parent genome (RPG) among selected backcross progenies ranged from 89 to 93%. Introgressed progenies possessed high concentration of proA (7.38-13.59 ?g/g), compared to 1.65-2.04 ?g/g in the recurrent parents. The reconstituted hybrids showed an average of 4.5-fold increase in proA with a range of 9.25-12.88 ?g/g, compared to original hybrids (2.14-2.48 ?g/g). Similar plant-, ear-, and grain- characteristics of improved versions of both inbreds and hybrids were observed when evaluated with their respective original versions. Mean lysine (0.334%) and tryptophan (0.080%) of the improved hybrids were at par with the original versions (lysine: 0.340%, tryptophan: 0.083%). Improved hybrids also possessed similar grain yield potential (6,301-8,545 kg/ha) with their original versions (6,135-8,479 kg/ha) evaluated at two locations. This is the first study of staking crtRB1-, lcyE-, and o2-, favorable alleles in single genetic background. The improved inbreds can be effectively used as potential donor for independent and/or simultaneous introgression of crtRB1, lcyE, and o2 in the future breeding programme. These biofortified maize hybrids, rich in proA, lysine and tryptophan will hold great promise for nutritional security.
| S-EPMC5826225 | biostudies-literature