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Background: In living organisms, small heat shock proteins (sHSPs) are triggered in response to stress situations. This family of proteins is large in plants and, in the case of tomato (Solanum lycopersicum), 33 genes have been identified, most of them related to heat stress response and to the ripening process. Transcriptomic and proteomic studies have revealed complex patterns of expression for these genes. In this work, we investigate the coregulation of these genes by performing a computational analysis of their promoter architecture to find regulatory motifs known as heat shock elements (HSEs). We leverage the presence of sHSP members that originated from tandem duplication events and analyze the promoter architecture diversity of the whole sHSP family, focusing on the identification of HSEs. Results: We performed a search for conserved genomic sequences in the promoter regions of the sHSPs of tomato, plus several other proteins (mainly HSPs) that are functionally related to heat stress situations or to ripening. Several computational analyses were performed to build multiple sequence motifs and identify transcription factor binding sites (TFBS) homologous to HSF1AE and HSF21 in Arabidopsis. We also investigated the expression and interaction of these proteins under two heat stress situations in whole tomato plants and in protoplast cells, both in the presence and in the absence of heat shock transcription factor A2 (HsfA2). The results of these analyses indicate that different sHSPs are up-regulated depending on the activation or repression of HsfA2, a key regulator of HSPs. Further, the analysis of protein-protein interaction between the sHSP protein family and other heat shock response proteins (Hsp70, Hsp90 and MBF1c) suggests that several sHSPs are mediating alternative stress response through a regulatory subnetwork that is not dependent on HsfA2. Conclusions: Overall, this study identifies two regulatory motifs (HSF1AE and HSF21) associated with the sHSP family in tomato which are considered genomic HSEs. The study also suggests that, despite the apparent redundancy of these proteins, which has been linked to gene duplication, tomato sHSPs showed different up-regulation and different interaction patterns when analyzed under different stress situations. [ABSTRACT FROM AUTHOR]

New cultivation techniques associated with the use of genetic materials with high productive potential have provided the need for studies related to the accumulation of dry matter and nutrients by tomato plants. The objective of this study was to evaluate the accumulation of dry matter and macronutrients by the tomato 'Caeté' in Chapada do Apodi-CE. The accumulation of dry matter and nutrients in the vegetative part (stems and leaves) and in the fruits was quantified by sampling the plants at 14, 28, 42, 56, 70, 84 and 99 days after the transplant (DAT). The total accumulation of macronutrients in the plant followed the accumulation of dry matter, with greater demands in the fruiting phase. The accumulation of macronutrients at 99 DAT followed by the following decreasing order: K> N> Ca> S> Mg> P. [ABSTRACT FROM AUTHOR]

A fusion gene coding for trehalose-6-phosphate synthase/phosphatase (TPSP) derived from Escherichia coli was introduced into tomato plants (Solanum lycopersicum cv. Joyful) to generate overexpressing lines and enhance the level of trehalose in the seeds. Trehalose has been implicated in tolerance to abiotic stress in plants, so the response to heat stress of seeds from the transgenic lines and seeds from wild-type plants were compared. Wild-type heat-treated seeds had a germination rate of 17%, while four transgenic lines had germination rates over 50% following heat stress. In one heat-treated transgenic line, 100% of the seeds germinated, and therefore had a germination rate six times that of seeds from wild-type plants. Moreover, quantitative PCR revealed that the expression of diverse genes that respond to heat stress was enhanced in TPSP transgenic seeds compared to wild-type seeds 150 min after the onset of heat stress. The enhanced germination rate and expression of these genes in the transgenic seeds were essentially mimicked in wild-type seeds treated with 1 mM exogenous trehalose. Therefore, accumulated trehalose and associated metabolites may act as signaling molecules that enhance the expression of heat stress-responsive genes and confer heat-stress tolerance to seeds. [ABSTRACT FROM AUTHOR]

The extent of genetic variability among 19 accessions of tomato was studied using genetic variability parameters. Four-week old seedlings were transplanted in a well levelled field with 0.6 x 0.6 spacing and replicated three times in randomized complete block design. High significant differences among the accessions for all attributes studied. Cluster analysis based on 37 agro-morphological attributes separated accessions into two distinct groups according to the fruit types. Values for genotypic and phenotypic coefficients of variation showed variability among the accessions. Correlation analysis showed fruit/plant is positively and significantly correlated to plant height, number of branches/plant and leaf length. Very high genetic advance and heritability estimates for leaf length, leaf width, days to flower, days to 50% flowering, number of fruits/plant, fruit length, fruit diameter, fruit weight and 1000 seed weight suggest simple inheritance system and thus amenability for these attributes to selection in tomato improvement. [ABSTRACT FROM AUTHOR]

Salinity is a global environmental problem, restricting crop production in a vast area of agricultural land. Although epigallocatechin-3-gallate (EGCG), the most abundant polyphenol in tea plants, has a strong antioxidative action in vitro, the role of EGCG in the plant response to salt stress remains unknown. In the present study, using a series of EGCG concentrations (10, 100, and 1000 µM), we showed that EGCG could alleviate salt stress-induced inhibition in seed germination and root growth in tomato. Exogenous EGCG increased not only the seed germination rate, but also the germination energy and germination index under salt stress. The 150 mM NaCl treatment significantly increased lipid peroxidation in roots by excessive accumulation of reactive oxygen species (ROS). In contrast, EGCG treatment, particularly at 100 µM concentration, mitigated NaCl-induced oxidative stress as evidenced by the decreased H2O2 and malondialdehyde content in roots. Analysis of the antioxidant enzyme system reveals that EGCG increased the activity of superoxide dismutase, peroxidase, ascorbate peroxidase, and catalase under salt stress. Considering the beneficial effect of EGCG on seed germination, root growth, ROS scavenging, and antioxidant enzyme activity, the 100 µM EGCG treatment appears to be the most effective concentration of those tested under salt stress in tomato. Our results suggest that the EGCG-promoted tomato tolerance to salt stress is associated with the mitigation of oxidative stress through an efficient ROS scavenging mechanism by the action of enhanced antioxidant enzyme activity. Thus, the enhancement of plant tolerance by exogenous EGCG can potentially expand crop cultivation in saline soils. [ABSTRACT FROM AUTHOR]

