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High temperature stress is one of the major environmental factors that affect the growth and development of plants. Although WRKY transcription factors play a critical role in stress responses, there are few studies on the regulation of heat stress by WRKY transcription factors, especially in tomato. Here, we identified a group I WRKY transcription factor, SlWRKY3, involved in thermotolerance in tomato. First, SlWRKY3 was induced and upregulated under heat stress. Accordingly, overexpression of SlWRKY3 led to an increase, whereas knock-out of SlWRKY3 resulted in decreased tolerance to heat stress. Overexpression of SlWRKY3 accumulated less reactive oxygen species (ROS), whereas knock-out of SlWRKY3 accumulated more ROS under heat stress. This indicated that SlWRKY3 positively regulates heat stress in tomato. In addition, SlWRKY3 activated the expression of a range of abiotic stress-responsive genes involved in ROS scavenging, such as a SlGRXS1 gene cluster. Further analysis showed that SlWRKY3 can bind to the promoters of the SlGRXS1 gene cluster and activate their expression. Collectively, these results imply that SlWRKY3 is a positive regulator of thermotolerance through direct binding to the promoters of the SlGRXS1 gene cluster and activating their expression and ROS scavenging. [ABSTRACT FROM AUTHOR]

GRAS proteins are plant-specific transcription factors that play crucial roles in plant development and stress responses. However, their involvement in the ripening of economically important fruits and their transcriptional regulatory mechanisms remain largely unclear. Here, we demonstrated that SlGRAS4, encoding a transcription factor of the GRAS family, was induced by the tomato ripening process and regulated by ethylene. Overexpression of SlGRAS4 accelerated fruit ripening, increased the total carotenoid content and increased PSY1 expression in SlGRAS4-OE fruit compared to wild-type fruit. The expression levels of key ethylene biosynthesis genes (SlACS2, SlACS4, SlACO1, and SlACO3) and crucial ripening regulators (RIN and NOR) were increased in SlGRAS4-OE fruit. The negative regulator of tomato fruit ripening, SlMADS1, was repressed in OE fruit. Exogenous ethylene and 1-MCP treatment revealed that more endogenous ethylene was derived in SlGRAS4-OE fruit. More obvious phenotypes were observed in OE seedlings after ACC treatment. Yeast one-hybrid and dual-luciferase assays confirmed that SlGRAS4 can directly bind SlACO1 and SlACO3 promoters to activate their transcription, and SlGRAS4 can also directly repress SlMADS1 expression. Our study identified that SlGRAS4 acts as a new regulator of fruit ripening by regulating ethylene biosynthesis genes in a direct manner. This provides new knowledge of GRAS transcription factors involved in regulating fruit ripening. [ABSTRACT FROM AUTHOR]

Summary: Carotenoids are critically important to plants and humans. The ORANGE (OR) gene is a key regulator for carotenoid accumulation, but its physiological roles in crops remain elusive. In this study, we generated transgenic tomato ectopically overexpressing the Arabidopsis wild‐type OR (AtORWT) and a 'golden SNP'‐containing OR (AtORHis). We found that AtORHis initiated chromoplast formation in very young fruit and stimulated carotenoid accumulation at all fruit developmental stages, uncoupled from other ripening activities. The elevated levels of carotenoids in the AtOR lines were distributed in the same subplastidial fractions as in wild‐type tomato, indicating an adaptive response of plastids to sequester the increased carotenoids. Microscopic analysis revealed that the plastid sizes were increased in both AtORWT and AtORHis lines at early fruit developmental stages. Moreover, AtOR overexpression promoted early flowering, fruit set and seed production. Ethylene production and the expression of ripening‐associated genes were also significantly increased in the AtOR transgenic fruit at ripening stages. RNA‐Seq transcriptomic profiling highlighted the primary effects of OR overexpression on the genes in the processes related to RNA, protein and signalling in tomato fruit. Taken together, these results expand our understanding of OR in mediating carotenoid accumulation in plants and suggest additional roles of OR in affecting plastid size as well as flower and fruit development, thus making OR a target gene not only for nutritional biofortification of agricultural products but also for alteration of horticultural traits. [ABSTRACT FROM AUTHOR]

The effect of hypoxia on the antioxidant system during the storage of tomato fruits was investigated using plants with different expression levels of the encoding gene for the MT-sHSP23.6 protein. Fruits from two 'Micro-Tom' tomato plant genotypes (wild type and high expression of MT-sHSP23.6) were harvested at the breaker stage and subjected to normoxia (23 °C, dark) and hypoxia (daily nitrogen flow of 0.098 Mpa/10 min, 23 °C, darkness) treatments for 5 and 8 days, respectively. We evaluated the color tone (Hueº) of the fruits, as well as the components of the enzymatic antioxidant system, reactive oxygen species, non-enzymatic antioxidant system components and radical scavenging activity. Fruits from plants with high MT-sHSP23.6 expression had the highest activity for all the antioxidant enzymes under hypoxia conditions (25, 20, 76, and 48% higher than wild type for SOD, CAT, APX, and GPOD, respectively). They also had the lowest concentrations of superoxide anion under both normoxia and hypoxia storage. At the end of the normoxia storage period, the genotype with high MT-sHSP23.6 expression had the highest accumulation of lycopene (13%), whereas at the end of hypoxia storage, the wild type genotype had the highest lycopene accumulation (39%). Higher accumulation of phenolic compounds was observed in the post-hypoxia period for plants subjected to the hypoxia treatment in high MT-sHSP23.6 expression genotype (12% on the fourth day and 9% on the eighth day). In addition, ascorbic acid concentration was considerably higher on the third and fourth storage days (47% and 8%, respectively). During the hypoxia period, the genotype with high MT-sHSP23.6 expression exhibited the highest radical scavenging activity. High MT-sHSP23.6 expression stimulated the enzymatic antioxidant system during the hypoxia period and led to higher accumulation of phenolic compounds and ascorbic acid, evidence that this protein may be related to tolerance mechanisms in plants subjected to post-harvest abiotic stresses. [ABSTRACT FROM AUTHOR]

Summary: High‐throughput RNA sequencing has proven invaluable not only to explore gene expression but also for both gene prediction and genome annotation. However, RNA sequencing, carried out on tens or even hundreds of samples, requires easy and cost‐effective sample preparation methods using minute RNA amounts. Here, we present TranSeq, a high‐throughput 3′‐end sequencing procedure that requires 10‐ to 20‐fold fewer sequence reads than the current transcriptomics procedures. TranSeq significantly reduces costs and allows a greater increase in size of sample sets analyzed in a single experiment. Moreover, in comparison with other 3′‐end sequencing methods reported to date, we demonstrate here the reliability and immediate applicability of TranSeq and show that it not only provides accurate transcriptome profiles but also produces precise expression measurements of specific gene family members possessing high sequence similarity. This is difficult to achieve in standard RNA‐seq methods, in which sequence reads cover the entire transcript. Furthermore, mapping TranSeq reads to the reference tomato genome facilitated the annotation of new transcripts improving >45% of the existing gene models. Hence, we anticipate that using TranSeq will boost large‐scale transcriptome assays and increase the spatial and temporal resolution of gene expression data, in both model and non‐model plant species. Moreover, as already performed for tomato (ITAG3.0; www.solgenomics.net), we strongly advocate its integration into current and future genome annotations. [ABSTRACT FROM AUTHOR]

Ethylene plays an important role in the grape rachis, where its production can be 10 times higher than in the berry. VvACS1 is the only rachis-specific ACC synthase (ACS) gene, and its expression is coincident with ethylene production in the rachis of Vitis vinifera ‘Thompson seedless’. VvACS1 was cloned and ectopically expressed in tomato ( Solanum lycopersicum ‘Moneymaker’). Lateral buds were increased in two- or four-week-old 35s∷ VvACS1 transgenic tomato plants after transplanting. Compared with wild-type (WT) plants, the transgenic tomato plants showed higher expression of the VvACS1 gene in the flowers, leaves, rachis, and fruits. There was no obvious difference of ACS activity in the fruit of tomato, and only increased ACS activity in the rachis of tomato. Ethylene production was decreased in flowers, leaves, and fruits (seven weeks after full bloom), while the relative expression of endogenous tomato ACS1 and ACS6 genes was not down-regulated by the ectopic expression of VvACS1 . These results imply that post-transcriptional or post-translational regulation of ACS may occur, resulting in lower ethylene production in the transgenic tomato plants. Moreover, expression of VvACS1 in tomato resulted in decreased auxin and increased zeatin contents in the lateral buds, as well as reduced or delayed formation of adventitious roots in lateral bud cuttings. RNA-Seq and qRT-PCR analyses of rooted lateral bud cuttings indicated that the relative expression levels of the genes for zeatin O -glucosyltransferase-like, auxin repressed/dormancy-associated protein, and ERF transcription factors were higher in transgenic tomatoes than in WT, suggesting that ethylene may regulate auxin transport and distribution in shoots and that adventitious root formation employs coordination between auxin and ethylene. [ABSTRACT FROM AUTHOR]

