Phenolic Compounds and Capsaicinoids in Three Capsicum annuum Varieties: From Analytical Characterization to In Silico Hypotheses on Biological Activity.

The affinity of specific phenolic compounds (PCs) and capsaicinoids (CAPs) present in three Capsicum annuum varieties (Friariello, Cayenne and Dzuljunska Sipka) to the transient receptor potential vanilloid member 1 (TRPV1) was investigated by integrating an analytic approach for the simultaneous extraction and analysis through high-performance liquid chromatography coupled with ion trap mass spectrometry (HPLC/ITMS) and UV detection (HPLC-UV) of PCs and CAPs and structural bioinformatics based on the protein modelling and molecular simulations of protein-ligand docking. Overall, a total of 35 compounds were identified in the different samples and CAPs were quantified. The highest content of total polyphenols was recorded in the pungent Dzuljunska Sipka variety (8.91 ± 0.05 gGAE/Kg DW) while the lowest was found in the non-pungent variety Friariello (3.58 ± 0.02 gGAE/Kg DW). Protein modelling generated for the first time a complete model of the homotetrameric human TRPV1, and it was used for docking simulations with the compounds detected via the analytic approach, as well as with other compounds, as an inhibitor reference. The simulations indicate that different capsaicinoids can interact with the receptor, providing details on the molecular interaction, with similar predicted binding energy values. These results offer new insights into the interaction of capsaicinoids with TRPV1 and their possible actions.

Complete Sequence, Multichromosomal Architecture and Transcriptome Analysis of the Solanum tuberosum Mitochondrial Genome.

Mitochondrial genomes (mitogenomes) in higher plants can induce cytoplasmic male sterility and be somehow involved in nuclear-cytoplasmic interactions affecting plant growth and agronomic performance. They are larger and more complex than in other eukaryotes, due to their recombinogenic nature. For most plants, the mitochondrial DNA (mtDNA) can be represented as a single circular chromosome, the so-called master molecule, which includes repeated sequences that recombine frequently, generating sub-genomic molecules in various proportions. Based on the relevance of the potato crop worldwide, herewith we report the complete mtDNA sequence of two S. tuberosum cultivars, namely Cicero and Désirée, and a comprehensive study of its expression, based on high-coverage RNA sequencing data. We found that the potato mitogenome has a multi-partite architecture, divided in at least three independent molecules that according to our data should behave as autonomous chromosomes. Inter-cultivar variability was null, while comparative analyses with other species of the Solanaceae family allowed the investigation of the evolutionary history of their mitogenomes. The RNA-seq data revealed peculiarities in transcriptional and post-transcriptional processing of mRNAs. These included co-transcription of genes with open reading frames that are probably expressed, methylation of an rRNA at a position that should impact translation efficiency and extensive RNA editing, with a high proportion of partial editing implying frequent mis-targeting by the editing machinery.

Distinct gene networks drive differential response to abrupt or gradual water deficit in potato.

Water-limiting conditions affect dramatically plant growth and development and, ultimately, yield of potato plants (Solanum tuberosum L.). Therefore, understanding the mechanisms underlying the response to water deficit is of paramount interest to obtain drought tolerant potato varieties. Herein, potato 10K cDNA array slides were used to profile transcriptomic changes of two potato cell populations under abrupt (shocked cells) or gradual exposure (adapted cells) to polyethylene glycol (PEG)-mediated water stress. Data analysis identified >1000 differentially expressed genes (DEGs) in our experimental conditions. Noteworthy, our microarray study also suggests that distinct gene networks underlie the cellular response to shock or gradual water stress. On the basis of our experimental findings, it is possible to speculate that DEGs identified in shocked cells participate in early protective and sensing mechanisms to environmental insults, while the genes whose expression was modulated in adapted cells are directly involved in the acquisition of a new cellular homeostasis to cope with water stress conditions. To validate microarray data obtained for potato cells, the expression analysis of 21 selected genes of interest was performed by Real-Time Quantitative Reverse Transcription PCR (qRT-PCR). Intriguingly, the expression levels of these transcripts in 4-week old potato plants exposed to long-term water-deficit. qRT-PCR analysis showed that several genes were regulated similarly in potato cells cultures and tissues exposed to drought, thus confirming the efficacy of our simple experimental system to capture important genes involved in osmotic stress response. Highlighting the differences in gene expression between shock-like and adaptive response, our findings could contribute to the discussion on the biological function of distinct gene networks involved in the response to abrupt and gradual adaptation to water deficit.

Transcriptome and metabolome of synthetic Solanum autotetraploids reveal key genomic stress events following polyploidization.

Polyploids are generally classified as autopolyploids, derived from a single species, and allopolyploids, arising from interspecific hybridization. The former represent ideal materials with which to study the consequences of genome doubling and ascertain whether there are molecular and functional rules operating following polyploidization events. To investigate whether the effects of autopolyploidization are common to different species, or if species-specific or stochastic events are prevalent, we performed a comprehensive transcriptomic and metabolomic characterization of diploids and autotetraploids of Solanum commersonii and Solanum bulbocastanum. Autopolyploidization remodelled the transcriptome and the metabolome of both species. In S. commersonii, differentially expressed genes (DEGs) were highly enriched in pericentromeric regions. Most changes were stochastic, suggesting a strong genotypic response. However, a set of robustly regulated transcripts and metabolites was also detected, including purine bases and nucleosides, which are likely to underlie a common response to polyploidization. We hypothesize that autopolyploidization results in nucleotide pool imbalance, which in turn triggers a genomic shock responsible for the stochastic events observed. The more extensive genomic stress and the higher number of stochastic events observed in S. commersonii with respect to S. bulbocastanum could be the result of the higher nucleoside depletion observed in this species.