Growth and development are vital processes in the life cycles of plants. Brassinosteroids (BRs) are steroids that when exogenously applied can regulate several biological responses. The aim of this research was to investigate the possible interferences caused by the exogenous application of BR on growth and metabolism using two genotypes of the DWARF gene, MT-d and MT-D, that are BR-deficient and BR-efficient, respectively. The experiment had four treatments with two genotypes (BR-efficient and BR-deficient) and two levels of brassinosteroids (0 and 100 nM BR, described as − BR and + BR, respectively). This study revealed that the exogenous application of BR promoted improvement in growth, inducing increases in all variables of both genotypes evaluated. In general, BR-deficient plants sprayed with BR had effects more intense, confirming the benefits of this steroid on photosynthetic apparatus and gas exchange. The changes in the anatomical characteristics of the leaf are related to the contribution of BR on the influx and consequent fixation of CO2. In addition, modifications related to root anatomy occurred as a result of the BR action with the purpose of increasing the root protection and absorption of water and nutrients. Increases in photosynthetic pigments suggest that the role of BR is linked with chlorophyll biosynthesis and the maintenance of chloroplast integrity, resulting from associations with the increases found in the activities of antioxidant enzymes that modulate the accumulation of reactive oxygen species. [ABSTRACT FROM AUTHOR]

Highlights • The presence of the glycans at the sites N94 and N112 was crucial for the activity of R-TBN1. • Hyperglycosylated mutant D36S/E104N was twice as active as R-TBN1 wt; the partial demannosylation shifted the substrate specificity of R-TBN1. • R-TBN1 was posttranslationally modified by high-mannose glycans, when produced in N. benthamiana. • Highly homologous nucleases R-HBN1 and R-ABN1 were glycosylated in a similar way as R-TBN1. • N. benthamiana can glycosylate foreign plant non-specific nucleases mainly by high-mannose type of glycosylation. Abstract A unique analysis of an enzyme activity versus structure modification of the tomato nuclease R-TBN1 is presented. R-TBN1, the non-specific nuclease belonging to the S1-P1 nuclease family, was recombinantly produced in N. benthamiana. The native structure is posttranslationally modified by N -glycosylation at three sites. In this work, it was found that this nuclease is modified by high-mannose type N -glycosylation with a certain degree of macro- and microheterogeneity. To monitor the role of N -glycosylation in its activity, hypo- and hyperglycosylated nuclease mutants, R-TBN1 digested by α-mannosidase, and R-TBN1 deglycosylated by PNGase F were prepared. Deglycosylated R-TBN1 and mutant N94D/N112D were virtually inactive. Compared to R-TBN1 wt, both N94D and N112D mutants showed about 60% and 10% of the activity, respectively, while the N186D, D36S, and D36S/E104 N mutants were equally or even more active than R-TBN1 wt. The partial demannosylation of R-TBN1 did not affect the nuclease activity; moreover, a little shift in substrate specificity was observed. The results show two facts: 1) which sites must be occupied by a glycan for the proper folding and stability and 2) how N. benthamiana glycosylates the foreign nuclease. At the same time, the modifications can be interesting in designing the nuclease activity or specificity through its glycosylation. [ABSTRACT FROM AUTHOR]

The site-directed mutagenesis of tomato allene oxide synthase LeAOS3 (CYP74C3) on the "F/L toggle" site resulted in a partial conversion of LeAOS3 into epoxyalcohol synthase. [ABSTRACT FROM AUTHOR]

Parthenocarpy and fruit malformations are common among independent transgenic tomato lines, expressing genes encoding different pathogenesis-related (PR) protein and antimicrobal peptides. Abnormal phenotype developed independently of the expression and type of target genes, but distinctive features during flower and fruit development were detected in each transgenic line. We analyzed the morphology, anatomy, and cytoembryology of abnormal flowers and fruits from these transgenic tomato lines and compared them with flowers and fruits of wild tomatoes, line YaLF used for transformation, and transgenic plants with normal phenotype. We confirmed that the main cause of abnormal flower and fruit development was the alterations of determinate growth of generative meristem. These alterations triggered different types of anomalous growth, affecting the number of growing ectopic shoots and formation of new flowers. Investigation of the ovule ontogenesis did not show anomalies in embryo sac development, but fertilization did not occur and embryo sac degenerated. Nevertheless, the ovule continued to differentiate due to proliferation of endothelium cells. The latter substituted embryo sac and formed pseudoembryonic tissue. This process imitated embryogenesis and stimulated ovary growth, leading to the development of parthenocarpic fruit. We demonstrated that failed fertilization occurred due to defective male gametophyte formation, which was manifested in blocked division of the nucleus in the microspore and arrest of vegetative and generative cell formation. Maturing pollen grains were overgrown microspores, not competent for fertilization but capable to induce proliferation of endothelium and development of parthenocarpic ovary. Thus, our study provided new data on the structural transformations of reproductive organs during development of parthenocarpic fruits in transgenic tomato. [ABSTRACT FROM AUTHOR]

Reductions in growth and quality due to powdery mildew (PM) disease cause significant economic losses in tomato production. Oidium neolycopersici was identified as the fungal species responsible for tomato PM disease in South Korea in the present study, based on morphological and internal transcribed spacer DNA sequence analyses of PM samples collected from two remote regions (Muju and Miryang). The genes involved in resistance to this pathogen in the tomato accession 'KNU-12' (Solanum lycopersicum var. cerasiforme) were evaluated, and the inheritance of PM resistance in 'KNU-12' was found to be conferred via simple Mendelian inheritance of a mutant allele of the PM susceptibility locus Ol-2 (SlMlo1). Full-length cDNA analysis of this newly identified mutant allele (Slmlo1.1) showed that a 1-bp deletion in its coding region led to a frameshift mutation possibly resulting in SlMlo1 loss-of-function. An alternatively spliced transcript of Slmlo1.1 was observed in the cDNA sequences of 'KNU-12', but its direct influence on PM resistance is unclear. A derived cleaved amplified polymorphic sequence (dCAPS) and a high-resolution melting (HRM) marker were developed based on the 1-bp deletion in Slmlo1.1, and could be used for efficient marker-assisted selection (MAS) using 'KNU-12' as the source for durable and broad-spectrum resistance to PM. [ABSTRACT FROM AUTHOR]