Summary: Steroidal glycoalkaloids (SGAs) are specialized anti‐nutritional metabolites that accumulate in Solanum lycopersicum (tomato) and Solanum tuberosum (potato). A series of SGA biosynthetic genes is known to be upregulated in Solanaceae species by jasmonate‐responsive Ethylene Response Factor transcription factors, including JRE4 (otherwise known as GAME9), but the exact regulatory significance in planta of each factor has remained unaddressed. Here, via TILLING‐based screening of an EMS‐mutagenized tomato population, we isolated a JRE4 loss‐of‐function line that carries an amino acid residue missense change in a region of the protein important for DNA binding. In this jre4 mutant, we observed downregulated expression of SGA biosynthetic genes and decreased SGA accumulation. Moreover, JRE4 overexpression stimulated SGA production. Further characterization of jre4 plants revealed their increased susceptibility to the generalist herbivore Spodoptera litura larvae. This susceptibility illustrates that herbivory resistance is dependent on JRE4‐mediated defense responses, which include SGA accumulation. Ethylene treatment attenuated the jasmonate‐mediated JRE4 expression induction and downstream SGA biosynthesis in tomato leaves and hairy roots. Overall, this study indicated that JRE4 functions as a primary master regulator of SGA biosynthesis, and thereby contributes toward plant defense against chewing insects. [ABSTRACT FROM AUTHOR]

Circular RNAs (circRNAs) are an emerging class of non-coding RNAs in plants. Our aim is to identify the circRNAs with different expression levels between tomato fruits from two cultivars (‘Jinling Fenyu’ and ‘Jinling Moyu’) at two different developmental stages. Following high-throughput sequencing, 3796 circRNAs were identified, and 243 circRNAs were shared in the four samples. As compared with the fruit at mature green stage, the expression levels of 273 and 89 circRNAs were significantly altered in the fruit at turning stage from ‘Jinling Fenyu’ and ‘Jinling Moyu’, respectively. Moreover, the parental genes of the circRNAs with significantly different expression level were mainly involved in metabolic, cellular and single-organism process and played roles in catalytic activity and binding based on GO (Gene Ontology) analysis. The results suggested that circRNAs were widespread in tomato and were generated from different chromosomes and diverse genomic regions. Some circRNAs were specific in tomato fruits at different developmental stages, which enriches the number of circRNAs in plants involved in fruit coloration and ripening. This study provides the first genome-wide profile of circRNAs involved in tomato fruit coloration and lays a foundation for studying the potential biological functions of circRNAs involved in fruit ripening. [ABSTRACT FROM AUTHOR]

The ripening stage at harvest time determines the tomato fruit quality. After the fruit achieves its maximum size several metabolic changes of typically climacteric fruits are produced. Two cultivated genotypes of Solanum lycopersicum (Caimanta and 804627), with normal and altered fruit ripening, respectively and two accession, LA1385 of S. lycopersicum var. cerasiforme and LA722 of S. pimpinellifolium , with genes that prolong fruit shelf life, were tested to: 1) characterize and make a comparatively analysis for the transcriptome at different fruit ripening stages in genotypes that differ in fruit shelf life by cDNA-AFLP; and 2) provide further insight into the relationship between the extreme phenotypic differences for ripening among the genotypes through changes at transcriptomic level. Fruits at the breaker stage (B) were evaluated for fruit weight, firmness and fruit shelf life. The elapsed days between mature green (MG) and breaker stages Days (MG-B) as well as the elapsed days between B and red ripe (RR) stages Days (B-RR) were recorded. Comparison among ripening stages showed a great polymorphism related to the changes in gene expression. For all genotypes the transition from B to RR stages had higher polymorphism than the transition from MG to B. It was observed a great genetic variability for the phenotypic traits in agreement with the changes of gene expression. Moreover, it was observed that the transcriptome expression profiles in the initial and intermediate stages during ripening (MG and B) are more important to characterize genotypes. The wild species which have long shelf life do not show as drastic changes in gene expression as the cultivar with altered ripening that carrythe nor gene. These results suggest that the expressed or silenced genes could be involved, in some way, in the determination of the phenotypic traits evaluated in this study. [ABSTRACT FROM AUTHOR]

Abstract: Reliable reference genes are critical for relative quantification using quantitative real‐time PCR (qPCR). Ten tomato genes (Solanum lycopersicum) and their respective primer sets, which have been used over the last 6 years as references in expression studies, were evaluated for their performance using leaf tissue samples grown under semi‐controlled conditions and infected with grey mould (Botrytis cinerea) or late blight (Phytophthora infestans). The target genes coding for U6 snRNA‐associated Sm‐like protein LSm7, calcineurin B‐like protein and V‐type proton ATPase were the most stable expressed of all the genes tested in three experimental repetitions. Evaluation of candidate reference genes with geNorm and NormFinder softwares yielded the lowest mean values for their respective primer sets LSM7, SlCBL1 and SlATPase, suggesting stable expression. However, SlATPase primer set revealed a comparably high intra‐group variation and was thus not considered further. In follow‐up experiments with P. infestans, the geNorm and NormFinder values of primer sets LSM7 and SlCBL1 were even lower, indicating the stability of their expression also under these conditions. Primer efficiency differed by ‐18 to +5 percentage points from values presented in the literature. Our findings show that a reference primer set which delivers the best results in one system may be outperformed by another under different experimental conditions, thus recommending a reassessment of both expression stability and qPCR efficiency whenever the biological or technical experimental set‐up is changed. On the basis of our results, we recommend the use of LSM7 and SlCBL1 as reference primer sets for gene expression studies on plant tissue derived from open or semi‐controlled conditions. [ABSTRACT FROM AUTHOR]

Pectate lyase genes have been documented as excellent candidates for improvement of fruit firmness. However, implementation of pectate lyase in regulating fruit postharvest deterioration has not been fully explored. In this report, 22 individual pectate lyase genes in tomato were identified, and one pectate lyase gene Sl PL (Solyc03g111690) showed dominant expression during fruit maturation. RNA interference of Sl PL resulted in enhanced fruit firmness and changes in pericarp cells. More importantly, the Sl PL- RNAi fruit demonstrated greater antirotting and pathogen-resisting ability. Compared to wild-type, Sl PL- RNAi fruit had higher levels of cellulose and hemicellulose, whereas the level of water-soluble pectin was lower. Consistent with this, the activities of peroxidase, superoxide dismutase and catalase were higher in Sl PL- RNAi fruit, and the malondialdehyde concentration was lower. RNA-Seq results showed large amounts of differentially expressed genes involved in hormone signalling, cell wall modification, oxidative stress and pathogen resistance. Collectively, these data demonstrate that pectate lyase plays an important role in both fruit softening and pathogen resistance. This may advance knowledge of postharvest fruit preservation in tomato and other fleshy fruit. [ABSTRACT FROM AUTHOR]

Adverse environmental conditions with increasing soil salinization have become a major problem in realizing the potential yields of crops. Plants deploy various mechanisms to deal with salt stress facilitated by the expression of several stress related genes. One such group is the late embryogenesis abundant (LEA) protein gene family which has various functions including protection of cellular structures from dehydration. To investigate the molecular basis of high tolerance of Punjab Keshari cv of tomato, we performed a comprehensive analysis of the proximal promoters of candidate members Rab1A, Rab1B, Rab1C, Rab11a of Rab gene family which are essential components of stress response. Genes and protein structures were also studied to identify the active sites. The completion and enhanced annotations of tomato genome sequence has provided the opportunity for genome-wide in-depth analysis of gene expression. Analysis suggests presence of various common stress related cis-acting elements (CREs) in the putative promoters of members of this gene family. Our results strengthen the idea that plants are comprised of numerous genes with an exceptionally wide range of functionalities along with huge number of promoters and regulatory elements and more remains to be unravelled. With advances in bioinformatics, functional genomics will aid in understanding the molecular and physiological basis to improve the salinity tolerance for sustainable crop production. [ABSTRACT FROM AUTHOR]

