PRGdb 4.0: an updated database dedicated to genes involved in plant disease resistance process.

The Plant Resistance Genes database (PRGdb; http://prgdb.org/prgdb4/) has been greatly expanded, keeping pace with the increasing amount of available knowledge and data (sequenced proteomes, cloned genes, public analysis data, etc.). The easy-to-use style of the database website has been maintained, while an updated prediction tool, more data and a new section have been added. This new section will contain plant resistance transcriptomic experiments, providing additional easy-to-access experimental information. DRAGO3, the tool for automatic annotation and prediction of plant resistance genes behind PRGdb, has been improved in both accuracy and sensitivity, leading to more reliable predictions. PRGdb offers 199 reference resistance genes and 586.652 putative resistance genes from 182 sequenced proteomes. Compared to the previous release, PRGdb 4.0 has increased the number of reference resistance genes from 153 to 199, the number of putative resistance genes from 177K from 76 proteomes to 586K from 182 sequenced proteomes. A new section has been created that collects plant-pathogen transcriptomic data for five species of agricultural interest. Thereby, with these improvements and data expansions, PRGdb 4.0 aims to serve as a reference to the plant scientific community and breeders worldwide, helping to further study plant resistance mechanisms that contribute to fighting pathogens.

The Impact of Growing Area on the Expression of Fruit Traits Related to Sensory Perception in Two Tomato Cultivars.

Environmental conditions greatly influence the quality of tomato fruit by affecting the expression of genes, the abundance of metabolites, and the perception of sensorial attributes. In this study, a fruit transcriptome investigation, a sensory test, and a metabolomic analysis were performed to evaluate the impact of the environment on two popular tomato cultivars grown in two Italian regions. The transcriptional profile of each cultivar, cultivated in two different areas, highlighted differential expression in genes involved in pathways related to cell wall components such as pectin, lignin, and hemicellulose and sugars as well as in amino acids, phenylpropanoids, and pigment synthesis. The cultivation area mainly affects sensory attributes related to texture and flavor and the metabolic pattern of cell wall precursors, sugars, glutamate, aspartate, and carotenoids. In the two genotypes cultivated in the same environment, some attributes and fruit-related quality processes are similarly affected, while others are differently influenced based on the specific genetic makeup of the tomato. A combination of transcriptomic, sensory, and metabolomic data obtained from the two tomato genotypes revealed that the environment has a profound effect on specific sensory traits, providing information on factors that shape the specific characteristics and genetic targets for improving tomato fruit characteristics.

Genomic and metabolic profiling of two tomato contrasting cultivars for tolerance to Tuta absoluta.

MAIN CONCLUSION: Dissimilar patterns of variants affecting genes involved in response to herbivory, including those leading to difference in VOC production, were identified in tomato lines with contrasting response to Tuta absoluta. Tuta absoluta is one of the most destructive insect pest affecting tomato production, causing important yield losses both in open field and greenhouse. The selection of tolerant varieties to T. absoluta is one of the sustainable approaches to control this invasive leafminer. In this study, the genomic diversity of two tomato varieties, one tolerant and the other susceptible to T. absoluta infestation was explored, allowing us to identify chromosome regions with highly dissimilar pattern. Genes affected by potential functional variants were involved in several processes, including response to herbivory and secondary metabolism. A metabolic analysis for volatile organic compounds (VOCs) was also performed, highlighting a difference in several classes of chemicals in the two genotypes. Taken together, these findings can aid tomato breeding programs aiming to develop tolerant plants to T. absoluta.

Comparison of Tomato Transcriptomic Profiles Reveals Overlapping Patterns in Abiotic and Biotic Stress Responses.