Phylogenetics and population genetics are central disciplines in evolutionary biology. Both are based on the comparison of single DNA sequences, or a concatenation of a number of these. However, with the advent of next-generation DNA sequencing technologies, the approaches that consider large genomic data sets are of growing importance for the elucidation of evolutionary relationships among species. Among these approaches, the assembly and alignment-free methods which allow an efficient distance computation and phylogeny reconstruction are of great importance. However, it is not yet clear under what quality conditions and abundance of genomic data such methods are able to infer phylogenies accurately. In the present study we assess the method originally proposed by Fan et al. for whole genome data, in the elucidation of Tomatoes' chloroplast phylogenetics using short read sequences. We find that this assembly and alignment-free method is capable of reproducing previous results under conditions of high coverage, given that low frequency k-mers (i.e. error prone data) are effectively filtered out. Finally, we present a complete chloroplast phylogeny for the best data quality candidates of the recently published 360 tomato genomes. [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]

Limitations in access to electricity in rural areas and substantial cost of supplemental lightning necessitate breeding as response to low light conditions. Seven inbred lines of tomato (Solanum lycopersicum L.) and their F1 hybrids, including reciprocals, developed through a 7x7 full diallel cross were evaluated under two different levels of light. Mean square for light (L) effect was significant for total yield, average fruit weight and days to first flower. Variation attributable to Genotypes and genotype x light (GxL) interaction had significant effect on all studied traits except days to ripening for which GxL interaction was not significant. Diallel analysis across two environments indicated that general (GCA), specific (SCA) and reciprocal combining ability (REC) were significant for all characters implying importance of additive and non-additive gene action along with cytoplasmic effects on genetic expression of yield, yield components and earliness. Ratio of SCA variance to SCA variance and estimates of narrow sense heritability (h²n.s) demonstrated higher weight of additive effects in inheritance of yield, fruit number and days to ripening, while indicating predominance of non-additive effects for fruit weight and early flowering. Interactions GCAxL and SCAxL were significant for almost all studied features. A particular genotype could not be recommended for all traits, but variation among genotypes in response to ambient light was promising for feasibility of plant breeding for non-optimal light intensity and duration. [ABSTRACT FROM AUTHOR]

A conventional breeding approach was framed to combine lycopene enhancing mutant gene, dg or hp-2 dg and anthocyanin synthesizing gene, Aft in one genotype in maximizing the accumulation of both lycopene and anthocyanin pigment in tomato fruit. This breeding scheme involves one crossing, Alisa Craig AftAft  × BCT-115 dgdg followed by two selfings for selection of the desirable double homozygous genotype, AftAft dgdg , the “Purple tomato” based on morphological marker characters conditioned by two genes, dark green fruit of dgdg due to very high total chlorophyll content (18.19 mg/100 g fresh weight) and purple colour for anthocyanin pigment of Aft - gene. Lycopene content in ripe fruits of BCT-115 dgdg (6.22 mg/100 g fresh weight) was 62.41% higher compared to Alisa Craig Aft / Aft while that of Alisa Craig AftAft had high anthocyanin content (14.51 mg/ 100 g fresh weight) contrary to nil in BCT 115 dgdg . Segregation pattern confirmed the expected 3:1 ratio for single dominant Aft gene and 1:3 ratio for single recessive dg gene. The F 2 segregates could be grouped into 5 phenotypic classes in the ratio of 6:3:3:3:1 instead of usual 9:3:3:1 in digenic segregation due to variable expression of anthocyanin in the homozygous ( Aft Aft ) and heterozygous ( Aft aft ) genotype. Of the selfed progeny of the 34 desirable F 2 segregates (dark green fruit with intense anthocyanin pigmentation) clustered in the Group-1 family (F 3 population), 9 segregates bred true confirming the presence of two targeted genes in homozygous condition, AftAft dgdg , the “Purple tomato line”. This breeding approach proved to be a viable option to develop a new variety “Purple tomato” with unique combination of high lycopene (6.13 mg/100 g fresh weight), anthocyanin (20.73 mg/100 g fresh weight) and ascorbic acid (31.56 mg/100 g fresh weight) contents. [ABSTRACT FROM AUTHOR]

Summary: Genomic imprinting is a conspicuous feature of the endosperm, a triploid tissue nurturing the embryo and synchronizing angiosperm seed development. An unknown subset of imprinted genes (IGs) is critical for successful seed development and should have highly conserved functions. Recent genome‐wide studies have found limited conservation of IGs among distantly related species, but there is a paucity of data from closely related lineages. Moreover, most studies focused on model plants with nuclear endosperm development, and comparisons with properties of IGs in cellular‐type endosperm development are lacking. Using laser‐assisted microdissection, we characterized parent‐specific expression in the cellular endosperm of three wild tomato lineages (Solanum section Lycopersicon). We identified 1025 candidate IGs and 167 with putative homologs previously identified as imprinted in distantly related taxa with nuclear‐type endosperm. Forty‐two maternally expressed genes (MEGs) and 17 paternally expressed genes (PEGs) exhibited conserved imprinting status across all three lineages, but differences in power to assess imprinted expression imply that the actual degree of conservation might be higher than that directly estimated (20.7% for PEGs and 10.4% for MEGs). Regardless, the level of shared imprinting status was higher for PEGs than for MEGs, indicating dissimilar evolutionary trajectories. Expression‐level data suggest distinct epigenetic modulation of MEGs and PEGs, and gene ontology analyses revealed MEGs and PEGs to be enriched for different functions. Importantly, our data provide evidence that MEGs and PEGs interact in modulating both gene expression and the endosperm cell cycle, and uncovered conserved cellular functions of IGs uniting taxa with cellular‐ and nuclear‐type endosperm. [ABSTRACT FROM AUTHOR]