Background: Gene co-expression, the similarity of gene expression profiles under various experimental conditions, has been used as an indicator of functional relationships between genes, and many co-expression databases have been developed for predicting gene functions. These databases usually provide users with a co-expression network and a list of strongly co-expressed genes for a query gene. Several of these databases also provide functional information on a set of strongly co-expressed genes (i.e., provide biological processes and pathways that are enriched in these strongly co-expressed genes), which is generally analyzed via over-representation analysis (ORA). A limitation of this approach may be that users can predict gene functions only based on the strongly co-expressed genes. Results: In this study, we developed a new co-expression database that enables users to predict the function of tomato genes from the results of functional enrichment analyses of co-expressed genes while considering the genes that are not strongly co-expressed. To achieve this, we used the ORA approach with several thresholds to select co-expressed genes, and performed gene set enrichment analysis (GSEA) applied to a ranked list of genes ordered by the co-expression degree. We found that internal correlation in pathways affected the significance levels of the enrichment analyses. Therefore, we introduced a new measure for evaluating the relationship between the gene and pathway, termed the percentile (p)-score, which enables users to predict functionally relevant pathways without being affected by the internal correlation in pathways. In addition, we evaluated our approaches using receiver operating characteristic curves, which concluded that the p-score could improve the performance of the ORA. Conclusions: We developed a new database, named Co-expressed Pathways DataBase for Tomato, which is available at http://cox-path-db.kazusa.or.jp/tomato. The database allows users to predict pathways that are relevant to a query gene, which would help to infer gene functions. [ABSTRACT FROM AUTHOR]

Background: Unilateral incompatibility (UI) is an asymmetric reproductive barrier that unidirectionally prevents gene flow between species and/or populations. UI is characterized by a compatible interaction between partners in one direction, but in the reciprocal cross fertilization fails, generally due to pollen tube rejection by the pistil. Although UI has long been observed in crosses between different species, the underlying molecular mechanisms are only beginning to be characterized. The wild tomato relative Solanum habrochaites provides a unique study system to investigate the molecular basis of this reproductive barrier, as populations within the species exhibit both interspecific and interpopulation UI. Here we utilized a transcriptomic approach to identify genes in both pollen and pistil tissues that may be key players in UI. Results: We confirmed UI at the pollen-pistil level between a self-incompatible population and a self-compatible population of S. habrochaites. A comparison of gene expression between pollinated styles exhibiting the incompatibility response and unpollinated controls revealed only a small number of differentially expressed transcripts. Many more differences in transcript profiles were identified between UI-competent versus UI-compromised reproductive tissues. A number of intriguing candidate genes were highly differentially expressed, including a putative pollen arabinogalactan protein, a stylar Kunitz family protease inhibitor, and a stylar peptide hormone Rapid ALkalinization Factor. Our data also provide transcriptomic evidence that fundamental processes including reactive oxygen species (ROS) signaling are likely key in UI pollen-pistil interactions between both populations and species. Conclusions: Gene expression analysis of reproductive tissues allowed us to better understand the molecular basis of interpopulation incompatibility at the level of pollen-pistil interactions. Our transcriptomic analysis highlighted specific genes, including those in ROS signaling pathways that warrant further study in investigations of UI. To our knowledge, this is the first report to identify candidate genes involved in unilateral barriers between populations within a species. [ABSTRACT FROM AUTHOR]

Small RNAs are a class of short non-coding endogenous RNAs that play essential roles in many biological processes. Recent studies have reported that microRNAs (miRNAs) are also involved in ethylene signaling in plants. LeERF1 is one of the ethylene response factors (ERFs) in tomato that locates in the downstream of ethylene signal transduction pathway. To elucidate the intricate regulatory roles of small RNAs in ethylene signaling pathway in tomato, the deep sequencing and bioinformatics methods were combined to decipher the small RNAs landscape in wild and sense-/antisense-LeERF1 transgenic tomato fruits. Except for the known miRNAs, 36 putative novel miRNAs, 6 trans-acting short interfering RNAs (ta-siRNAs), and 958 natural antisense small interfering RNAs (nat-siRNAs) were also found in our results, which enriched the tomato small RNAs repository. Among these small RNAs, 9 miRNAs, and 12 nat-siRNAs were differentially expressed between the wild and transgenic tomato fruits significantly. A large amount of target genes of the small RNAs were identified and some of them were involved in ethylene pathway, including AP2 TFs, auxin response factors, F-box proteins, ERF TFs, APETALA2-like protein, and MADS-box TFs. Degradome sequencing further confirmed the targets of miRNAs and six novel targets were also discovered. Furthermore, a regulatory model which reveals the regulation relationships between the small RNAs and their targets involved in ethylene signaling was set up. This work provides basic information for further investigation of the function of small RNAs in ethylene pathway and fruit ripening. [ABSTRACT FROM AUTHOR]

Phenotypic plasticity is the phenomenon that one particular genotype produces different phenotypes under different environmental conditions, but its underlying molecular and genetic mechanisms are poorly understood. Plastic traits may be under the control of genes whose expression is modulated by environmental cues. In this study, we investigated phenotypic plasticity in tomato (Solanum lycopersicum) and its ancestral species S. pimpinellifolium by comparing the global gene expression of young seedlings grown under two distinct growth conditions. Our results show that more than 7000 genes exhibited differential expression in response to environmental changes from phytotron to a plastic greenhouse, and 98 environmentally sensitive genes displayed the same patterns of expression response across the two tomato species. We also found that growth conditions had a remarkable impact on transcriptome complexity, attributable to alternative splicing (AS), in which 665 splice variants showed differential expression in response to the environmental changes. Moreover, more splice variants and AS events per gene were detected in plastic greenhouse-grown seedlings than their phytotron counterparts, and these seedlings also had higher percentages of intron retention events. The identification of the conserved environmentally-sensitive genes and the splice variants in this study will be useful for further analysis of gene regulation of environmental response in tomato and other crops. [ABSTRACT FROM AUTHOR]

Background: Tomatoes provide a significant dietary source of the carotenoids, lycopene and β-carotene. During ripening, carotenoid accumulation determines the fruit colors while chlorophyll degradation. These traits have been, and continue to be, a significant focus for plant breeding efforts. Previous work has found strong evidence for a relationship between CYC-B gene expression and the orange color of fleshy fruit. Other work has identified a point mutation in SGR that impedes chlorophyll degradation and causes brown flesh color to be retained in some tomato varieties. Methods: We crossed two inbred lines, KNY2 (orange) and KNB1 (brown) and evaluated the relationship between these genes for their effect on fruit color. Phenotypes of F2 generation plants were analyzed and a novel 'orange-brown' fruit color was identified. Results: We confirm two SNPs, one in CYC-B and another in SGR gene sequence, associated with segregation of 'orange-brown' fruit color in F2 generation. The carotenoid and chlorophyll content of a fleshy fruit was assessed across the different phenotypes and showed a strong correlation with expression pattern of carotenoid biosynthesis genes and SGR function. The orange-brown fruit has high β-carotene and chlorophyll. Our results provide valuable information for breeders to develop tomato fruit of a novel color using molecular markers. [ABSTRACT FROM AUTHOR]

Gametogenesis is a key step in the production of ovules or pollen in higher plants. The molecular aspects of gametogenesis are well characterized in the model plant Arabidopsis; however, little information is known in tomato, which is a model plant for fleshy fruit development. In this study, we characterized a tomato (Solanum lycopersicum L.) γ-ray mutant, sexual sterility (Slses), that exhibited both male and female sterility. Morphological analysis revealed that the Slses mutant forms incomplete ovules and wilted anthers devoid of pollen grains at the anthesis stage. Genetic and next-generation sequencing analyses revealed that the Slses mutant carried a 13 bp deletion within the first exon of a homolog of SPOROCYTELESS/NOZZLE (SPL/NZZ), which plays an important role in gametogenesis in Arabidopsis. Complementation analysis in which the complete SlSES genomic region was introduced into the Slses mutant fully restored normal phenotypes, demonstrating that Solyc07g063670 is responsible for the Slses mutation. SlSES probably act as a transcriptional repressor because of an EAR motif at the C-terminal region. Gene expression levels of WUSCHEL (SlWUS) and INNER NO OUTER (SlINO), both of which are required for ovule development, were dramatically reduced in the early stages of pistil development in the Slses mutant, suggesting a positive regulatory role for SlSES in the transcription of gametogenesis genes and differences in the regulation of INO (SlINO) and integument development by SPL/NZZ (SLSES) between Arabidopsis and tomato. Taken together, our results indicate that SlSES is a novel tomato gametogenesis gene essential for both male and female gametogenesis. [ABSTRACT FROM AUTHOR]

Antimicrobial peptides (AMPs) are produced by all living organisms and play an important role in innate immunity because they are readily available and non-specific against invading pathogenic microorganisms. Snakin-2 (SN2) from tomato is a short, cationic peptide that forms lethal pores in biomembranes of microbes. In plant cells, SN2 is produced as a prepeptide with a signal sequence for ER targeting and an acidic region to decrease toxicity in the producing organism. Gene expression analysis by qRT-PCR in tomato plants demonstrated that SN2 is constitutively expressed, mostly in leaves and flowers. After fungal infection, wounding, or external application of phytohormones (such as methyl jasmonate, MeJa) operating in the JA-dependent defense response, a systemic reaction with an elevated expression of the SN2 gene is triggered in all parts of tomato plants. Abiotic stress factors like extreme temperatures or dehydration do not affect SN2 expression. Upon wounding, the expression of SN2 and LoxD are strongly enhanced in tomato fruits. Furthermore, we provide evidence that the protein level of bioactive SN2 is also increased upon application of methyl jasmonate in tomato seedlings. [ABSTRACT FROM AUTHOR]

