Katia Petroni

Recent advances on the regulation of anthocyanin synthesis in reproductive organs

Anthocyanins represent the major red, purple, violet and blue pigments in many flowers and fruits. They attract pollinators and seed dispersers and defend plants against abiotic and biotic stresses. Anthocyanins are produced by a specific branch of the flavonoid pathway, which is differently regulated in monocot and dicot species. In the monocot maize, the anthocyanin biosynthesis genes are activated as a single unit by a ternary complex of MYB-bHLH-WD40 transcription factors (MBW complex). In the dicot Arabidopsis, anthocyanin biosynthesis genes can be divided in two subgroups: early biosynthesis genes (EBGs) are activated by co-activator independent R2R3-MYB transcription factors, whereas late biosynthesis genes (LBGs) require an MBW complex. In addition to this, a complex regulatory network of positive and negative feedback mechanisms controlling anthocyanin synthesis in Arabidopsis has been described. Recent studies have broadened our understanding of the regulation of anthocyanin synthesis in flowers and fruits, indicating that a regulatory system based on the cooperation of MYB, bHLH and WD40 proteins that control floral and fruit pigmentation is common to many dicot species.

Function search in a large transcription factor gene family in Arabidopsis: assessing the potential of reverse genetics to identify insertional mutations in R2R3 MYB genes.

More than 92 genes encoding MYB transcription factors of the R2R3 class have been described in Arabidopsis. The functions of a few members of this large gene family have been described, indicating important roles for R2R3 MYB transcription factors in the regulation of secondary metabolism, cell shape, and disease resistance, and in responses to growth regulators and stresses. For the majority of the genes in this family, however, little functional information is available. As the first step to characterizing these genes functionally, the sequences of >90 family members, and the map positions and expression profiles of >60 members, have been determined previously. An important second step in the functional analysis of the MYB family, through a process of reverse genetics that entails the isolation of insertion mutants, is described here. For this purpose, a variety of gene disruption resources has been used, including T-DNA–insertion populations and three distinct populations that harbor transposon insertions. We report the isolation of 47 insertions into 36 distinct MYB genes by screening a total of 73 genes. These defined insertion lines will provide the foundation for subsequent detailed functional analyses for the assignment of specific functions to individual members of the R2R3 MYB gene family.

The Promiscuous Life of Plant NUCLEAR FACTOR Y Transcription Factors

The CCAAT box is one of the most common cis-elements present in eukaryotic promoters and is bound by the transcription factor NUCLEAR FACTOR Y (NF-Y). NF-Y is composed of three subunits, NF-YA, NF-YB, and NF-YC. Unlike animals and fungi, plants have significantly expanded the number of genes encoding NF-Y subunits. We provide a comprehensive classification of NF-Y genes, with a separation of closely related, but distinct, histone fold domain proteins. We additionally review recent experiments that have placed NF-Y at the center of many developmental stress-responsive processes in the plant lineage.

How Can Research on Plants Contribute to Promoting Human Health

One of the most pressing challenges for the next 50 years is to reduce the impact of chronic disease. Unhealthy eating is an increasing problem and underlies much of the increase in mortality from chronic diseases that is occurring worldwide. Diets rich in plant-based foods are strongly associated with reduced risks of major chronic diseases, but the constituents in plants that promote health have proved difficult to identify with certainty. This, in turn, has confounded the precision of dietary recommendations. Plant biochemistry can make significant contributions to human health through the identification and measurement of the many metabolites in plant-based foods, particularly those known to promote health (phytonutrients). Plant genetics and metabolic engineering can be used to make foods that differ only in their content of specific phytonutrients. Such foods offer research tools that can provide significant insight into which metabolites promote health and how they work. Plant science can reduce some of the complexity of the diet-health relationship, and through building multidisciplinary interactions with researchers in nutrition and the pathology of chronic diseases, plant scientists can contribute novel insight into which foods reduce the risk of chronic disease and how these foods work to impact human health.

Plants, diet, and health.

Chronic disease is a major social challenge of the twenty-first century. In this review, we examine the evidence for discordance between modern diets and those on which humankind evolved as the cause of the increasing incidence of chronic diseases, and the evidence supporting consumption of plant foods as a way to reduce the risk of chronic disease. We also examine the evidence for avoiding certain components of plant-based foods that are enriched in Western diets, and review the mechanisms by which different phytonutrients are thought to reduce the risk of chronic disease. This body of evidence strongly suggests that consuming more fruits and vegetables could contribute both to medical nutrition therapies, as part of a package of treatments for conditions like type 2 diabetes, heart disease, cancer, and obesity, and to the prevention of these diseases. Plant science should be directed toward improving the quality of plant-based foods by building on our improved understanding of the complex relationships between plants, our diet, and our health.

Blood orange juice inhibits fat accumulation in mice

Objective:To analyze the effect of the juice obtained from two varieties of sweet orange (Citrus sinensis L. Osbeck), Moro (a blood orange) and Navelina (a blond orange), on fat accumulation in mice fed a standard or a high-fat diet (HFD).Methods:Obesity was induced in male C57/Bl6 mice by feeding a HFD. Moro and Navelina juices were provided instead of water. The effect of an anthocyanin-enriched extract from Moro oranges or purified cyanidin-3-glucoside (C3G) was also analyzed. Body weight and food intake were measured regularly over a 12-week period. The adipose pads were weighted and analyzed histologically; total RNA was also isolated for microarray analysis.Results:Dietary supplementation of Moro juice, but not Navelina juice significantly reduced body weight gain and fat accumulation regardless of the increased energy intake because of sugar content. Furthermore, mice drinking Moro juice were resistant to HFD-induced obesity with no alterations in food intake. Only the anthocyanin extract, but not the purified C3G, slightly affected fat accumulation. High-throughput gene expression analysis of fat tissues confirmed that Moro juice could entirely rescue the high fat-induced transcriptional reprogramming.Conclusion:Moro juice anti-obesity effect on fat accumulation cannot be explained only by its anthocyanin content. Our findings suggest that multiple components present in the Moro orange juice might act synergistically to inhibit fat accumulation.