In a previous study, when resistant tomato plants (cv. Motelle) carrying the Sm gene were challenged with S. lycopersici , the SLNLC1 gene was significantly upregulated. In this study, to verify the function of the SLNLC1 gene response to disease resistance against S. lycopersici , virus-induced gene silencing (VIGS) was used to downregulate the expression level of the SLNLC1 gene in resistant tomato plants inoculated with S. lycopersici . After inoculation with S. lycopersici , a susceptible phenotype was observed in the silenced SLNLC1 -resistant plants. Through microscopy, impaired hypersensitive response (HR) and decreased ROS accumulation were also observed in the silenced SLNLC1 plants. In addition, the production of lignin and callose were decreased in the silenced SLNLC1 plants. Taken together, these results indicated that silencing the SLNLC1 gene attenuated the resistance of tomato plants resistant to S. lycopersici. [ABSTRACT FROM AUTHOR]

We assembled three complete mitochondrial genomes (mitogenomes), two of Solanum lycopersicum and one of Solanum pennellii, and analyzed their intra- and interspecific variations. The mitogenomes were 423,596–446,257 bp in length. Despite numerous rearrangements between the S. lycopersicum and S. pennellii mitogenomes, over 97% of the mitogenomes were similar to each other. These mitogenomes were compared with plastid and nuclear genomes to investigate genetic material transfers among DNA-containing organelles in tomato. In all mitogenomes, 9,598 bp of plastome sequences were found. Numerous nuclear copies of mitochondrial DNA (NUMTs) and plastid DNA (NUPTs) were observed in the S. lycopersicum and S. pennellii nuclear genomes. Several long organellar DNA fragments were tightly clustered in the nuclear genome; however, the NUMT and NUPT locations differed between the two species. Our results demonstrate the recent occurrence of frequent endosymbiotic gene transfers in tomato genomes. [ABSTRACT FROM AUTHOR]

Saline stress severely affects the growth and productivity of plants. The activation of hormonal signaling cascades and reactive oxygen species (ROS) in response to salt stress are important for cellular detoxification. Jasmonic acid (JA) and the enzyme SOD (superoxide dismutase), are well recognized markers of salt stress in plants. In this study, the application of chitosan-polyvinyl alcohol hydrogels (Cs-PVA) and copper nanoparticles (Cu NPs) on the growth and expression of defense genes in tomato plants under salt stress was evaluated. Our results demonstrate that Cs-PVA and Cs-PVA + Cu NPs enhance plant growth and also promote the expression of JA and SOD genes in tomato (Solanum lycopersicum L.), under salt stress. We propose that Cs-PVA and Cs-PVA + Cu NPs mitigate saline stress through the regulation of oxidative and ionic stress. [ABSTRACT FROM AUTHOR]

Plants are sessile organisms, hence to face environmental constrains they developed strategies that rely on the activation of stress-response genes under the control of specific transcription factors. The plant hormone ethylene mediates physiological, developmental and stress responses through the activation of Ethylene Response Factors (ERFs) which belong to a large multigene family of transcription factors. While an increasing number of studies supports the involvement of ERFs in abiotic stress responses, so far the specific role of ERF family members in different abiotic stress conditions remains unexplored. The present work investigates the expression profile of a set of ERFs, representative of different ERF types, in tomato plants subjected to cold, heat, salt, drought and flooding conditions. The study revealed that a group of ERFs is preferentially associated with cold and heat stress responses while another set is expressed in response to salt, water and flooding stresses. Transactivation assays indicated that ERFs can regulate the expression of abiotic stress genes regardless of whether or not they harbor conserved GCC or DRE cis -elements in their promoter region. The outcome of the study provides clue on which ERFs should be targeted when aiming to improve adaptation to a particular stress type. [ABSTRACT FROM AUTHOR]

The impact of extreme and sustained high temperatures on plant growth has become increasingly prominent. Heat shock cognate 70-kDa proteins play an important role in plant heat tolerance. In this study, we identified and characterized the tomato ortholog of LrgB ( SlLrgB ), and demonstrate that it interacts with Hsc70.1. Similar to other genes that encode chloroplast-localized proteins, the expression of SlLrgB is upregulated in green tissues and suppressed by heat shock. Functional analyses utilizing transgenic plants indicate that SlLrgB contributes to chlorophyll metabolism. Both the overexpression and the RNA interference-mediated suppression of SlLrgB led to chlorotic leaves, reduced plant height, smaller size and decreases in pigment levels in ripening fruits. However, the starch levels in the SlLrgB -RNAi lines were significantly increased and the heat tolerance of SlLrgB -RNAi was obvious elevated. Downregulating the expression of Hsc70.1 by VIGS in tomato led to retarded growth, chlorotic leaves, and increased expression of SlLrgB. Based on these data, we suggest that SlLrgB regulates chlorophyll metabolism and the assimilation and partitioning of carbon. We also suggest that Hsc70.1 and SlLrgB contribute to heat tolerance and that Hsc70.1 negatively regulates SlLrgB . [ABSTRACT FROM AUTHOR]

Cell wall metabolism during fruit ripening is a highly organized process that involves complex interplay among various cell wall hydrolases. Among these cell wall hydrolases, β-galactosidase has been identified to participate in cell wall metabolism via its ability to catalyze galactosyl metabolism from the large and complex side chains of cell walls. In this study, the galactose content in the pericarp increased during persimmon fruit ripening, but cell wall galactosyl residues decreased, indicating a relationship between galactose metabolism and persimmon fruit ripening. Expression of a previously isolated β-galactosidase gene, DkGAL1 , increased 25.01-fold during fruit ripening. Heterologous expression of DkGAL1 under the CaMV 35S promoter in tomato accelerated on-plant and postharvest fruits ripening. The fruit firmness of one of transgenic line, OE-18, was 23.83% lower than that of WT at the breaker stage. The transgenic fruits produced more ethylene by promoting the expression of ethylene synthesis-related genes and cell wall degradation-related genes. Overexpression of DkGAL1 in tomato also reduced cell-to-cell adhesion and promoted both wider intercellular spaces and less cell compaction in transgenic fruit structures. Moreover, DkGAL1 was involved in seed germination and radicle elongation in transgenic tomato seeds. These results confirm the role of DkGAL1 in fruit ripening and suggest that this gene alters galactose metabolism in the fruit, which can promote ripening and reduce cellular adhesion. In addition, the role of DkGAL1 is not limited to fruit softening; DkGAL1 was also involved in seed germination and radicle elongation in transgenic tomato seeds. [ABSTRACT FROM AUTHOR]