Targeted manipulation of phenylalanine (Phe) synthesis is a potentially powerful strategy to boost biologically and economically important metabolites, including phenylpropanoids, aromatic volatiles and other specialized plant metabolites. Here, we use two transgenes to significantly increase the levels of aromatic amino acids, tomato flavour-associated volatiles and antioxidant phenylpropanoids. Overexpression of the petunia MYB transcript factor, ODORANT1 ( ODO1), combined with expression of a feedback-insensitive E. coli 3-deoxy-D-arabino-heptulosonate 7-phosphate synthase ( AroG), altered the levels of multiple primary and secondary metabolites in tomato fruit, boosting levels of multiple secondary metabolites. Our results indicate that coexpression of AroG and ODO1 has a dual effect on Phe and related biosynthetic pathways: (i) positively impacting tyrosine (Tyr) and antioxidant related metabolites, including ones derived from coumaric acid and ferulic acid; (ii) negatively impacting other downstream secondary metabolites of the Phe pathway, including kaempferol-, naringenin- and quercetin-derived metabolites, as well as aromatic volatiles. The metabolite profiles were distinct from those obtained with either single transgene. In addition to providing fruits that are increased in flavour and nutritional chemicals, coexpression of the two genes provides insights into regulation of branches of phenylpropanoid metabolic pathways. [ABSTRACT FROM AUTHOR]

The objective of this study was to investigate the phenotypic and molecular variability in a F2 generation derived from a SCH (Second Cycle Hybrid) in order to detect QTLs for some fruit traits of tomato. Genome coverage at different levels was achieved by three types of molecular markers (polypeptides, sequence-related amplified polymorphism-SRAP and amplified restriction fragment polymorphism - AFLP). Different degrees of polymorphism were detected by SRAP and AFLP at the DNA structure level and also by polypeptides at the DNA expression level. The first two markers, associated with phenotypic variation, detected QTLs involved in important agronomic traits such as fruit shelf life, soluble solids content, pH, and titratable acidity. New gene blocks originated by recombination during the first cycle of crossing were detected. This study confirmed that the observed phenotypic differences represent a new gene rearrangement and that these new gene blocks are responsible for the presence of the genetic variability detected for these traits. [ABSTRACT FROM AUTHOR]

Inorganic nitrogen is a key element for plant growth under salt stress. A comparative study including physiological responses, ion content, transcript regulation of ammonium/nitrate transporters ( AMTs / NRTs ) as well as key enzymes for nitrogen assimilation was undertaken in wild salt-tolerant tomatoes ( Solanum pennellii ) and cultivated tomatoes ( Solanum lycopersicon ) exposed to 100 mM NaCl for 1 and 7 days. In comparison to S. lycopersicon , S. pennellii was more salt tolerant as evidenced by its higher survival rate, lower biomass reduction, and less salt injury (reduced electrolyte leakage and proline accumulation). In root tissues of both species, salt exposure (7 days) reduced the mRNA expression levels of low affinity nitrate transporters ( NRT1.1 and NRT1.2 ). This was associated with a decline in both nitrate content and expression level of the nitrate reductase gene ( NR ). Salt-stressed root tissues of S. pennellii showed relatively higher mRNA expression of the high affinity ammonium transporters ( AMT1.1 and AMT1.2 ) compared to S. lycopersicon. The root ammonium content was increased only in S. lycopersicon going hand in hand with a reduction in mRNA level of cytosolic glutamine synthetase ( GS1 ) after 7 days of salt stress, whereas the expression level of GS1 was unchanged in S. pennellii , suggesting a lower salt-induced inhibition in ammonium assimilation in this species. Our comparative study demonstrated that the salt-tolerant and salt-sensitive tomato species show differential contribution of the nitrogen transporters and key genes associated with nitrogen assimilation under salt stress. While the reduction in the expression of key components of NO 3 − uptake ( NRT1.1, NRT1.2 ) and assimilation ( NR gene) in both species, likely contributed to the reduction in plant growth under salt stress, the observed salt tolerance for S. pennellii was associated with relative higher mRNA expression of ammonium uptake and assimilation genes. These results provide crucial knowledge for tomato breeding employing salt-tolerant wild species in salt-induced nitrogen- deficient environments. [ABSTRACT FROM AUTHOR]

The aluminum-activated malate transporter (ALMT) family of proteins transports malate and/or inorganic anions across plant membranes. To demonstrate the possible role of ALMT genes in tomato fruit development, we focused on SlALMT4 and SlALMT5, the two major genes expressed during fruit development. Predicted proteins were classified into clade 2 of the family, many members of which localize to endomembranes. Tissue-specific gene expression was determined using transgenic tomato expressing the β-glucuronidase reporter gene controlled by their own promoters. Both the genes were expressed in vascular bundles connecting to developing seeds in fruit and in the embryo of mature seeds. Further, SlALMT5 was expressed in embryo in developing seeds in fruit. Subcellular localization of both proteins to the endoplasmic reticulum (ER) was established by transiently expressing the green fluorescent protein fusions in plant protoplasts. SlALMT5 probably localized to other endomembranes as well. Localization of SlALMT5 to the ER was also confirmed by immunoblot analysis. The transport function of both SlALMT proteins was investigated electrophysiologically in Xenopus oocytes. SlALMT5 transported malate and inorganic anions such as nitrate and chloride, but not citrate. SlALMT4 also transported malate, but the results were less consistent perhaps because it did not localize strongly to the plasma membrane. To elucidate the physiological role of SlALMT5 further, we overexpressed SlALMT5 in tomato. Compared with the wild type, overexpressors exhibited higher malate and citrate contents in mature seeds, but not in fruit. We conclude that the malate transport function of SlALMT5 expressed in developing fruit influences the organic acid contents in mature seeds. [ABSTRACT FROM AUTHOR]

Metacaspases play critical roles in developmentally regulated and environmentally induced programmed cell death in plants. In this study, we systematically identified and analyzed metacaspase gene family in tomato ( Solanum lycopersicum ). The results illustrated that tomato possesses eight metacaspase genes ( SlMC1 − 8 ) located on chromosomes 1, 3, 5, 9, and 10. SlMC1 − 6 belonged to type I metacaspases and had 5 exon/4 intron structures. SlMC7 and 8 were type II metacaspases and had 2 and 3 exons, respectively. Expression analysis revealed distinct expression patterns of SlMCs in various tomato tissues. Cis -regulatory element prediction showed that there were many hormone- and stress-related cis -regulatory elements in SlMCs promoter regions. Quantitative real-time PCR analysis further demonstrated that most of the SlMCs were regulated by drought, cold, salt, methyl viologen, and ethephon treatments. This study provides insights into the characteristics of SlMC genes and laid the foundation for further functional analysis of these genes in tomato. [ABSTRACT FROM AUTHOR]

Local translocation of small RNAs between cells is proved. Long distance translocation between rootstock and scion is also well documented in the homo-grafting system, but the process in distant-grafting is widely unexplored where rootstock and scion belonging to different genera. Micro RNAs are a class of small, endogenous, noncoding, gene silencing RNAs that regulate target genes of a wide range of important biological pathways in plants. In this study, tomato was grafted onto goji (Lycium chinense Mill.) to reveal the insight of miRNAs regulation and expression patterns within a distant-grafting system. Goji is an important traditional Chinese medicinal plant with enriched phytochemicals. Illumina sequencing technology has identified 68 evolutionary known miRNAs of 37 miRNA families. Moreover, 168 putative novel miRNAs were also identified. Compared with control tomato, 43 (11 known and 32 novels) and 163 (33 known and 130 novels) miRNAs were expressed significantly different in shoot and fruit of grafted tomato, respectively. The fruiting stage was identified as the most responsive in the distant-grafting approach and 123 miRNAs were found as up-regulating in the grafted fruit which is remarkably higher compare to the grafted shoot tip (28). Potential targets of differentially expressed miRNAs were found to be involved in diverse metabolic and regulatory pathways. ADP binding activities, molybdopterin synthase complex and RNA helicase activity were found as enriched terms in GO (Gene Ontology) analysis. Additionally, "metabolic pathways" was revealed as the most significant pathway in KEGG (Kyoto Encyclopedia of Genes and Genomes) analysis. The information of the small RNA transcriptomes that are obtained from this study might be the first miRNAs elucidation for a distant-grafting system, particularly between goji and tomato. The results from this study will provide the insights into the molecular aspects of miRNA-mediated regulation in the medicinal plant goji, and in grafted tomato. Noteworthy, it would provide a basis how miRNA signals could exchange between rootstock and scion, and the relevance to diverse biological processes. [ABSTRACT FROM AUTHOR]

