The dominant allele Aft induces a shift from flavonol to anthocyanin production in response to UV-B radiation in tomato fruit

The introgression of the A ft allele into domesticated tomato induced a shift from flavonol to anthocyanin production in response to UV-B radiation, while the hp - 1 allele negatively influenced the response of flavonoid biosynthesis to UV-B. Introgression of the dominant allele Anthocyanin fruit (Aft) from Solanum chilense induces anthocyanin accumulation in the peel of tomato (Solanum lycopersicum L.) fruit. UV-B radiation can influence plant secondary metabolism regulating the expression of several genes, among which those involved in flavonoid biosynthesis. Here, we investigated whether post-harvest UV-B treatment could up-regulate flavonoid production in tomato fruits and whether the Aft allele could affect flavonoid biosynthesis under UV-B radiation. Mature green fruits of an anthocyanin-rich tomato mutant line (SA206) and of its wild-type reference, cv. Roma, were daily subjected to post-harvest UV-B treatment until full ripening. Up-regulation of CHS and CHI transcription by UV-B treatment induced flavonoid accumulation in the peel of cv. Roma. Conversely, UV-B decreased the total flavonoid content and CHS transcript levels in the SA206 peel. SA206 being a double mutant containing also hp-1 allele, we investigated also the behavior of hp-1 fruit. The decreased peel flavonoid accumulation and gene transcription in response to UV-B suggest that hp-1 allele is involved in the marked down-regulation of the flavonoid biosynthesis observed in SA206 fruit. Interestingly, in SA206, UV-B radiation promoted the synthesis of delphinidin, petunidin, and malvidin by increasing F3'5'H and DFR transcription, but it decreased rutin production, suggesting a switch from flavonols to anthocyanins. Finally, although UV-B radiation does not reach the inner fruit tissues, it down-regulated flavonoid biosynthesis in the flesh of both genotypes. This study provides, for the first time, evidence that the presence of the functional Aft allele, under UV-B radiation, redirects flavonoid synthesis towards anthocyanin production and suggests that the hp-1 allele negatively influences the response of flavonoid biosynthesis to UV-B

Remarkable antioxidant and anti-inflammatory properties of wastes from fermented pomegranate and ellagic acid release profile under simulated digestion conditions.

Fruit and vegetable fermentation techniques are widely used as methodologies to preserve food. Fermentation processes lead to considerable amounts of residues that could contain bioactive compounds deriving from those originally present in the raw materials and modified by fermentation, or accumulate single components at high concentrations. Wastes deriving from production of \u201cwines\u201d by fermentation of Punica granatum, fermented pomegranate wastes (FPW), showed a marked antioxidant activity in a series of conventional chemical tests. HPLC /MS analyses of the methanol extracts showed the presence of ellagic acid (EA) as the main component at levels up to 40% on a w/w basis together with minor species that were further investigated by 1D and 2D NMR spectroscopy without fractionation. Experiments using murine macrophages showed a stronger effect of the FPW extracts in reducing the LPS-induced expression of pro-inflammatory genes IL-1\u3b2, IL-6, TNF-\uf061 and iNOS compared to the fresh fruit extracts to a degree correlating with EA levels. Under simulated gastro intestinal conditions, EA was slowly released from FPW up to 30% of the overall content over 2 hrs only at the slightly alkaline pHs simulating the small intestine environment, suggesting a potential of the material in nutraceuticals and other applications

The Arabidopsis <i>NF-YA3</i> and <i>NF-YA8</i> Genes Are Functionally Redundant and Are Required in Early Embryogenesis

<div><p>Nuclear factor Y (NF-Y) is a trimeric transcription factor composed of three distinct subunits called NF-YA, NF-YB and NF-YC. In <i>Arabidopsis thaliana</i>, NF-Y subunits are known to play roles in many processes, such as gametogenesis, embryogenesis, seed development, drought resistance, ABA signaling, flowering time, primary root elongation, Endoplasmic Reticulum (ER) stress response and blue light responses. Here, we report that the closely related <i>NF-YA3</i> and <i>NF-YA8</i> genes control early embryogenesis. Detailed GUS and <i>in situ</i> analyses showed that <i>NF-YA3</i> and <i>NF-YA8</i> are expressed in vegetative and reproductive tissues with the highest expression being during embryo development from the globular to the torpedo embryo stage. Plants from the <i>nf-ya3</i> and <i>nf-ya8</i> single mutants do not display any obvious phenotypic alteration, whereas <i>nf-ya3 nf-ya8</i> double mutants are embryo lethal. Morphological analyses showed that the <i>nf-ya3 nf-ya8</i> embryos fail to undergo to the heart stage and develop into abnormal globular embryos with both proembryo and suspensor characterized by a disordered cell cluster with an irregular shape, suggesting defects in embryo development. The suppression of both <i>NF-YA3</i> and <i>NF-YA8</i> gene expression by RNAi experiments resulted in defective embryos that phenocopied the <i>nf-ya3 nf-ya8</i> double mutants, whereas complementation experiments partially rescued the abnormal globular <i>nf-ya3 nf-ya8</i> embryos, confirming that <i>NF-YA3</i> and <i>NF-YA8</i> are required in early embryogenesis. Finally, the lack of GFP expression of the auxin responsive <i>DR5rev::GFP</i> marker line in double mutant embryos suggested that mutations in both <i>NF-YA3</i> and <i>NF-YA8</i> affect auxin response in early developing embryos. Our findings indicate that <i>NF-YA3</i> and <i>NF-YA8</i> are functionally redundant genes required in early embryogenesis <i>of Arabidopsis thaliana</i>.</p></div