Glycine-rich RNA-binding proteins (GR-RBPs) are involved in RNA processing and also some of them are output signals of the circadian clock. In tomato, one GR-RBP gene family (LeGRP1) is composed by three highly homologous genes (LeGRP1a-c); each one rendering three transcriptional products: the un-spliced pre-RNA (preLegrp1a-c), the mature mRNA (mLegrp1a-c) and the alternatively spliced mRNA (asLegrp1a-c). To get insight into their regulation and impact on RNA metabolism in fruits, Solanum lycopersicum cv. Micro-Tom was transformed with preLeGRP1a fused to the polygalacturonase promoter, which drives expression to fruits from the mature green stage. Our results demonstrated a complex positive regulation of LeGRPs, in which LeGRP1a overexpression led to the induction of the others LeGRP1 members. Even though the LeGRP1 transcription and the content of three LeGRPs proteins were affected, the overall LeGRP protein circadian rhythm profile was similar in transgenic and wild type (WT) fruits. However, when the fruits were kept at a chilling temperature after harvest, total protein content was significantly higher in transgenic than in WT fruits, and the content of some free amino acids was modified. The results obtained suggest a probable role of LeGRP1s: structural rearrangements and/or stabilization of mRNA to allow efficient processing of fruits under cold conditions. [ABSTRACT FROM AUTHOR]

The NAC (NAM, ATAF1/2, and CUC2) transcription factor family participates in responses to various kinds of environmental stimuli in plants. However, the roles of NAC protein in cold resistance, especially in the cold resistance of tomatoes, are not completely understood. This study examined the roles of a tomato (Solanum lycopersicum) NAC transcription factor (SlNAC35) in resisting chilling using transgenic tomatoes. GUS staining and expression analysis revealed that SlNAC35 expression was induced at 4 °C, thereby suggesting its involvement in plant responses to chilling stress. Moreover, transgenic lines over-expressing SlNAC35 exhibited high chlorophyll content, fresh mass, and low accumulation of reactive oxygen species and membrane damage under chilling stress. These results indicated that SlNAC35 overexpression enhanced the chilling tolerance of transgenic tomatoes. High expressions of cold tolerance markers SlCOR518 and SlCOR413IM1 were observed under chilling stress in transgenic lines. This observation suggested that SlNAC35 overexpression enhanced the chilling tolerance of transgenic lines by involving the c-repeat binding factor-cold stress response (CBF-COR) signaling pathway and by regulating SlCOR expression. [ABSTRACT FROM AUTHOR]

Vitamin B6 (VitB6) is an essential cofactor for >140 biochemical reactions. Also, VitB6 is a potent antioxidant and helps plants cope with both biotic and abiotic stress conditions. However, the role of VitB6 in plant disease resistance has yet to be confirmed using molecular biology approaches. Here, we analyzed the expression patterns of VitB6 biosynthetic genes, including the de novo ( PDX1 [ PDX1.2 and 1.3 ] and PDX2 ) and the salvage ( SOS4 ) pathways during the response to Erwinia carotovora subsp. carotovora . By quantitative PCR, we found that the most significant upregulation in the transcript profile of PDX2 , which showed a 9.2-fold increase in expression at 12 h post inoculation (hpi) compared to 24–48 hpi. We also detected significant upregulation of PDX1.2 and PDX1.3 , which were 6.6- and 4.3-fold upregulated at 24 hpi compared to 12 hpi, while SOS4 showed only low-level expression. Also, at 24 hpi, a significant increase in superoxide dismutase, catalase, peroxidase, and polyphenol oxidase activities was observed in plants. Our findings confirm that the expression of de novo and salvage pathway genes is induced by E. carotovora and that this plays an important role in the regulation of defense response by modulating cellular antioxidant capacity. [ABSTRACT FROM AUTHOR]

Early blight caused by Alternaria solani is a major production constraint in the cultivation of tomato. Success through conventional breeding has been elusive for the development of resistant varieties from wild sources due to transfer of several unacceptable quality and yield attributes from them. Therefore, transformation of tomato with gene encoding pathogen cell wall degrading enzyme chitinase isolated from Trichoderma harzianum was carried out and transgenic plants were generated. Two homozygous lines N3-5 and N8-4 with 2.5× and 4.0× higher chitinase expression, respectively, were shortlisted and were assessed at fully grown (mature) stage in net house bioassays with Alternaria solani through in vitro detached leaf as well as challenging with fungal inoculum and were compared with disease reaction at the seedling stage. Screening at the seedling stage in T2 generation in the earlier study revealed that one of the transgenic lines N3-5 exhibited moderately resistant reaction with significant reduction in lesion size as compared to the untransformed control under in vitro detached leaf bioassay and challenge inoculation assays. Screening at fully grown stage through challenge inoculation in T3 and T4 generation recorded an intermediate disease reaction (between susceptible and resistant) with significantly lower disease incidence of early blight as compared to susceptible reaction of control plants. In the in vitro detached leaf assay, these lines exhibited significantly reduced lesion size compared to control at 65 days but this difference was eliminated at 74 and 85 days signifying thereby the role of plant age. [ABSTRACT FROM AUTHOR]

Choline monooxygenase (CMO) is a key enzyme involved in betaine synthesis and our preliminary work has shown that the SlCMO gene promoter (pC5: − 267 to + 128 base pair), cloned from Suaeda liaotungensis, is salt-inducible. In the present study, pC5-SlCMO was transferred into tomato (Solanum lycopersicon L. ‘Micro-Tom’) plants via Agrobacterium mediation. Homozygous transgenic plants were selected using quantitative real-time polymerase chain reaction. The expression of SlCMO in pC5-SlCMO transgenic plants was induced by salinity. Under salt tolerance, betaine content, chlorophyll content, and net photosynthetic rate were higher in transgenic plants than in wild-type (WT) plants. Proline content was lower in transgenic plants than in WT plants. Under normal conditions, seed germination, length of the whole plant, dry weight, and fruit products of transgenic plants were the same as in WT plants. These results demonstrated that the pC5 promoter can drive increased expression of SlCMO in transgenic tomato plants under salt stress and increase salt tolerance without affecting plant growth and yield. [ABSTRACT FROM AUTHOR]