Heavy metals are significant abiotic stress factor, affecting various response mechanisms in plants. These responses include: changes in membrane composition, production of small molecules and free radicals, and alterations in the activities of antioxidant enzymes and their gene expressions. For this reason, lipid peroxidation levels (MDA), catalase enzyme activity, and gene expression profiles, quantified by real-time PCR, were analyzed in tomato plants exposed to various concentrations (0, 80, 160, 320, 640 and 1280 µM) of Cd2+ and Pb2+ . All concentration of Cd2+ or Pb+2 contamination led to increased lipid peroxidation and catalase enzyme activity, except for 320 and 640 µM Cd+2 contamination levels. As a result, gene expression patterns at the mRNA level and changes in MDA content under different concentrations of Pb+2 and Cd+2 contamination revealed a positive correlation, although no correlation was found between gene expression patterns at the mRNA level and catalase enzyme activity. These results might be explained by the regulation of genes at the transcriptional, posttranscriptional, and also translational or posttranslational levels. [ABSTRACT FROM AUTHOR]

Heavy metals such as Cd are considered to be the most important pollutants in soil contamination. Cd is a non-essential element adversely affecting plant growth and development, and it has caused some physiological and molecular changes. Metallothioneins (MTs) are low molecular weight, cysteine-rich, and metal binding proteins. In this study, we aimed to evaluate the MT gene expression levels and minerals uptake in the tissues of Solanum lycopersicum exposed to Cd. The transcriptional expression of the MT genes was determined by real-time quantitative PCR. The MT genes were regulated by the Cd and the mineral elements uptake changed tissue type and applied doses. The MT1 and MT2 transcript levels increased in the roots, the leaves and the fruits of the tomato. The MT3 and MT4 transcript pattern changed according to the tissue types. The Cd treatment on the growth medium increased the Mg, Ca, and Fe content in both the leaves and fruits of the tomato. However, the Cd affected the mineral levels in the roots depending on the mineral types and doses. Also, the Cd content increased in the roots, the leaves, and the fruits of the tomato, respectively. The results presented in this study show that Cd has synergistic and/or antagonistic effects on minerals depending on the tissue types. These results indicate that the MT1 and MT2 expression pattern increased together with the Mg, Ca, and Fe content in both the leaves and the fruits of the tomato. [ABSTRACT FROM AUTHOR]

Drought stress has adverse impacts on plant production and productivity. MicroRNAs (miRNAs) are one class of noncoding RNAs regulating gene expression post-transcriptionally. In this study, we employed small RNA and degradome sequencing to systematically investigate the tissuespecific miRNAs responsible to drought stress, which are understudied in tomato. For this purpose, root and upground tissues of two different drought-responsive tomato genotypes (Lycopersicon esculentum as sensitive and L. esculentum var. cerasiforme as tolerant) were subjected to stress with 5% polyethylene glycol for 7 days. A total of 699 conserved miRNAs belonging to 578 families were determined and 688 miRNAs were significantly differentially expressed between different treatments, tissues and genotypes. Using degradome sequencing, 44 target genes were identified associated with 36 miRNA families. Drought-related miRNAs and their targets were enriched functionally by Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analyses. Totally, 53 miRNAs targeted 23 key drought stress- and tissue development-related genes, including DRP (dehydration-responsive protein), GTs (glycosyltransferases), ERF (ethylene responsive factor), PSII (photosystem II) protein, HD-ZIP (homeodomain-leucine zipper), MYB and NAC-domain transcription factors. miR160, miR165, miR166, miR171, miR398, miR408, miR827, miR9472, miR9476 and miR9552 were the key miRNAs functioning in regulation of these genes and involving in tomato response to drought stress. Additionally, plant hormone signal transduction pathway genes were differentially regulated by miR169, miR172, miR393, miR5641, miR5658 and miR7997 in both tissues of both sensitive and tolerant genotypes. These results provide new insight into the regulatory role of miRNAs in drought response with plant hormone signal transduction and drought-tolerant tomato breeding. [ABSTRACT FROM AUTHOR]

Background: The INNER NO OUTER (INO) gene is expressed in the outermost cell layer of the outer integument of bitegmic ovules and is essential for this organ's growth. The role and cross-species functional conservation of INO orthologs were examined in members of the Solanaceae, which have unitegmic ovules. Unitegmy has evolved several times in disparate angiosperm lineages. INO expression has been observed in the outermost cell layers of all examined unitegmic ovules, but the functional role of INO in unitegmic ovules has not previously been evaluated. Results: INO orthologs were unambiguously identified in tobacco and tomato by sequence homology. Expression of the tomato INO gene was limited to the outer cell layer of the single integument indicating that this single integument has properties of the outer integument. Expression occurred only after integument initiation, later than observed in ovules of other examined angiosperms. Virus-induced knock-down of expression of the INO ortholog in tobacco inhibited growth of the outer cell layer of the integument leading to a decrease in both integument extension and curvature of the ovule. The altered ovules closely resemble those of the aberrant testa shape (ats) ino mutant combination in Arabidopsis where we see the effect of the ino mutation on a single fused integument produced by the ats mutation. Despite significant sequence identity and similar expression patterns, the tomato INO coding region was not able to complement the Arabidopsis ino mutant. Conclusions: The similarity of effects of ino mutations on the unitegmic ovules of tobacco and the fused integuments of the Arabidopsis ats mutant show that: 1) INO orthologs play the same role in promoting integument growth in ovules of tobacco and Arabidopsis; and 2) the unitegmic ovules of tobacco (and hence other solanaceous species) are most likely the result of a congenital fusion of two ancestral integuments. Our results further indicate that INO has a conserved role in growth of the outermost cell layer of integuments. The curvature of solanaceous ovules is driven by unequal growth of the outer layers of the single integument that likely correspond to an ancestral outer integument. [ABSTRACT FROM AUTHOR]

To establish a successful infection viruses need to overcome plant innate immune responses and redirect host gene expression for their multiplication and diffusion. Tomato yellow leaf curl Sardinia virus (TYLCSV) is a geminivirus, which causes significant economic losses in tomato. The multifunctional replication associated geminivirus protein (Rep) has an important role during viral infection. In particular, the Rep central domain spanning from aa 120 to 180 is known to interact with viral and host factors. In this study, we used long serial analysis of gene expression to analyse the transcriptional profiles of transgenic tomato plants expressing the first 210 amino acids of TYLCSV Rep (Rep210) and TYLCSV-infected wild-type tomato plants (Wt-Ty). Also, we compared these profiles with those of transgenic Rep130 tomatoes. Comparison of Wt-Ty and Rep210 libraries with the wild-type one identified 118 and 203 differentially expressed genes (DEGs), respectively. Importantly, 55% of Wt-Ty DEGs were in common with Rep210, and no ones showed opposite expression. Conversely, a negligible overlap was found between Rep130 DEGs and Wt-Ty and Rep210 ones. TYLCSV- and Rep210-repressed genes, but not induced ones, overlapped with the leaf senescence process. Interestingly, TYLCSV upregulates expression of genes involved in the negative regulation of programmed cell death (PCD), several of which were also regulated by the abscisic acid. Rep210 upregulated genes related to defence response, immune system processes and negative regulation of PCD. Collectively, our results support a model in which the Rep central domain has a pivotal role in redirecting host plant gene expression. [ABSTRACT FROM AUTHOR]

Herbaceous species can modify leaf structure during the growing season in response to drought stress and water loss. Evolution can select combinations of traits in plants for efficient water use in restricted environments. We investigated plant traits that mediate adaptation and acclimation to water stress in two herbaceous drought-tolerant species. Anatomical, morphological and physiological traits related to stems and leaves were examined under optimal watering ( OW) and a long period of restricted watering ( RW) in 11 accessions from three Solanaceae species ( Solanum chilense, S. peruvianum and S. lycopersicum). The relationships between these traits were tested using linear regression and PCA. There were significant differences in anatomical traits between the species under both OW and RW, where leaf area correlated with stem diameter. Proline and total carbohydrates accumulated highly in S. chilense and S. peruvianum, respectively, and these osmolytes were strongly correlated with increased osmotic potential. Stomatal density varied between species but not between acclimation treatments, while stomatal rate was significantly higher in wild tomatoes. There was a strong positive relationship between stem growth rate and a group of traits together expressed as total stomatal number. Total stomata is described by integration of leaf area, stomatal density, height and internode length. It is proposed that constitutive adaptations and modifications through acclimation that mediate RW play an important role in tolerance to drought stress in herbaceous plants. The capacity for growth under drought stress was not associated with any single combination of traits in wild tomatoes, since the two species differed in relative levels of expression of various phenotypic traits. [ABSTRACT FROM AUTHOR]