Until a few years ago, many studies focused on the transcriptomic response to single stresses. However, tomato cultivations are often constrained by a wide range of biotic and abiotic stress that can occur singularly or in combination, and several genes can be involved in the defensive mechanism response. Therefore, we analyzed and compared the transcriptomic responses of resistant and susceptible genotypes to seven biotic stresses (Cladosporium fulvum, Phytophthora infestans, Pseudomonas syringae, Ralstonia solanacearum, Sclerotinia sclerotiorum, Tomato spotted wilt virus (TSWV) and Tuta absoluta) and five abiotic stresses (drought, salinity, low temperatures, and oxidative stress) to identify genes involved in response to multiple stressors. With this approach, we found genes encoding for TFs, phytohormones, or participating in signaling and cell wall metabolic processes, participating in defense against various biotic and abiotic stress. Moreover, a total of 1474 DEGs were commonly found between biotic and abiotic stress. Among these, 67 DEGs were involved in response to at least four different stresses. In particular, we found RLKs, MAPKs, Fasciclin-like arabinogalactans (FLAs), glycosyltransferases, genes involved in the auxin, ET, and JA pathways, MYBs, bZIPs, WRKYs and ERFs genes. Detected genes responsive to multiple stress might be further investigated with biotechnological approaches to effectively improve plant tolerance in the field.

NB-LRR Lineage-Specific Equipment Is Sorted Out by Sequence Pattern Adaptation and Domain Segment Shuffling.

The nucleotide-binding and leucine-rich repeat (NB-LRR) genes, also known as resistance (R)-genes, play an important role in the activation of immune responses. In recent years, large-scale studies have been performed to highlight the diversification of plant NB-LRR repertories. It is well known that, to provide new functionalities, NB-LRR sequences are subject to duplication, domain fusions and acquisition and other kinds of mutations. Although some mechanisms that govern NB-LRR protein domain adaptations have been uncovered, to retrace the plant-lineage-specific evolution routes of R protein structure, a multi-genome comparative analysis was performed. This study allowed us to define groups of genes sharing homology relationships across different species. It is worth noting that the most populated groups contained well-characterized R proteins. The arsenal profile of such groups was investigated in five botanical families, including important crop species, to underline specific adaptation signatures. In addition, the dissection of 70 NB domains of well-characterized R-genes revealed the NB core motifs from which the three main R protein classes have been diversified. The structural remodeling of domain segments shaped the specific NB-LRR repertoires observed in each plant species. This analysis provided new evolutionary and functional insights on NB protein domain shuffling. Taken together, such findings improved our understanding of the molecular adaptive selection mechanisms occurring at plant R loci.

Tomato transcriptomic response to Tuta absoluta infestation.

BACKGROUND: The South America pinworm, Tuta absoluta, is a destructive pest of tomato that causes important losses worldwide. Breeding of resistant/tolerant tomato cultivars could be an effective strategy for T. absoluta management but, despite the economic importance of tomato, very limited information is available about its response to this treat. To elucidate the defense mechanisms to herbivore feeding a comparative analysis was performed between a tolerant and susceptible cultivated tomato at both morphological and transcriptome level to highlight constitutive leaf barriers, molecular and biochemical mechanisms to counter the effect of T. absoluta attack. RESULTS: The tolerant genotype showed an enhanced constitutive barrier possibly as result of the higher density of trichomes and increased inducible reactions upon mild infestation thanks to the activation/repression of key transcription factors regulating genes involved in cuticle formation and cell wall strength as well as of antinutritive enzymes, and genes involved in the production of chemical toxins and bioactive secondary metabolites. CONCLUSIONS: Overall, our findings suggest that tomato resilience to the South America pinworm is achieved by a combined strategy between constitutive and induced defense system. A well-orchestrated modulation of plant transcription regulation could ensure a trade-off between defense needs and fitness costs. Our finding can be further exploited for developing T. absoluta tolerant cultivars, acting as important component of integrated pest management strategy for more sustainable production.

Evolutionary conservation of MLO gene promoter signatures.

BACKGROUND: Powdery mildew (PM) is a widespread fungal disease of plants in temperate climates, causing significant economic losses in agricultural settings. Specific homologs of the MLO gene family are PM susceptibility factors, as their loss-of function results in durable PM resistance (mlo resistance) in several plant species. The role of MLO susceptibility genes in plant-pathogen interactions is still elusive, however it is known that they are strongly upregulated following PM infection. RESULTS: In this study, we investigated the structure of 414 Putative Promoter Regions (PPRs) of MLO genes and highlighted motif and regulatory element patterns related to genomic relationships among species and phylogenetic distance among homologs. A TC box-like motif and a thymine-rich motif were found to be overrepresented in MLO genes transcriptionally upregulated upon infection with PM fungi. As proof of concept, we showed that the expression of a melon (Cucumis melo L.) gene enriched for the motifs above mentioned was strongly upregulated upon infection with the PM fungus Podosphaera xanthii. CONCLUSION: While identifying a candidate MLO susceptibility gene in melon, this study provides insight on the transcriptional control of MLO genes and indicates diagnostic features useful to identify MLO susceptibility genes across species affected by the PM disease.