Eloquent information about the genetic basis of inheritance is important for any breeding program. Therefore, a diallel study was conducted under the influence of tomato leaf curl virus (TLCV) disease, using the eleven advanced lines of tomato. Firstly, information regarding percent disease index (PDI) was determined via artificial screening with viruliferous whiteflies. Later, these lines were crossed in a half diallel mating design to produce fifty-five one-way hybrids. These hybrids and parental genotypes were evaluated for morphological and biochemical traits under open field conditions. Using the Griffing approach (Method II and Model I), the basis of the inheritance of traits was determined. Furthermore, a Bayesian model was applied to the total yield descriptor. Correlation data indicated that total yield was not correlated with any other trait. The significant general combining ability (GCA) and specific combining ability (SCA) values indicate exploitable genetic variation. The broad-sense heritability values were larger than narrow-sense heritability, showing that selection will be efficient for the improvement of these traits. Hybrid combinations H23, H42 and H49 can be considered efficient for the selection of multiple traits, including yield. Overall, this study provides useful information regarding the genetics of important traits of tomato under TLCV infestation. [ABSTRACT FROM AUTHOR]

Auxin and strigolactone (SL) are two important phytohormones involved in shoot branching and morphology. Tomato (Solanum lycopersicum L.), a member of the Solanaceae family, is one of the most popular food crops with high economic value in the world. To seek a better understanding of the responses to exogenous hormones, transcriptome analyses of the tomato shoots treated with exogenous auxin and SL, separately or together, were performed. A total of 2326, 260 and 1379 differential expressed genes (DEGs) were identified under the IAA, GR24 and IAA+GR24 treatments, respectively. Network analysis pointed out two enriched interaction clusters, including “ethylene biosynthesis” and “photosynthesis”. Several ethylene biosynthesis and metabolism-related genes were up-regulated under both IAA and IAA+GR24 treatments, suggesting their involvement in the regulation of ethylene biosynthesis. Besides, auxin-SLs-triggered the expression of several CAB genes may lead to systemic increases in the induction of photosynthesis. Several auxin-activated metabolic pathways could be reduced by the GR24 treatment, indicated that the crosstalk between auxin and SLs may be involved in the metabolic regulation of tomato. Further analysis showed that SLs affect the responses of tomato shoots to auxin by inducing the expression of a series of auxin downstream genes. On the other hand, auxin regulated the biosynthesis of SLs by affecting the genes in the “Carotenoid biosynthesis” pathway. Our data will give us an opportunity to reveal the crosstalk between auxin and SLs in the shoots of tomato. [ABSTRACT FROM AUTHOR]

Despite a range of initiatives to reduce global carbon emission, the mean global temperature is increasing due to climate change. Since rising temperatures pose a serious threat of food insecurity, it is important to further explore important biological molecules that can confer thermotolerance to plants. Recently, melatonin has emerged as a universal abiotic stress regulator that can enhance plant tolerance to high temperature. Nonetheless, such regulatory roles of melatonin were unraveled mainly by assessing the effect of exogenous melatonin on plant tolerance to abiotic stress. Here, we generated melatonin deficient tomato plants by silencing of a melatonin biosynthetic gene, CAFFEIC ACID O-METHYLTRANSFERASE 1 (COMT1), to unveil the role of endogenous melatonin in photosynthesis under heat stress. We examined photosynthetic pigment content, leaf gas exchange, and a range of chlorophyll fluorescence parameters. The results showed that silencing of COMT1 aggravated heat stress by inhibiting both the light reactions and the carbon fixation reactions of photosynthesis. The photosynthetic pigment content, light absorption flux, trapped energy flux, energy dissipation, density of active reaction center per photosystem II (PSII) cross-section, the photosynthetic electron transport rate, the maximum photochemical efficiency of PSII photochemistry, and the rate of CO2 assimilation all decreased in COMT1-silenced plants compared with that of non-silenced plants particularly under heat stress. However, exogenous melatonin alleviated heat-induced photosynthetic inhibition in both genotypes, indicating that melatonin is essential for maintaining photosynthetic capacity under stressful conditions. These findings provide genetic evidence on the vital role of melatonin in photosynthesis and thus may have useful implication in horticultural crop management in the face of climate change. [ABSTRACT FROM AUTHOR]

Abstract: The abscission of tomato leaves occurs in the petiole abscission zone, and its late stage includes two spatially divided processes: cell separation and programmed cell death (PCD). Both of these processes are regulated by ethylene. The last step in ethylene biosynthesis is conversion of 1‐aminocyclopropane‐1‐carboxylic acid to ethylene, which is catalysed by the enzyme 1‐aminocyclopropane‐1‐carboxylate oxidase (ACO); however, the location of ACO in the leaf petiole abscission zone is not known. The tomato gene LeLX encodes ribonuclease LX, which is a marker for PCD and is induced by ethylene during abscission, but its association with ACO has not been explored. In a tomato transgenic line 1‐7 with inhibited expression of LeLX showing delayed leaf abscission, the morphology and ultrastructure of the leaf petiole abscission zone was examined. In this zone of the cv.’VF36’ and of a transgenic line 1‐7, spatiotemporal differences in expression of LeACO1 and LeACO4 were analysed and ACO protein was detected immunohistochemically. In comparison to wild‐type plants, there were no obvious morphological and ultrastructural features in the abscission zone of plants of a transgenic line 1‐7 before and after abscission induction. LeACO1 expression was low before abscission induction, and increased 24 h after induction, although with no apparent spatial pattern. In contrast, LeACO4 was expressed before abscission induction, and its transcript level declined 24 h after induction on the distal side of the abscission zone fracture. In the LeLX‐inhibited transgenic line, there were no significant differences in LeACO1 and LeACO4 expression in the petiole abscission zone, in comparison to wild‐type plants. In addition, the ACO protein was immunolocalised to the vascular tissues that traverse the petiole abscission zone in plants of wild type and of a transgenic line 1‐7; and additionally in the plane of future abscission zone fracture of transgenic‐line plants. The results suggest temporal differential expression of the LeACO genes in tomato leaf petioles and vascular localisation of ACO1 protein. Additionally, the results indicate that expression of LeACO genes is not affected by suppression of the LeLX expression. [ABSTRACT FROM AUTHOR]