Plant expansins are capable of inducing pH-dependent cell wall extension and stress relaxation. They may be useful as targets for crop improvement to enhance fruit development and stress resistance. Tomato is a major agricultural crop and a model plant for studying fruit development. Because only some tomato expansins have been studied, a genome-wide analysis of the tomato expansin family is necessary. In this study, we identified 25 SlEXPAs, eight SlEXPBs, one SlEXLA, four SlEXLBs, and five short homologs in the tomato genome. 25 of these genes were identified as being expressed. Bioinformatic analysis showed that although tomato expansins share similarities with those from other plants, they also exhibit specific features regarding genetic structure and amino acid sequences, which indicates a unique evolutionary process. Segmental and tandem duplication events have played important roles in expanding the tomato expansin family. Additionally, the 3-exon/2-intron structure may form the basic organization of expansin genes. We identified new expansin genes preferentially expressed in fruits ( SlEXPA8, SlEXPB8, and SlEXLB1), roots ( SlEXPA9, SlEXLB2, and SlEXLB4), and floral organs. Among the analyzed genes those that were inducible by hormone or stress treatments, including SlEXPA3, SlEXPA7, SlEXPB1- B2, SlEXPB8, SlEXLB1- LB2, and SlEXLB4. Our findings may further clarify the biological activities of tomato expansins, especially those related to fruit development and stress resistance, and contribute to the genetic modification of tomato plants to improve crop quality and yield. [ABSTRACT FROM AUTHOR]

The gene expression stability of candidate reference genes in the roots and leaves of Solanum lycopersicum inoculated with arbuscular mycorrhizal fungi was investigated. Eight candidate reference genes including elongation factor 1 α ( EF1 ), glyceraldehyde-3-phosphate dehydrogenase ( GAPDH ), phosphoglycerate kinase ( PGK ), protein phosphatase 2A ( PP2Acs ), ribosomal protein L2 ( RPL2 ), β-tubulin ( TUB ), ubiquitin ( UBI ) and actin ( ACT ) were selected, and their expression stability was assessed to determine the most stable internal reference for quantitative PCR normalization in S. lycopersicum inoculated with the arbuscular mycorrhizal fungus Rhizophagus irregularis . The stability of each gene was analysed in leaves and roots together and separated using the geNorm and NormFinder algorithms. Differences were detected between leaves and roots, varying among the best-ranked genes depending on the algorithm used and the tissue analysed. PGK , TUB and EF1 genes showed higher stability in roots, while EF1 and UBI had higher stability in leaves. Statistical algorithms indicated that the GAPDH gene was the least stable under the experimental conditions assayed. Then, we analysed the expression levels of the LePT4 gene, a phosphate transporter whose expression is induced by fungal colonization in host plant roots. No differences were observed when the most stable genes were used as reference genes. However, when GAPDH was used as the reference gene, we observed an overestimation of LePT4 expression. In summary, our results revealed that candidate reference genes present variable stability in S. lycopersicum arbuscular mycorrhizal symbiosis depending on the algorithm and tissue analysed. Thus, reference gene selection is an important issue for obtaining reliable results in gene expression quantification. [ABSTRACT FROM AUTHOR]

Long noncoding RNAs (lnc RNAs) regulate gene expression and biological processes. With the development of high-throughput RNA sequencing technology, lnc RNAs have been extensively studied in recent years. Nevertheless, the expression and evolution of lnc RNAs in plants remain poorly understood., Here, we identified 413 and 709 multi-exon noncoding transcripts from 353 and 595 loci of the cultivar tomato Heinz1706 and its wild relative LA1589, respectively. Systematic comparison of the sequence and expression of lnc RNAs showed that they are poorly conserved in Solanaceae, with only < 0.4% lnc RNAs present in all sequenced genomes of tomato and potato. Sequence analysis of Lycopersicon-specific lnc RNA loci in Solanum lycopersicum and S. pennellii showed that the origins of these molecules are associated with transposable elements ( TEs)., Lnc RNA-314, a fruit-specific lnc RNA expressed in S. lycopersicum and S. pimpinellifolium, but not in S. pennellii, originated through two evolutionary events: speciation of S. pennellii resulted in insertion of a long terminal repeat ( LTR) retrotransposon into chromosome 10 and contributed to most of the transcribed region of lnc RNA-314; and a large deletion in Lycopersicon generated the promoter region and part of the transcribed region of lnc RNA-314., These results provide novel insights into the evolution of lnc RNAs in plants. [ABSTRACT FROM AUTHOR]

Ascorbate (AsA) is an antioxidant that can scavenge the reactive oxygen species (ROS) produced when plants encounter stressful conditions. Here, it was revealed by a yeast one-hybrid assay that a tomato (Solanum lycopersicum) HD-Zip I family transcription factor, SlHZ24, binds to the promoter of an AsA biosynthetic gene encoding GDP-D-mannose pyrophosphorylase 3 (SlGMP3). Both the transient expression system and electrophoretic mobility shift assay (EMSA) showed that SlHZ24 binds to a regulatory cis-element in the SlGMP3 promoter, and further overexpression of SlHZ24 in transgenic tomato lines resulted in increased AsA levels. In contrast, suppressing expression of the gene using RNA interference (RNAi) had the opposite effect. These data suggest that SlHZ24 can positively regulate the accumulation of AsA, and in support of this it was shown that SlGMP3 expression increased in the SlHZ24-overexpressing lines and declined in SlHZ24-RNAi lines. SlHZ24 also affected the expression of other genes in the D-mannose/L-galactose pathway, such as genes encoding GDP-mannose-3',5'-epimerase 2 (SlGME2), GDP-L-galactose phosphorylase (SlGGP) and SlGMP4. The EMSA indicated that SlHZ24 bound to the promoters of SlGME2 and SlGGP, suggesting multi-targeted regulation of AsA biosynthesis. Finally, SlHZ24-overexpressing plants showed less sensitivity to oxidative stress; we therefore conclude that SlHZ24 promotes AsA biosynthesis, which in turn enhances oxidative stress tolerance. [ABSTRACT FROM AUTHOR]

Background: Salinity-alkalinity stress is known to adversely affect a variety of processes in plants, thus inhibiting growth and decreasing crop yield. Polyamines protect plants against a variety of environmental stresses. However, whether exogenous spermidine increases the tolerance of tomato seedlings via effects on chloroplast antioxidant enzymes and chlorophyll metabolism is unknown. In this study, we examined the effect of exogenous spermidine on chlorophyll synthesis and degradation pathway intermediates and related enzyme activities, as well as chloroplast ultrastructure, gene expression, and antioxidants in salinity-alkalinity-stressed tomato seedlings. Results: Salinity-alkalinity stress disrupted chlorophyll metabolism and hindered uroorphyrinogen III conversion to protoporphyrin IX. These effects were more pronounced in seedlings of cultivar Zhongza No. 9 than cultivar Jinpengchaoguan. Under salinity-alkalinity stress, exogenous spermidine alleviated decreases in the contents of total chlorophyll and chlorophyll a and b in seedlings of both cultivars following 4 days of stress. With extended stress, exogenous spermidine reduced the accumulation of δ-aminolevulinic acid, porphobilinogen, and uroorphyrinogen III and increased the levels of protoporphyrin IX, Mg-protoporphyrin IX, and protochlorophyllide, suggesting that spermidine promotes the conversion of uroorphyrinogen III to protoporphyrin IX. The effect occurred earlier in cultivar Jinpengchaoguan than in cultivar Zhongza No. 9. Exogenous spermidine also alleviated the stress-induced increases in malondialdehyde content, superoxide radical generation rate, chlorophyllase activity, and expression of the chlorophyllase gene and the stress-induced decreases in the activities of antioxidant enzymes, antioxidants, and expression of the porphobilinogen deaminase gene. In addition, exogenous spermidine stabilized the chloroplast ultrastructure in stressed tomato seedlings. Conclusions: The tomato cultivars examined exhibited different capacities for responding to salinity-alkalinity stress. Exogenous spermidine triggers effective protection against damage induced by salinity-alkalinity stress in tomato seedlings, probably by maintaining chloroplast structural integrity and alleviating salinity-alkalinity-induced oxidative damage, most likely through regulation of chlorophyll metabolism and the enzymatic and non-enzymatic antioxidant systems in chloroplast. Exogenous spermidine also exerts positive effects at the transcription level, such as down-regulation of the expression of the chlorophyllase gene and up-regulation of the expression of the porphobilinogen deaminase gene. [ABSTRACT FROM AUTHOR]