Structure of the T-DNA insertional mutants and semi-quantitative RT-PCR analyses.

<p>A) Schematic representation of the <i>NF-YA3</i> and <i>NF-YA8</i> genes. Position of the T-DNA insertion sites for <i>nf-ya3</i> and <i>nf-ya8</i> mutant alleles are indicated. Exons are boxed and lines between boxes represent introns. White boxes correspond to coding sequence exons, black boxes correspond to 5′ UTR and 3′UTR. LB, left border; RB, right border. As determined by sequencing of the T-DNA/gene junctions, the duplicated genomic region of 54 nucleotides in the <i>nf-ya8</i> mutant allele is shown. The highly conserved domain, composed by Subunit Association Domain (SAD) and DNA binding domain (BD) and the linker region (lined box), is indicated. B) RT-PCR of <i>NF-YA3</i> and <i>NF-YA8</i> transcript in <i>nf-ya3</i> and <i>nf-ya8</i> mutants. Total RNA was extracted from developing siliques of wild-type and mutant plants and analyzed by RT-PCR. All cDNA samples were standardized using a set of primers specific for the <i>ACTIN 2</i> (<i>ACT2</i>) gene.</p

Percentage of normal and arrested seeds of <i>nf-ya3/nf-ya3 NF-YA8/nf-ya8</i> mutant plants.

a<p>Chi-square test for a 3:1 phenotypic segregation hypothesis.</p

A biochemical and molecular dissection of fruit anthocyanins in anthocyanin-rich tomato mutant after post-harvest UV-B treatment

Phenylpropanoid consumption is well-known to be associated with a reduced risk of several human diseases. Many studies demonstrated that UV-B may influence phenylpropanoid metabolism. This work investigates how UV-B radiation can modulate the transcription of different genes involved in the phenylpropanoid pathway, in fruits of the tomato line SA206-1- 2-2 (with the anthocyanin fruit Aft gene), characterised by purple spots on fruit surface, and in its wild type, cv Roma. Fruits, harvested at mature green stage, were irradiated with UV-B (1h a day, 6.08 kJ m-2 d-1) until red ripe stage. UV-B treatment induced an overall stimulation of the flavonoid biosynthetic pathway in Roma peel. In SA206 peel, early biosynthetic genes (CHS, F3H and F3’H) were down- regulated, leading to a decrease in downstream metabolites. However, F3’5’H and DFR transcription, as well as delphinidin, petunidin and malvidin levels, increased, indicating a metabolic shift towards anthocyanin synthesis in response to UV-B irradiation

Expression pattern of DR5rev::GFP in wild-type and nf-ya3 nf-ya8 embryo.

<p>(A,B,C) <i>DR5rev::GFP</i> expression in wild-type embryos. Embryo at proembryo stage (A), globular stage (B) and at cotyledon stage (C). (D,E,F) <i>DR5rev::GFP</i> expression in <i>nf-ya3 nf-ya8</i> defective embryos. Confocal laser scanning images; bars = 25 µm.</p

Phenotypic analysis of <i>nf-ya3/nf-ya3 NF-YA8/nf-ya8</i> siliques.

<p>A) Siliques of wild-type plants with normal seeds. B) Siliques of <i>nf-ya3/nf-ya3 NF-YA8/nf-ya8</i> mutant plants with arrested seeds (arrow). Bar = 500 µm.</p

Complementation of <i>nf-ya3/nf-ya3 nf-ya8/nf-ya8</i> double mutants with <i>35S::CDS-NF-YA3</i> construct.

a<p><i>nf-ya3/nf-ya3 NF-YA8/nf-ya8</i> T1 plants carrying the <i>35S::CDS-NF-YA3</i> complementation construct.</p>b<p>Arrested seeds expected in case of full complementation.</p>c<p>Chi-square test for 1/16 (one copy of construct inserted) and 1/64 (two copies of construct inserted) segregation hypothesis.</p

Segregation analysis of F3 progeny from <i>nf-ya3/nf-ya3 NF-YA8/nf-ya8</i> plants.

a<p>Chi-square test for a 2∶1 segregation hypothesis.</p