The SEPALLATA (SEP) MADS-box transcription factors play essential roles in reproductive growth, especially in floral organ differentiation. Here, SlCMB1 , a tomato SEP MADS-box gene, was isolated. SlCMB1 is noticeably expressed in inflorescences and flowers. Its transcript levels were higher in sepals than in other floral organs and decreased during sepal development. Tomato plants with reduced SlCMB1 mRNA levels displayed longer, branched and indeterminate inflorescences that exhibited a transition from reproductive to vegetative growth and enlarged and abnormally fused sepals. The transcript levels of genes known to regulate the development of inflorescence architecture and sepal size in tomato were dramatically changed. In addition, the expression levels of cell elongation-related and gibberellin biosynthetic genes also showed significant differences between the transgenic lines and the wild type, and the GA content of the peduncle in the transgenic lines was higher than that in the wild type. Yeast two-hybrid assay showed that SlCMB1 could interact individually with MC, J, AP2a and SlMBP21. Overall, our results indicate that SlCMB1 is an important regulator involved in the development of inflorescence architecture and sepal size in tomato plants. [ABSTRACT FROM AUTHOR]

Jasmonic acid (JA) has been mostly studied in responses to biotic stresses, such as herbivore attack and pathogenic infection. More recently, the involvement of JA in abiotic stresses including salinity was highlighted; yet, its role in salt stress remained unclear. In the current study, we compared the physiological and biochemical responses of wild-type (WT) tomato ( Solanum lycopersicum ) cv Castlemart and its JA-deficient mutant defenseless-1 ( def-1 ) under salt stress to investigate the role of JA. Plant growth, photosynthetic pigment content, ion accumulation, oxidative stress-related parameters, proline accumulation and total phenolic compounds, in addition to both enzymatic and non-enzymatic antioxidant activities, were measured in both genotypes after 14 days of 100 mM NaCl treatment. Although we observed in both genotypes similar growth pattern and sodium, calcium and potassium levels in leaves under salt stress, def-1 plants exhibited a more pronounced decrease of nitrogen content in both leaves and roots and a slightly higher level of sodium in roots compared to WT plants. In addition, def-1 plants exposed to salt stress showed reactive oxygen species (ROS)-associated injury phenotypes. These oxidative stress symptoms in def-1 were associated with lower activity of both enzymatic antioxidants and non-enzymatic antioxidants. Furthermore, the levels of the non-enzymatic ROS scavengers proline and total phenolic compounds increased in both genotypes exposed to salt stress, with a higher amount of proline in the WT plants. Overall the results of this study suggest that endogenous JA mainly enhanced tomato salt tolerance by maintaining ROS homeostasis. [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]

Agronomic performance of transgenic tomato overexpressing functional genes has rarely been investigated in the field. In an attempt to improve low-phosphate (P) stress tolerance of tomato ( Solanum lycopersicum ) plants and promote tomato fruit production in the field, an expression vector containing cDNA to an Arabidopsis 14-3-3 protein, General Regulatory Factor 9 ( GRF9 ), driven by a cauliflower mosaic virus 35S promoter, was transferred into tomato plants. Transgenic expression of GRF9 was ascertained by quantitative real-time PCR analysis. The degree of low-P tolerance in transgenic plants was found to be significantly greater than that in wild-type plants, and reflected in improved root development and enhanced P content under hydroponic conditions. For transgenic tomato, roots had higher P uptake, as evidenced by tissue P content and relative expression of the genes LePT1 and LePT2 in both normal and low-P hydroponic solutions. GRF9 overexpressors had greatly enhanced proton extrusion from roots and heightened activity of the plasma-membrane H + -ATPase (PM H + -ATPase) in roots under low-P hydroponic conditions. Thus, in addition to enhanced root development, higher expression of genes coding for phosphate transporters and improved capacity for acidification in the rhizosphere emerged as key mechanisms underpinning improved P acquisition in transgenic tomato plants in soil. Subsequent field trials measuring tomato fruit production at two P levels, indicated that GRF9 can indeed improve total tomato production and may play a role in early fruit maturity. Our results suggest that the heterologous Arabidopsis GRF9 gene can confer resistance to P deficiency in transgenic tomato plants and promote fruit production. [ABSTRACT FROM AUTHOR]

Circadian period and phase of cultivated tomato (Solanum lycopersicum) were changed during domestication, likely adapting the species to its new agricultural environments. Whereas the delayed circadian phase is mainly caused by allelic variation of EID1, the genetic basis of the long circadian period has remained elusive. Here we show that a partial deletion of the clock gene LNK2 is responsible for the period lengthening in cultivated tomatoes. We use resequencing data to phylogenetically classify hundreds of tomato accessions and investigate the evolution of the eid1 and Ink2 mutations along successive domestication steps. We reveal signatures of selection across the genomic region of LNK2 and different patterns of fixation of the mutant alleles. Strikingly, LNK2 and EID1 are both involved in light input to the circadian clock, indicating that domestication specifically targeted this input pathway. In line with this, we show that the clock deceleration in the cultivated tomato is light-dependent and requires the phytochrome B1 photoreceptor. Such conditional variation in circadian rhythms may be key for latitudinal adaptation in a variety of species, including crop plants and livestock. [ABSTRACT FROM AUTHOR]