Favorable temperatures for tomato (Solanum lycopersicumL.) cultivation are between 25 and 30°C during the day and 20°C at night. Average global temperatures are increasing by approximately 0.3°C per decade. A 2–4°C increase over the optimal (25°C) temperature adversely affects plant growth, flowering, gamete development, embryo development, and seed germination; inhibits the ability of pollinated flowers to develop into seeded fruit; inhibits fruit ripening; and reduces yield. Drought resulting from insufficient rainfall and/or altered precipitation patterns is another important climatic stress factor for tomato, often accompanied by relatively high temperature, in which evapotranspiration is affected and photosynthetic kinetics and crop productivity reduced. Application of genomics in tomato, to discover novel and high-throughput genetic and molecular technologies as an indirect selection tool, provides the potential for increasing efficiency of breeding by reducing the genotype × environment interaction, facilitating efficient introgression of superior alleles from wild species into plants, promoting gene-pyramiding control of quantitative traits, and enhancing development of stress-tolerant, higher yielding varieties. Advances in genetics and genomics have improved the understanding of structural and functional aspects of plant genomes. TheCBF/NHX1/DREB1genes have been used successfully to engineer drought tolerance in tomato. These genes are transcription factors implicated in tomato response to drought and heat stress. There are few tomato varieties/species which can easily adapt to stress tolerance as environmental conditions change. Some tomato varieties and wild species are heat and drought tolerant. The focus of research is to use tools and techniques to deliver global integration and overcome effects of environmental stresses on tomato. [ABSTRACT FROM PUBLISHER]

Genome-scale analyses of variation, gene expression, and metabolite accumulation in ancestral, early domesticates, and modern tomatoes by Zhu et al. identify genes underlying fruit chemistry and demonstrate that alleles affecting metabolic quality have been bred into modern varieties as a result of linkage drag. Similar metabolic hitchhikers are likely ubiquitous in other domesticated species. [ABSTRACT FROM AUTHOR]

Plant-invading microbes betray their presence to a plant by exposure of antigenic molecules such as small, secreted proteins called 'effectors'. In Fusarium oxysporum f. sp . lycopersici ( Fol) we identified a pair of effector gene candidates, AVR2- SIX5, whose expression is controlled by a shared promoter., The pathogenicity of AVR2 and SIX5 Fol knockouts was assessed on susceptible and resistant tomato ( Solanum lycopersicum) plants carrying I-2. The I-2 NB- LRR protein confers resistance to Fol races carrying AVR2., Like Avr2, Six5 was found to be required for full virulence on susceptible plants. Unexpectedly, each knockout could breach I-2-mediated disease resistance. So whereas Avr2 is sufficient to induce I-2-mediated cell death, Avr2 and Six5 are both required for resistance. Avr2 and Six5 interact in yeast two-hybrid assays as well as in planta. Six5 and Avr2 accumulate in xylem sap of plants infected with the reciprocal knockouts, showing that lack of I-2 activation is not due to a lack of Avr2 accumulation in the SIX5 mutant., The effector repertoire of a pathogen determines its host specificity and its ability to manipulate plant immunity. Our findings challenge an oversimplified interpretation of the gene-for-gene model by showing requirement of two fungal genes for immunity conferred by one resistance gene. [ABSTRACT FROM AUTHOR]

In plants, epigenetic changes have been identified as regulators of developmental events during normal growth as well as environmental stress exposures. Flavonoid biosynthetic and antioxidant pathways play a significant role in plant defence during their exposure to environmental cues. The aim of this study was to unravel whether genes encoding enzymes of flavonoid biosynthetic and antioxidant pathways are under epigenetic regulation, particularly DNA methylation, during salt stress. For this, a repressor of silencing from Arabidopsis, AtROS, was overexpressed in transgenic tobacco. Generated transgenics were evaluated to examine the influence of AtROS1 on methylation status of promoters as well as on coding regions of genes encoding enzymes of flavonoids biosynthesis and antioxidant pathways. Overexpression of AtROS1 increases the demethylation levels of both promoters as well as coding regions of genes encoding chalcone synthase, chalcone isomerase, flavanone 3-hydroxylase, flavonol synthase, dihydroflavonol 4-reductase, and anthocyanidin synthase of the flavonoid biosynthetic pathway, and glutathione S-transferase, ascorbate peroxidase, glutathione peroxidase, and glutathione reductase of the antioxidant pathway during control conditions. The level of demethylation was further increased at promoters as well as coding regions of these genes during salt-stress conditions. Transgenic tobacco overexpressing AtROS1 showed tolerance to salt stress that could have been due to the higher expression levels of the genes encoding enzymes of the flavonoid biosynthetic and antioxidant pathways. This is the first comprehensive study documenting the epigenetic regulation of flavonoid biosynthetic and antioxidant pathways during salt-stress exposure of plants. [ABSTRACT FROM AUTHOR]

Although glutathione S-transferases (GST, EC 2.5.1.18) are involved in response to abiotic stress, limited information is available regarding gene function in tomato. In this study, a GST gene from tomato, designated LeGSTU2, was cloned and functionally characterized. Expression profile analysis results showed that it was expressed in roots and flowers, and the transcription was induced by salt, osmotic, and heat stress. The gene was then introduced to Arabidopsis by Agrobacterium tumefaciens-mediated transformation. Transgenic Arabidopsis plants were normal in terms of growth and maturity compared with wild-type plants. Transgenic plants also showed an enhanced resistance to salt and osmotic stress induced by NaCl and mannitol. The increased tolerance of transgenic plants was correlated with the changes in proline, malondialdehyde and antioxidative emzymes activities. Our results indicated that the gene from tomato plays a positive role in improving tolerance to salinity and drought stresses in Arabidopsis. [ABSTRACT FROM AUTHOR]

Soil management affects chemical and microbiological properties of soils in ways that impact plant growth. We examined effects of amendment treatments and contrasting management treatments on chemical and microbiological soil properties and on expression of selected defense genes in laboratory-grown tomatoes ( Solanum lycopersicum ). Treatments imposed on a conventionally managed Maury silt loam included: compost, manure, vetch ( Vicia villosa ), inorganic N, and non-treated control. Non-amended, organically managed Maury soil, collected from the same location was also studied. Compost, manure, and vetch increased total organic C (TOC) and permanganate oxidizable C (POXC) in soil. Organically managed soil also had greater TOC and POXC than the control treatment. Soil N was increased by compost, manure, vetch and inorganic N treatment. Microbial communities from compost, vetch and manure treatments differed from each other and from communities in the other treatments. Defense genes: ChiB (chitinase), Osm (osmotin), and GluA (β-1,3-glucanase) were generally expressed less in plants from manure treated soil and organically managed soil. The PR1b (pathogenesis-related protein PR1b) gene was expressed more in plants from compost, inorganic N, and vetch treatments. Defense gene expression was negatively related to Gram-negative bacterial biomarkers, which were greatest in manure treated soil and organically managed soil. These results suggest that increased relative abundance of Gram-negative members of soil microbial communities in manure treated or organically managed soils may indirectly reduce the steady state expression of defense genes in plants, allowing plants to shift resources from defense to other beneficial functions such as fruit or biomass production. [ABSTRACT FROM AUTHOR]

SKP1 is a core component of SCF complex, a major type of E3 ubiquitin ligase catalyzing the last step in ubiquitin-mediated protein degradation pathway. In present study, SKP1 gene family in Solanum pimpinellifolium ( SSK ), a wild species of tomato, was investigated. A total of 19 SSK genes were identified through homologous search. Their chromosomal locations, gene structures, phylogeny, expression profiles, sub-cellular localizations and protein–protein interaction patterns with putative F-box proteins were analyzed in detail. The high homology and similar expression patterns among clustered SSK genes in chromosome suggested that they may have evolved from duplication events and are functionally redundant. Sub-cellular localization indicated that most of the SSK proteins are distributed in both cytosol and nucleus, except for SSK8 , which is detected in cytosol only. Tissue-specific expression patterns suggested that many SSK genes may be involved in tomato fruit development. Furthermore, several SSK genes were found to be responsive to heat stress and salicylic acid treatment. Based on phylogenetic analysis, expression profiles and protein interaction property, we proposed that tomato SSK1 and SSK2 might have similar function to ASK1 and ASK2 in Arabidopsis. [ABSTRACT FROM AUTHOR]

