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Temporal analysis reveals a key role for VTE5 in vitamin E biosynthesis in olive fruit during on-tree development.

ABSTRACT: The aim of this work was to generate a high resolution temporal mapping of the biosynthetic pathway of vitamin E in olive fruit (Olea europaea cv. "Koroneiki") during 17 successive on-tree developmental stages. Fruit material was collected from the middle of June until the end of January, corresponding to 6-38 weeks after flowering (WAF). Results revealed a variable gene regulation pattern among 6-38 WAF studied and more pronounced levels of differential regulation of gene expression for the first and intermediate genes in the biosynthetic pathway (VTE5, geranylgeranyl reductase, HPPD, VTE2, HGGT and VTE3) compared with the downstream components of the pathway (VTE1 and VTE4). Notably, expression of HGGT and VTE2 genes were significantly suppressed throughout the developmental stages examined. Metabolite analysis indicated that the first and intermediate stages of development (6-22 WAF) have higher concentrations of tocochromanols compared with the last on-tree stages (starting from 24 WAF onwards). The concentration of ?-tocopherol (16.15 ± 0.60-32.45 ± 0.54 mg/100 g F.W.) were substantially greater (up to 100-fold) than those of ?-, ?-, and ?-tocopherols (0.13 ± 0.01-0.25 ± 0.03 mg/100 g F.W., 0.13 ± 0.01-0.33 ± 0.04 mg/100 g F.W., 0.14 ± 0.01-0.28 ± 0.01 mg/100 g F.W., respectively). In regard with tocotrienol content, only ?-tocotrienol was detected. Overall, olive fruits (cv. "Koroneiki") exhibited higher concentrations of vitamin E until 22 WAF as compared with later WAF, concomitant with the expression profile of phytol kinase (VTE5), which could be used as a marker gene due to its importance in the biosynthesis of vitamin E. To the best of our knowledge, this is the first study that explores the complete biosynthetic pathway of vitamin E in a fruit tree crop of great horticultural importance such as olive, linking molecular gene expression analysis with tocochromanol content.

SUBMITTER: Georgiadou EC

PROVIDER: S-EPMC4617049 | biostudies-literature | 2015

REPOSITORIES: biostudies-literature

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Temporal analysis reveals a key role for VTE5 in vitamin E biosynthesis in olive fruit during on-tree development.

Georgiadou Egli C EC Ntourou Thessaloniki T Goulas Vlasios V Manganaris George A GA Kalaitzis Panagiotis P Fotopoulos Vasileios V

Frontiers in plant science 20151021

The aim of this work was to generate a high resolution temporal mapping of the biosynthetic pathway of vitamin E in olive fruit (Olea europaea cv. "Koroneiki") during 17 successive on-tree developmental stages. Fruit material was collected from the middle of June until the end of January, corresponding to 6-38 weeks after flowering (WAF). Results revealed a variable gene regulation pattern among 6-38 WAF studied and more pronounced levels of differential regulation of gene expression for the fir ...[more]

PMID: 26557125

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Project description:The term vitamin E refers to a group of eight lipophilic compounds known as tocochromanols. The tocochromanols are divided into two groups, that is, tocopherols and tocotrienols, with four forms each, namely α-, β-, γ-, and δ-. In order to explore the temporal biosynthesis of tocochromanols in olive (Olea europaea cv. 'Koroneiki') fruit during on-tree development and ripening over successive growing years, a combined array of analytical, molecular, bioinformatic, immunoblotting, and antioxidant techniques were employed. Fruits were harvested at eight successive developmental stages [10-30 weeks after flowering (WAF)], over three consecutive years. Intriguingly, climatic conditions affected relative transcription levels of vitamin E biosynthetic enzymes; a general suppression to induction pattern (excluding VTE5) was monitored moving from the 1st to the 3rd growing year, probably correlated to decreasing rainfall levels and higher temperature, particularly at the fruit ripening stage. A gradual diminution of VTE5 protein content was detected during the fruit development of each year, with a marked decrease occurring after 16 WAF. Alpha-tocopherol was the most abundant metabolite with an average percentage of 96.82 ± 0.23%, 91.13 ± 0.95%, and 88.53 ± 0.96% (during the 1st, 2nd, and 3rd year, respectively) of total vitamin E content in 10-30 WAF. The concentrations of α-tocopherol revealed a generally declining pattern, both during the on-tree ripening of the olive fruit and across the 3 years, accompanied by a parallel decline of the total antioxidant capacity of the drupe. Contrarily, all other tocochromanols demonstrated an inverse pattern with lowest levels being recorded during the 1st year. It is likely that, in a defense attempt against water deficit conditions and increased air temperature, transcription of genes involved in vitamin E biosynthesis (excluding VTE5) is up-regulated in olive fruit, probably leading to the blocking/deactivating of the pathway through a negative feedback regulatory mechanism.