Drought and flooding have distinct effects on herbivore-induced responses and resistance in Solanum dulcamara.

In the field, biotic and abiotic stresses frequently co-occur. As a consequence, common molecular signalling pathways governing adaptive responses to individual stresses can interact, resulting in compromised phenotypes. How plant signalling pathways interact under combined stresses is poorly understood. To assess this, we studied the consequence of drought and soil flooding on resistance of Solanum dulcamara to Spodoptera exigua and their effects on hormonal and transcriptomic profiles. The results showed that S. exigua larvae performed less well on drought-stressed plants than on well-watered and flooded plants. Both drought and insect feeding increased abscisic acid and jasmonic acid (JA) levels, whereas flooding did not induce JA accumulation. RNA sequencing analyses corroborated this pattern: drought and herbivory induced many biological processes that were repressed by flooding. When applied in combination, drought and herbivory had an additive effect on specific processes involved in secondary metabolism and defence responses, including protease inhibitor activity. In conclusion, drought and flooding have distinct effects on herbivore-induced responses and resistance. Especially, the interaction between abscisic acid and JA signalling may be important to optimize plant responses to combined drought and insect herbivory, making drought-stressed plants more resistant to insects than well-watered and flooded plants.

Genomic analysis of the native European Solanum species, S. dulcamara.

BACKGROUND: Solanum dulcamara (bittersweet, climbing nightshade) is one of the few species of the Solanaceae family native to Europe. As a common weed it is adapted to a wide range of ecological niches and it has long been recognized as one of the alternative hosts for pathogens and pests responsible for many important diseases in potato, such as Phytophthora. At the same time, it may represent an alternative source of resistance genes against these diseases. Despite its unique ecology and potential as a genetic resource, genomic research tools are lacking for S. dulcamara. We have taken advantage of next-generation sequencing to speed up research on and use of this non-model species. RESULTS: In this work, we present the first large-scale characterization of the S. dulcamara transcriptome. Through comparison of RNAseq reads from two different accessions, we were able to predict transcript-based SNP and SSR markers. Using the SNP markers in combination with genomic AFLP and CAPS markers, the first genome-wide genetic linkage map of bittersweet was generated. Based on gene orthology, the markers were anchored to the genome of related Solanum species (tomato, potato and eggplant), revealing both conserved and novel chromosomal rearrangements. This allowed a better estimation of the evolutionary moment of rearrangements in a number of cases and showed that chromosomal breakpoints are regularly re-used. CONCLUSION: Knowledge and tools developed as part of this study pave the way for future genomic research and exploitation of this wild Solanum species. The transcriptome assembly represents a resource for functional analysis of genes underlying interesting biological and agronomical traits and, in the absence of the full genome, provides a reference for RNAseq gene expression profiling aimed at understanding the unique biology of S. dulcamara. Cross-species orthology-based marker selection is shown to be a powerful tool to quickly generate a comparative genetic map, which may speed up gene mapping and contribute to the understanding of genome evolution within the Solanaceae family.

Revealing impaired pathways in the an11 mutant by high-throughput characterization of Petunia axillaris and Petunia inflata transcriptomes.

Petunia is an excellent model system, especially for genetic, physiological and molecular studies. Thus far, however, genome-wide expression analysis has been applied rarely because of the lack of sequence information. We applied next-generation sequencing to generate, through de novo read assembly, a large catalogue of transcripts for Petunia axillaris and Petunia inflata. On the basis of both transcriptomes, comprehensive microarray chips for gene expression analysis were established and used for the analysis of global- and organ-specific gene expression in Petunia axillaris and Petunia inflata and to explore the molecular basis of the seed coat defects in a Petunia hybrida mutant, anthocyanin 11 (an11), lacking a WD40-repeat (WDR) transcription regulator. Among the transcripts differentially expressed in an11 seeds compared with wild type, many expected targets of AN11 were found but also several interesting new candidates that might play a role in morphogenesis of the seed coat. Our results validate the combination of next-generation sequencing with microarray analyses strategies to identify the transcriptome of two petunia species without previous knowledge of their genome, and to develop comprehensive chips as useful tools for the analysis of gene expression in P. axillaris, P. inflata and P. hybrida.

An EST database from saffron stigmas.

BACKGROUND: Saffron (Crocus sativus L., Iridaceae) flowers have been used as a spice and medicinal plant ever since the Greek-Minoan civilization. The edible part - the stigmas - are commonly considered the most expensive spice in the world and are the site of a peculiar secondary metabolism, responsible for the characteristic color and flavor of saffron. RESULTS: We produced 6,603 high quality Expressed Sequence Tags (ESTs) from a saffron stigma cDNA library. This collection is accessible and searchable through the Saffron Genes database http://www.saffrongenes.org. The ESTs have been grouped into 1,893 Clusters, each corresponding to a different expressed gene, and annotated. The complete set of raw EST sequences, as well as of their electopherograms, are maintained in the database, allowing users to investigate sequence qualities and EST structural features (vector contamination, repeat regions). The saffron stigma transcriptome contains a series of interesting sequences (putative sex determination genes, lipid and carotenoid metabolism enzymes, transcription factors). CONCLUSION: The Saffron Genes database represents the first reference collection for the genomics of Iridaceae, for the molecular biology of stigma biogenesis, as well as for the metabolic pathways underlying saffron secondary metabolism.