Citrus is a perennial woody crop of global economic importance. However, the regulatory mechanism of fruit ripening in citrus remains poorly understood. To explore the regulatory roles of transcription factors in fruit ripening, CgDREB ( d ehydration r esponsive e lement b inding) gene was characterized and was functionally overexpressed in tomato ( Solanum lycopersicum, cv. Ailsa Craig), resulting in smaller fruit size. Therefore, two independent lines with the highest expression of CgDREB were selected to study the metabolite profile in the flesh and peel of tomato fruits during ripening using gas chromatography–mass spectrometry. The results revealed that the levels of organic acids and sugars generally increased in both the flesh and peel of tomato. In addition, a significant increase of ethylene production was observed at the breaker stage, indicating a strong correlation between fruit ripening and ethylene release. Taken together, our results provide insights into the important role of CgDREB gene in regulating fruit ripening properties. [ABSTRACT FROM AUTHOR]

Fruit formation and early development involve a range of physiological and morphological transformations of the various constituent tissues of the ovary. These developmental changes vary considerably according to tissue type, but molecular analyses at an organ-wide level inevitably obscure many tissue-specific phenomena. We used laser-capture microdissection coupled to high-throughput RNA sequencing to analyze the transcriptome of ovaries and fruit tissues of the wild tomato species Solanum pimpinellifolium. This laser-capture microdissection-high-throughput RNA sequencing approach allowed quantitative global profiling of gene expression at previously unobtainable levels of spatial resolution, revealing numerous contrasting transcriptome profiles and uncovering rare and cell type-specific transcripts. Coexpressed gene clusters linked specific tissues and stages to major transcriptional changes underlying the ovary-to-fruit transition and provided evidence of regulatory modules related to cell division, photosynthesis, and auxin transport in internal fruit tissues, together with parallel specialization of the pericarp transcriptome in stress responses and secondary metabolism. Analysis of transcription factor expression and regulatory motifs indicated putative gene regulatory modules that may regulate the development of different tissues and hormonal processes. Major alterations in the expression of hormone metabolic and signaling components illustrate the complex hormonal control underpinning fruit formation, with intricate spatiotemporal variations suggesting separate regulatory programs. [ABSTRACT FROM AUTHOR]

Main conclusion: This study provided a comparative genomic analysis of the LEA gene family, and these may provide valuable information for their functional investigations in the future. Late embryogenesis abundant (LEA) proteins are a group of proteins that accumulate in response to cellular dehydration in many organisms. Here, we identified 27 LEA genes in tomato. A strong correlation between phylogeny, gene structure, and motif composition was found. The predicted SlLEA genes were non-randomly distributed within their chromosomes, and segmental and tandem duplications were probably important for their expansion. Many cis-elements potentially mediating transcription in response to abiotic stress were also found in the 1,000 bp upstream sequence of the promoter region. An additional intragenic recombination played an important role in the evolution of SlLEA genes. Selection analysis also identified some significant site-specific constraints that acted on the evolution of most LEA paralogs. Expression analysis using both microarray data and quantitative real-time PCR indicated that SlLEA genes were widely expressed in various tissues, and that a few members responded to some abiotic stresses. Our study provides useful information on the LEA genes in tomato and will facilitate their further characterization to better understand their functions. [ABSTRACT FROM AUTHOR]

Background: Next Generation Sequencing technologies (NGS) unexpectedly pushed forward the capability of solving genome organization and of widely depicting gene expression. However, although the flourishing of tools to process the NGS data, versatile and user-friendly computational environments for integrative and comparative analyses of the results from the increasing amount of collections are still required. The gene expression of tomato tissues has been widely investigated in the years, thanks to both EST sequencing and different microarray platforms. However, the resulting collections are heterogeneous in terms of experimental approaches, genotypes and conditions, making the data far from representing a gene expression atlas for the species. Therefore, the recent release of NGS transcriptome collections from several tissues and stages from physiological conditions for specific tomato genotypes provides a relevant resource to be appropriately exploited to address key questions on gene expression patterns, such as those related to fruit ripening and development in tomato. The organization of the results from the processed collections in web accessible environments, enriched with tools for their exploration, may represent a precious opportunity for the scientific research in tomato and a reference example for similar efforts. Description: Here we present the architecture and the facilities of NexGenEx-, a web based platform that offers processed NGS transcriptome collections and enables immediate analyses of the results. The platform allows gene expression investigations, profiling and comparisons, and exploits different resources. Specifically, we present here the platform partition NexGenEx-Tom, dedicated to the organization of results from tomato NGS based transcriptomes. Conclusion: In the current version, NexGenEx-Tom includes processed and normalized NGS expression data from three collections covering several tissue/stages from different genotypes. Beyond providing a user-friendly interface, the platform was designed with the aim to easily be expanded to include other NGS based transcriptome collections. It can also integrate different genome releases, possibly from different cultivars or genotypes, but even from different species. The platform is proposed as an example effort in tomato, and is described as a profitable approach for the exploitation of these challenging and precious datasets. [ABSTRACT FROM AUTHOR]

Background Reciprocal hybrids showing different phenotypes have been well documented in previous studies, and many factors accounting for different phenotypes have been extensively investigated. However, less is known about whether the profiles of small RNAs differ between reciprocal hybrids and how these small RNAs affect gene expression and phenotypes. To better understand this mechanism, the role of small RNAs on phenotypes in reciprocal hybrids was analysed. Results Reciprocal hybrids between Solanum lycopersicum cv. Micro-Tom and S. pimpinellifolium line WVa700 were generated. Significantly different phenotypes between the reciprocal hybrids were observed, including fruit shape index, single fruit weight and plant height. Then, through the high-throughput sequencing of small RNAs, we found that the expression levels of 76 known miRNAs were highly variable between the reciprocal hybrids. Subsequently, a total of 410 target genes were predicted to correspond with these differentially expressed miRNAs. Furthermore, gene ontology (GO) annotation indicated that those target genes are primarily involved in metabolic processes. Finally, differentially expressed miRNAs, such as miR156f and 171a, and their target genes were analysed by qRTPCR, and their expression levels were well correlated with the different phenotypes. Conclusions This study showed that the profiles of small RNAs differed between the reciprocal hybrids, and differentially expressed genes were also observed based on the different phenotypes. The qRT-PCR results of target genes showed that differentially expressed miRNAs negatively regulated their target genes. Moreover, the expression of target genes was well correlated with the observations of different phenotypes. These findings may aid in elucidating small RNAs contribute significantly to different phenotypes through epigenetic modification during reciprocal crossing. [ABSTRACT FROM AUTHOR]

Three independent transgenic Arabidopsis lines expressing SlTIP2;2 from Solanum lycopersicum L. cv. Lichun under the control of its endogenous promoter were used to analyze the expression of SlTIP2;2 and the salt stress tolerance under NaCl concentration gradient treatment. The expression patterns of SlTIP2;2 were shown to be tissue-specific and NaCl dose-dependent under salt stress. SlTIP2;2-transformed Arabidopsis plants exhibited enhanced salt stress tolerance, and the physiological parameters suggested that SlTIP2;2 has close links with the ion homeostasis and antioxidant enzymes activities in salt-stressed transgenic Arabidopsis. Moreover, SlTIP2;2 expression significantly affected the Na and K fluxes from the root meristematic zones and resulted in remarkable changes in the morphology of the pith ray cells in the inflorescence stems of transgenic Arabidopsis. Based on the yeast growth assay, β-galactosidase activity testing and bimolecular fluorescence complementation, SlTIP1;1, SlTIP2;1 and an UDP-galactose transporter were confirmed to interact with SlTIP2;2, which may greatly broaden our understanding of the physiological functions of aquaporins. [ABSTRACT FROM AUTHOR]

Recent studies have reported that decreased level of DNA cytosine methylation in the global genome was closely related to the initiation of tomato ( Solanum lycopersicum ) fruit ripening. However, genome-scale analysis of cytosine-5 DNA methyltransferases (C5-MTases) and demethylases in tomato has not been engaged. In this study, 7 C5-MTases and 3 demethylases were identified in tomato genome, which probably contributed to DNA cytosine methylation level in tomato. The 7 C5-MTases were categorized into 4 subgroups, and the 3 demethylases were classified into 2 subgroups based on phylogenetic analyses. Comprehensive analysis of their structure and genomic localization was also performed in this paper. According to online RNA-seq data, 4 S. lycopersicum C5-MTase (SlC5-MTase) genes ( SlMET , SlDRM1L1 , SlDRM5 , SlMET3L ) were expressed higher than others, and one DNA demethylase gene ( SlDML ) was significantly changed during tomato fruit development and ripening. Furthermore, all these five gene expressions at breaker (BK) stage changed with 1-methylcyclopropene (1-MCP) treatment, indicating that they were regulated by ethylene directly or indirectly in tomato fruit. In addition, subcellular localization analysis indicated that SlDRM1L1 and SlDRM5 located in the nucleus might have responsibility for RNA-directed DNA methylation (RdDM). Collectively, this paper provided a framework for gene discovery and functional characterization of C5-MTases and DNA demethylases in other Solanaceae species. [ABSTRACT FROM AUTHOR]