The reduced expression of endogenous duplications (REED) in the maize R gene family is mediated by DNA methylation

The duplicated R and Sn genes regulate the maize anthocyanin biosynthetic pathway and encode tissue‐specific products that are homologous to helix‐loop‐helix transcriptional activators. As a consequence of their coupling in the genome, Sn is partially silenced. Genomic restriction analysis failed to reveal gross structural DNA alterations between the strong original phenotype and the weak derivatives. However, the differences in pigmentation were inversely correlated with differences in the methylation of the Sn promoter. Accordingly, treatment with 5‐azacytidine (AZA), a demethylating agent, restored a strong pigmentation pattern that was transmitted to the progeny and that was correlated with differential expression of the Sn transcript. Genomic sequencing confirmed that methylation of the Sn promoter was more apparent in the less pigmented seedlings and was greatly reduced in the AZA revertants. In addition, some methylcytosines were located in non‐symmetrical C sequences. These findings provide an insight into Sn and R interaction, a process that we have termed Reduced Expression of Endogenous Duplications (REED). We propose that increasing the copy number of regulatory genes by endogenous duplication leads to such epigenetic mechanisms of silencing. Further understanding of the REED process may have broader implications for gene regulation and may identify new levels of regulation within eukaryotic genomes.

Flavonoid profiling and biosynthetic gene expression in flesh and peel of two tomato genotypes grown under UV-B-depleted conditions during ripening.

The effect of shielding solar ultraviolet B radiation on the accumulation of some flavonoids and their precursor hydroxycinnamic acids in tomato (Solanum lycopersicum) was evaluated by liquid chromatography-electrospray ionization tandem mass spectrometry (LC-ESI-MS/MS). In particular, flesh and peel of two tomato hybrids, DRW 5981 and Esperanza, were separately analyzed. The hybrids have been chosen for their different responses to the light, since it was previously reported that they show different pigmentation and opposite behavior under UV-B in terms of carotenoids and ascorbic acid content at different ripening stages. To determine the effect of UV-B radiation during tomato ripening, we also measured the expression of some flavonoid biosynthetic genes by real-time reverse transcription-polymerase chain reaction (RT-PCR) analysis. The results allowed us to conclude that UV-B radiation deeply and differentially affects the content of the considered flavonoids and hydroxycinnamic acids as well as the expression of some of their biosynthetic genes in both flesh and peel during the ripening process. On the other hand, the collected data clearly showed that this influence varies between different genotypes. We conclude that the use of specific plastic covers able to eliminate UV-B radiation may be an environmentally friendly approach to modulate the expression of structural genes and, in turn, to enhance healthy antioxidant compounds in fruits of specific tomato cultivars.

Interactions and CCAAT-Binding of Arabidopsis thaliana NF-Y Subunits

Background NF-Y is a transcription factor that recognizes with high specificity and affinity the widespread CCAAT box promoter element. It is formed by three subunits: NF-YA and the NF-YB/NF-YC- heterodimer containing histone fold domains (HFDs). We previously identified a large NF-Y gene family in Arabidopsis thaliana, composed of 29 members, and characterized their expression patterns in various plant tissues. Methods We used yeast Two-hybrids assays (Y2H), pull-down and Electrophoretic Mobility Shift Assay (EMSA) in vitro experiments with recombinant proteins to dissect AtNF-YB/AtNF-YC interactions and DNA-binding with different AtNF-YAs. Results Consistent with robust conservation within HFDs, we show that heterodimerization is possible among all histone-like subunits, including the divergent and related LEC1/AtNF-YB9 and L1L/AtNF-YB6 required for embryo development. DNA-binding to a consensus CCAAT box was investigated with specific AtNF-YB/AtNF-YC combinations and observed with some, but not all AtNF-YA subunits. Conclusions Our results highlight (i) the conserved heterodimerization capacity of AtNF-Y histone-like subunits, and (ii) the different affinities of AtNF-YAs for the CCAAT sequence. Because of the general expansion of NF-Y genes in plants, these results most likely apply to other species.

Solar UV-B Radiation Influences Carotenoid Accumulation of Tomato Fruit through Both Ethylene-Dependent and - Independent Mechanisms

The effect of UV-B shielding on ethylene production in ripening tomato fruits and the contribution of ethylene and UV-B radiation on carotenoid accumulation and profile during ripening were assessed to get more insight about the interplay between these two regulatory factors. To this aim, rin and nor tomato mutants, unable to produce ripening ethylene, and cv Ailsa Craig were cultivated under control or UV-B depleted conditions until full fruit ripening. The significantly decreased ethylene evolution following UV-B depletion, evident only in Ailsa Craig, suggested the requirement of functional rin and nor genes for UVB-mediated ethylene production. Carotenoid content and profile were found to be controlled by both ethylene and UV-B radiation. This latter influenced carotenoid metabolism either in an ethylene-dependent or -independent way, as indicated by UVB-induced changes also in nor and rin carotenoid content and confirmed by correlation plots between ethylene evolution and carotenoid accumulation performed separately for control and UV-B shielded fruits. In conclusion, natural UV-B radiation influences carotenoid metabolism in a rather complex way, involving ethylene-dependent and -independent mechanisms, which seem to act in an antagonistic way.