Project description:BackgroundOlive (Olea europaea L.) fruits contain numerous secondary metabolites, primarily phenolics, terpenes and sterols, some of which are particularly interesting for their nutraceutical properties. This study will attempt to provide further insight into the profile of olive phenolic compounds during fruit development and to identify the major genetic determinants of phenolic metabolism.ResultsThe concentration of the major phenolic compounds, such as oleuropein, demethyloleuropein, 3-4 DHPEA-EDA, ligstroside, tyrosol, hydroxytyrosol, verbascoside and lignans, were measured in the developing fruits of 12 olive cultivars. The content of these compounds varied significantly among the cultivars and decreased during fruit development and maturation, with some compounds showing specificity for certain cultivars. Thirty-five olive transcripts homologous to genes involved in the pathways of the main secondary metabolites were identified from the massive sequencing data of the olive fruit transcriptome or from cDNA-AFLP analysis. Their mRNA levels were determined using RT-qPCR analysis on fruits of high- and low-phenolic varieties (Coratina and Dolce d'Andria, respectively) during three different fruit developmental stages. A strong correlation was observed between phenolic compound concentrations and transcripts putatively involved in their biosynthesis, suggesting a transcriptional regulation of the corresponding pathways. OeDXS, OeGES, OeGE10H and OeADH, encoding putative 1-deoxy-D-xylulose-5-P synthase, geraniol synthase, geraniol 10-hydroxylase and arogenate dehydrogenase, respectively, were almost exclusively present at 45 days after flowering (DAF), suggesting that these compounds might play a key role in regulating secoiridoid accumulation during fruit development.ConclusionsMetabolic and transcriptional profiling led to the identification of some major players putatively involved in biosynthesis of secondary compounds in the olive tree. Our data represent the first step towards the functional characterisation of important genes for the determination of olive fruit quality.

Project description:BackgroundOlea europaea L. is a traditional tree crop of the Mediterranean basin with a worldwide economical high impact. Differently from other fruit tree species, little is known about the physiological and molecular basis of the olive fruit development and a few sequences of genes and gene products are available for olive in public databases. This study deals with the identification of large sets of differentially expressed genes in developing olive fruits and the subsequent computational annotation by means of different software.ResultsmRNA from fruits of the cv. Leccino sampled at three different stages [i.e., initial fruit set (stage 1), completed pit hardening (stage 2) and veraison (stage 3)] was used for the identification of differentially expressed genes putatively involved in main processes along fruit development. Four subtractive hybridization libraries were constructed: forward and reverse between stage 1 and 2 (libraries A and B), and 2 and 3 (libraries C and D). All sequenced clones (1,132 in total) were analyzed through BlastX against non-redundant NCBI databases and about 60% of them showed similarity to known proteins. A total of 89 out of 642 differentially expressed unique sequences was further investigated by Real-Time PCR, showing a validation of the SSH results as high as 69%. Library-specific cDNA repertories were annotated according to the three main vocabularies of the gene ontology (GO): cellular component, biological process and molecular function. BlastX analysis, GO terms mapping and annotation analysis were performed using the Blast2GO software, a research tool designed with the main purpose of enabling GO based data mining on sequence sets for which no GO annotation is yet available. Bioinformatic analysis pointed out a significantly different distribution of the annotated sequences for each GO category, when comparing the three fruit developmental stages. The olive fruit-specific transcriptome dataset was used to query all known KEGG (Kyoto Encyclopaedia of Genes and Genomes) metabolic pathways for characterizing and positioning retrieved EST records. The integration of the olive sequence datasets within the MapMan platform for microarray analysis allowed the identification of specific biosynthetic pathways useful for the definition of key functional categories in time course analyses for gene groups.ConclusionThe bioinformatic annotation of all gene sequences was useful to shed light on metabolic pathways and transcriptional aspects related to carbohydrates, fatty acids, secondary metabolites, transcription factors and hormones as well as response to biotic and abiotic stresses throughout olive drupe development. These results represent a first step toward both functional genomics and systems biology research for understanding the gene functions and regulatory networks in olive fruit growth and ripening.

Project description:Vitamin D deficiency poses a major global health challenge, shaped by environmental and genetic determinants. In this genetic context, the recently discovered rs11542462 nonsense variant in the short-chain dehydrogenase/reductase 42E1 (SDR42E1) gene emerges as a critical regulator, though its biological implications remain largely unexplored. Our comprehensive bioinformatic assessments revealed pronounced expression of SDR42E1 in skin keratinocytes and the analogous HaCat human keratinocyte cell lines, prompting us to select the latter as an experimental model. Employing CRISPR/Cas9 technology and an integrated multi-omics approach, combining transcriptomics and proteomics, we explored SDR42E1's functions, focusing on its localization, role in lipid and steroid biosynthesis, and the consequent effects on vitamin D biosynthesis. We highlighted the significant cytoplasmic presence of the SDR42E1 protein, essential for lipid and steroid production. Our whole transcriptomics analysis of the SDR42E1 depleted model showed a notable disruption in the steroid biosynthesis pathway by 1.6-fold (P = 0.03). This disruption was accompanied by considerable alterations in genes involved in vitamin D synthesis, a finding supported by our proteomics data. Notably, SERPINB2 (P = 2.17E−103), EBP (P = 2.46E−13), and DHCR7 (P = 8.03E−09) showed significant elevations by ~2-3 fold, while ALPP (P < 2.2E−308), SLC7A5 (P = 1.96E−215), and CYP26A1 (P = 1.06E−08) decreased by ~1.5-3 fold. These alterations led to an accumulation of 7-dehydrocholesterol, preventing its conversion to vitamin D3, as evidenced by our drug enrichment analysis (P = 4.39E−06). This investigation unveils SDR42E1's pivotal role in vitamin D homeostasis and its broader impact on metabolic pathways. Our findings provide a foundation for future research and highlight the potential of precision medicine in addressing vitamin D deficiency, offering significant insights into the genetic regulation of vitamin D biosynthesis.

Dataset Information

Temporal analysis reveals a key role for VTE5 in vitamin E biosynthesis in olive fruit during on-tree development.

Publications

Temporal analysis reveals a key role for VTE5 in vitamin E biosynthesis in olive fruit during on-tree development.

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