The Solanum commersonii Genome Sequence Provides Insights into Adaptation to Stress Conditions and Genome Evolution of Wild Potato Relatives.

Here, we report the draft genome sequence of Solanum commersonii, which consists of ∼830 megabases with an N50 of 44,303 bp anchored to 12 chromosomes, using the potato (Solanum tuberosum) genome sequence as a reference. Compared with potato, S. commersonii shows a striking reduction in heterozygosity (1.5% versus 53 to 59%), and differences in genome sizes were mainly due to variations in intergenic sequence length. Gene annotation by ab initio prediction supported by RNA-seq data produced a catalog of 1703 predicted microRNAs, 18,882 long noncoding RNAs of which 20% are shown to target cold-responsive genes, and 39,290 protein-coding genes with a significant repertoire of nonredundant nucleotide binding site-encoding genes and 126 cold-related genes that are lacking in S. tuberosum. Phylogenetic analyses indicate that domesticated potato and S. commersonii lineages diverged ∼2.3 million years ago. Three duplication periods corresponding to genome enrichment for particular gene families related to response to salt stress, water transport, growth, and defense response were discovered. The draft genome sequence of S. commersonii substantially increases our understanding of the domesticated germplasm, facilitating translation of acquired knowledge into advances in crop stability in light of global climate and environmental changes.

Unraveling the complexity of transcriptomic, metabolomic and quality environmental response of tomato fruit.

BACKGROUND: The environment has a profound influence on the organoleptic quality of tomato (Solanum lycopersicum) fruit, the extent of which depends on a well-regulated and dynamic interplay among genes, metabolites and sensorial attributes. We used a systems biology approach to elucidate the complex interacting mechanisms regulating the plasticity of sensorial traits. To investigate environmentally challenged transcriptomic and metabolomic remodeling and evaluate the organoleptic consequences of such variations we grown three tomato varieties, Heinz 1706, whose genome was sequenced as reference and two "local" ones, San Marzano and Vesuviano in two different locations of Campania region (Italy). RESULTS: Responses to environment were more pronounced in the two "local" genotypes, rather than in the Heinz 1706. The overall genetic composition of each genotype, acting in trans, modulated the specific response to environment. Duplicated genes and transcription factors, establishing different number of network connections by gaining or losing links, play a dominant role in shaping organoleptic profile. The fundamental role of cell wall metabolism in tuning all the quality attributes, including the sensorial perception, was also highlighted. CONCLUSIONS: Although similar fruit-related quality processes are activated in the same environment, different tomato genotypes follow distinct transcriptomic, metabolomic and sensorial trajectories depending on their own genetic makeup.

Fusarium oxysporum f.sp. radicis-lycopersici induces distinct transcriptome reprogramming in resistant and susceptible isogenic tomato lines.

BACKGROUND: Fusarium oxysporum f.sp. radicis-lycopersici (FORL) is one of the most destructive necrotrophic pathogens affecting tomato crops, causing considerable field and greenhouse yield losses. Despite such major economic impact, little is known about the molecular mechanisms regulating Fusarium oxysporum f.sp. radicis-lycopersici resistance in tomato. RESULTS: A transcriptomic experiment was carried out in order to investigate the main mechanisms of FORL response in resistant and susceptible isogenic tomato lines. Microarray analysis at 15 DPI (days post inoculum) revealed a distinct gene expression pattern between the two genotypes in the inoculated vs non-inoculated conditions. A model of plant response both for compatible and incompatible reactions was proposed. In particular, in the incompatible interaction an activation of defense genes related to secondary metabolite production and tryptophan metabolism was observed. Moreover, maintenance of the cell osmotic potential after the FORL challenging was mediated by a dehydration-induced protein. As for the compatible interaction, activation of an oxidative burst mediated by peroxidases and a cytochrome monooxygenase induced cell degeneration and necrosis. CONCLUSIONS: Our work allowed comprehensive understanding of the molecular basis of the tomato-FORL interaction. The result obtained emphasizes a different transcriptional reaction between the resistant and the susceptible genotype to the FORL challenge. Our findings could lead to the improvement in disease control strategies.

Structure, evolution and functional inference on the Mildew Locus O (MLO) gene family in three cultivated Cucurbitaceae spp.

BACKGROUND: The powdery mildew disease affects thousands of plant species and arguably represents the major fungal threat for many Cucurbitaceae crops, including melon (Cucumis melo L.), watermelon (Citrullus lanatus L.) and zucchini (Cucurbita pepo L.). Several studies revealed that specific members of the Mildew Locus O (MLO) gene family act as powdery mildew susceptibility factors. Indeed, their inactivation, as the result of gene knock-out or knock-down, is associated with a peculiar form of resistance, referred to as mlo resistance. RESULTS: We exploited recently available genomic information to provide a comprehensive overview of the MLO gene family in Cucurbitaceae. We report the identification of 16 MLO homologs in C. melo, 14 in C. lanatus and 18 in C. pepo genomes. Bioinformatic treatment of data allowed phylogenetic inference and the prediction of several ortholog pairs and groups. Comparison with functionally characterized MLO genes and, in C. lanatus, gene expression analysis, resulted in the detection of candidate powdery mildew susceptibility factors. We identified a series of conserved amino acid residues and motifs that are likely to play a major role for the function of MLO proteins. Finally, we performed a codon-based evolutionary analysis indicating a general high level of purifying selection in the three Cucurbitaceae MLO gene families, and the occurrence of regions under diversifying selection in candidate susceptibility factors. CONCLUSIONS: Results of this study may help to address further biological questions concerning the evolution and function of MLO genes. Moreover, data reported here could be conveniently used by breeding research, aiming to select powdery mildew resistant cultivars in Cucurbitaceae.

Genetic variability and evolutionary diversification of membrane ABC transporters in plants.

BACKGROUND: ATP-binding cassette proteins have been recognized as playing a crucial role in the regulation of growth and resistance processes in all kingdoms of life. They have been deeply studied in vertebrates because of their role in drug resistance, but much less is known about ABC superfamily functions in plants. RESULTS: Recently released plant genome sequences allowed us to identify 803 ABC transporters in four vascular plants (Oryza. sativa, Solanum lycopersicum, Solanum tuberosum and Vitis vinifera) and 76 transporters in the green alga Volvox carteri, by comparing them with those reannotated in Arabidopsis thaliana and the yeast Saccharomyces cerevisiae. Retrieved proteins have been phylogenetically analysed to infer orthologous relationships. Most orthologous relationships in the A, D, E and F subfamilies were found, and interesting expansions within the ABCG subfamily were observed and discussed. A high level of purifying selection is acting in the five ABC subfamilies A, B, C, D and E. However, evolutionary rates of recent duplicate genes could influence vascular plant genome diversification. The transcription profiles of ABC genes within tomato organs revealed a broad functional role for some transporters and a more specific activity for others, suggesting the presence of key ABC regulators in tomato. CONCLUSIONS: The findings achieved in this work could contribute to address several biological questions concerning the evolution of the relationship between genomes of different species. Plant ABC protein inventories obtained could be a valuable tool both for basic and applied studies. Indeed, interpolation of the putative role of gene functions can accelerate the discovering of new ABC superfamily members.

PRGdb 2.0: towards a community-based database model for the analysis of R-genes in plants.

The Plant Resistance Genes database (PRGdb; http://prgdb.org) is a comprehensive resource on resistance genes (R-genes), a major class of genes in plant genomes that convey disease resistance against pathogens. Initiated in 2009, the database has grown more than 6-fold to recently include annotation derived from recent plant genome sequencing projects. Release 2.0 currently hosts useful biological information on a set of 112 known and 104 310 putative R-genes present in 233 plant species and conferring resistance to 122 different pathogens. Moreover, the website has been completely redesigned with the implementation of Semantic MediaWiki technologies, which makes our repository freely accessed and easily edited by any scientists. To this purpose, we encourage plant biologist experts to join our annotation effort and share their knowledge on resistance-gene biology with the rest of the scientific community.

Patchwork sequencing of tomato San Marzano and Vesuviano varieties highlights genome-wide variations.

BACKGROUND: Investigation of tomato genetic resources is a crucial issue for better straight evolution and genetic studies as well as tomato breeding strategies. Traditional Vesuviano and San Marzano varieties grown in Campania region (Southern Italy) are famous for their remarkable fruit quality. Owing to their economic and social importance is crucial to understand the genetic basis of their unique traits. RESULTS: Here, we present the draft genome sequences of tomato Vesuviano and San Marzano genome. A 40x genome coverage was obtained from a hybrid Illumina paired-end reads assembling that combines de novo assembly with iterative mapping to the reference S. lycopersicum genome (SL2.40). Insertions, deletions and SNP variants were carefully measured. When assessed on the basis of the reference annotation, 30% of protein-coding genes are predicted to have variants in both varieties. Copy genes number and gene location were assessed by mRNA transcripts mapping, showing a closer relationship of San Marzano with reference genome. Distinctive variations in key genes and transcription/regulation factors related to fruit quality have been revealed for both cultivars. CONCLUSIONS: The effort performed highlighted varieties relationships and important variants in fruit key processes useful to dissect the path from sequence variant to phenotype.

PRGdb: a bioinformatics platform for plant resistance gene analysis.

PRGdb is a web accessible open-source (http://www.prgdb.org) database that represents the first bioinformatic resource providing a comprehensive overview of resistance genes (R-genes) in plants. PRGdb holds more than 16,000 known and putative R-genes belonging to 192 plant species challenged by 115 different pathogens and linked with useful biological information. The complete database includes a set of 73 manually curated reference R-genes, 6308 putative R-genes collected from NCBI and 10463 computationally predicted putative R-genes. Thanks to a user-friendly interface, data can be examined using different query tools. A home-made prediction pipeline called Disease Resistance Analysis and Gene Orthology (DRAGO), based on reference R-gene sequence data, was developed to search for plant resistance genes in public datasets such as Unigene and Genbank. New putative R-gene classes containing unknown domain combinations were discovered and characterized. The development of the PRG platform represents an important starting point to conduct various experimental tasks. The inferred cross-link between genomic and phenotypic information allows access to a large body of information to find answers to several biological questions. The database structure also permits easy integration with other data types and opens up prospects for future implementations.

Molecular tools for exploring polyploid genomes in plants.

Polyploidy is a very common phenomenon in the plant kingdom, where even diploid species are often described as paleopolyploids. The polyploid condition may bring about several advantages compared to the diploid state. Polyploids often show phenotypes that are not present in their diploid progenitors or exceed the range of the contributing species. Some of these traits may play a role in heterosis or could favor adaptation to new ecological niches. Advances in genomics and sequencing technology may create unprecedented opportunities for discovering and monitoring the molecular effects of polyploidization. Through this review, we provide an overview of technologies and strategies that may allow an in-depth analysis of polyploid genomes. After introducing some basic aspects on the origin and genetics of polyploids, we highlight the main tools available for genome and gene expression analysis and summarize major findings. In the last part of this review, the implications of next generation sequencing are briefly discussed. The accumulation of knowledge on polyploid formation, maintenance, and divergence at whole-genome and subgenome levels will not only help plant biologists to understand how plants have evolved and diversified, but also assist plant breeders in designing new strategies for crop improvement.

Multilevel evolution shapes the function of NB-LRR encoding genes in plant innate immunity.

A sophisticated innate immune system based on diverse pathogen receptor genes (PRGs) evolved in the history of plant life. To reconstruct the direction and magnitude of evolutionary trajectories of a given gene family, it is critical to detect the ancestral signatures. The rearrangement of functional domains made up the diversification found in PRG repertoires. Structural rearrangement of ancient domains mediated the NB-LRR evolutionary path from an initial set of modular proteins. Events such as domain acquisition, sequence modification and temporary or stable associations are prominent among rapidly evolving innate immune receptors. Over time PRGs are continuously shaped by different forces to find their optimal arrangement along the genome. The immune system is controlled by a robust regulatory system that works at different scales. It is important to understand how the PRG interaction network can be adjusted to meet specific needs. The high plasticity of the innate immune system is based on a sophisticated functional architecture and multi-level control. Due to the complexity of interacting with diverse pathogens, multiple defense lines have been organized into interconnected groups. Genomic architecture, gene expression regulation and functional arrangement of PRGs allow the deployment of an appropriate innate immunity response.

The Tomato Interspecific NB-LRR Gene Arsenal and Its Impact on Breeding Strategies.

Tomato (Solanum lycopersicum L.) is a model system for studying the molecular basis of resistance in plants. The investigation of evolutionary dynamics of tomato resistance (R)-loci provides unique opportunities for identifying factors that promote or constrain genome evolution. Nucleotide-binding domain and leucine-rich repeat (NB-LRR) receptors belong to one of the most plastic and diversified families. The vast amount of genomic data available for Solanaceae and wild tomato relatives provides unprecedented insights into the patterns and mechanisms of evolution of NB-LRR genes. Comparative analysis remarked a reshuffling of R-islands on chromosomes and a high degree of adaptive diversification in key R-loci induced by species-specific pathogen pressure. Unveiling NB-LRR natural variation in tomato and in other Solanaceae species offers the opportunity to effectively exploit genetic diversity in genomic-driven breeding programs with the aim of identifying and introducing new resistances in tomato cultivars. Within this motivating context, we reviewed the repertoire of NB-LRR genes available for tomato improvement with a special focus on signatures of adaptive processes. This issue is still relevant and not thoroughly investigated. We believe that the discovery of mechanisms involved in the generation of a gene with new resistance functions will bring great benefits to future breeding strategies.

Tomato genomic prediction for good performance under high-temperature and identification of loci involved in thermotolerance response.

Many studies showed that few degrees above tomato optimum growth temperature threshold can lead to serious loss in production. Therefore, the development of innovative strategies to obtain tomato cultivars with improved yield under high temperature conditions is a main goal both for basic genetic studies and breeding activities. In this paper, a F4 segregating population was phenotypically evaluated for quantitative and qualitative traits under heat stress conditions. Moreover, a genotyping by sequencing (GBS) approach has been employed for building up genomic selection (GS) models both for yield and soluble solid content (SCC). Several parameters, including training population size, composition and marker quality were tested to predict genotype performance under heat stress conditions. A good prediction accuracy for the two analyzed traits (0.729 for yield production and 0.715 for SCC) was obtained. The predicted models improved the genetic gain of selection in the next breeding cycles, suggesting that GS approach is a promising strategy to accelerate breeding for heat tolerance in tomato. Finally, the annotation of SNPs located in gene body regions combined with QTL analysis allowed the identification of five candidates putatively involved in high temperatures response, and the building up of a GS model based on calibrated panel of SNP markers.

Accelerating Tomato Breeding by Exploiting Genomic Selection Approaches.

Genomic selection (GS) is a predictive approach that was built up to increase the rate of genetic gain per unit of time and reduce the generation interval by utilizing genome-wide markers in breeding programs. It has emerged as a valuable method for improving complex traits that are controlled by many genes with small effects. GS enables the prediction of the breeding value of candidate genotypes for selection. In this work, we address important issues related to GS and its implementation in the plant context with special emphasis on tomato breeding. Genomic constraints and critical parameters affecting the accuracy of prediction such as the number of markers, statistical model, phenotyping and complexity of trait, training population size and composition should be carefully evaluated. The comparison of GS approaches for facilitating the selection of tomato superior genotypes during breeding programs is also discussed. GS applied to tomato breeding has already been shown to be feasible. We illustrated how GS can improve the rate of gain in elite line selection, and descendent and backcross schemes. The GS schemes have begun to be delineated and computer science can provide support for future selection strategies. A new promising breeding framework is beginning to emerge for optimizing tomato improvement procedures.