Summary: Current breeding relies mostly on random mutagenesis and recombination to generate novel genetic variation. However, targeted genome editing is becoming an increasingly important tool for precise plant breeding. Using the CRISPR‐Cas system combined with the bean yellow dwarf virus rolling circle replicon, we optimized a method for targeted mutagenesis and gene replacement in tomato. The carotenoid isomerase (CRTISO) and phytoene synthase 1 (PSY1) genes from the carotenoid biosynthesis pathway were chosen as targets due to their easily detectable change of phenotype. We took advantage of the geminiviral replicon amplification as a means to provide a large amount of donor template for the repair of a CRISPR‐Cas‐induced DNA double‐strand break (DSB) in the target gene, via homologous recombination (HR). Mutagenesis experiments, performed in the Micro‐Tom variety, achieved precise modification of the CRTISO and PSY1 loci at an efficiency of up to 90%. In the gene targeting (GT) experiments, our target was a fast‐neutron‐induced crtiso allele that contained a 281‐bp deletion. This deletion was repaired with the wild‐type sequence through HR between the CRISPR‐Cas‐induced DSB in the crtiso target and the amplified donor in 25% of the plants transformed. This shows that efficient GT can be achieved in the absence of selection markers or reporters using a single and modular construct that is adaptable to other tomato targets and other crops. [ABSTRACT FROM AUTHOR]

Several expression systems for multiple guide RNA (gRNAs) have been developed in the CRISPR/Cas9 (clustered regularly interspaced short palindromic repeats/CRISPR associated protein 9) system to induce multiple-gene modifications in plants. Here, we evaluated mutation efficiencies in the tomato genome using multiplex CRISPR/Cas9 vectors consisting of various Cas9 expression promoters with multiple gRNA expression combinations. In transgenic tomato calli induced with these vectors, mutation patterns varied depending on the promoters used to express Cas9. By using the tomato ELONGATION FACTOR-1α (SlEF1α) promoter to drive Cas9, occurrence of various types of mutations with high efficiency was detected in the tomato genome. Furthermore, sequence analysis showed that the majority of mutations using the multiplex system with the SlEF1α promoter corresponded to specific mutation pattern of deletions produced by self-ligation at two target sites of CRISPR/Cas9 with low mosaic mutations. These results suggest that optimizing the Cas9 expression promoter used in CRISPR/Cas9-mediated mutation improves multiplex genome editing, and could be used effectively to disrupt functional domains precisely in the tomato genome. [ABSTRACT FROM AUTHOR]

Summary: Chloroplast development and chlorophyll(Chl)metabolism in unripe tomato contribute to the growth and quality of the fruit, however these mechanisms are poorly understood. In this study, we initially investigated seven homeobox‐containing transcription factors (TFs) with specific ripening‐associated expression patterns using virus‐induced gene silencing (VIGS) technology and found that inhibiting the expression of one of these TFs, BEL1‐LIKE HOMEODOMAIN11 (SlBEL11), significantly increased Chl levels in unripe tomato fruit. This enhanced Chl accumulation was further validated by generating stable RNA interference (RNAi) transgenic lines. RNA sequencing (RNA‐seq) of RNAi‐SlBEL11 fruit at the mature green (MG) stage showed that 48 genes involved in Chl biosynthesis, photosynthesis and chloroplast development were significantly upregulated compared with the wild type (WT) fruit. Genomic global scanning for Homeobox TF binding sites combined with RNA‐seq differential gene expression analysis showed that 22 of these 48 genes were potential target genes of SlBEL11 protein. These genes included Chl biosynthesis‐related genes encoding for protochlorophyllide reductase (POR), magnesium chelatase H subunit (CHLH) and chlorophyllide a oxygenase (CAO), and chloroplast development‐related genes encoding for chlorophyll a/b binding protein (CAB), homeobox protein knotted 2 (TKN2) and ARABIDOPSIS PSEUDO RESPONSE REGULATOR 2‐LIKE (APRR2‐like). Electrophoretic mobility shift assay (EMSA) and chromatin immunoprecipitation quantitative polymerase chain reaction (PCR) (ChIP‐qPCR) assays were employed to verify that SlBEL11 protein could bind to the promoters for TKN2, CAB and POR. Taken together, our findings demonstrated that SlBEL11 plays an important role in chloroplast development and Chl synthesis in tomato fruit. [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]

Fruit ripening is a complex biological process affecting fruit quality. In tomato the fruit ripening process is delicately regulated by transcription factors (TFs). Among these, the TOMATO AGAMOUS-LIKE 1 (TAGL1) gene plays an important role in both the development and ripening of fruit. In this study, the TAGL1 gene was successfully silenced by virus-induced gene silencing technology (VIGS), and the global gene expression and metabolites profiles of TAGL1-silenced fruits were analyzed by RNA-sequence analysis (RNA-seq) and liquid chromatography–mass spectrometry (LC-MS/MS). The TAGL1-silenced fruits phenotypically displayed an orange pericarp, which was in accordance with the results expected from the down-regulation of genes associated with carotenoid synthesis. Levels of several amino acids and organic acids were lower in the TAGL1-silenced fruits than in the wild-type fruits, whereas, α-tomatine content was greatly increased (more than 10-fold) in the TAGL1-silenced fruits compared to wild-type fruits. The findings of this study showed that TAGL1 not only regulates the ripening of tomato fruits, but also affects the synthesis and levels of nutrients in the fruit. [ABSTRACT FROM AUTHOR]

Background: Pollen development is central for plant reproduction and is assisted by changes of the transcriptome and proteome. At the same time, pollen development and viability is largely sensitive to stress, particularly to elevated temperatures. The transcriptomic and proteomic changes during pollen development and of different stages in response to elevated temperature was targeted to define the underlying molecular principles. Results: The analysis of the transcriptome and proteome of Solanum lycopersicum pollen at tetrad, post-meiotic and mature stage before and after heat stress yielded a decline of the transcriptome but an increase of the proteome size throughout pollen development. Comparison of the transcriptome and proteome led to the discovery of two modes defined as direct and delayed translation. Here, genes of distinct functional processes are under the control of direct and delayed translation. The response of pollen to elevated temperature occurs rather at proteome, but not as drastic at the transcriptome level. Heat shock proteins, proteasome subunits, ribosomal proteins and eukaryotic initiation factors are most affected. On the example of heat shock proteins we demonstrate a decoupling of transcript and protein levels as well as a distinct regulation between the developmental stages. Conclusions: The transcriptome and proteome of developing pollen undergo drastic changes in composition and quantity. Changes at the proteome level are a result of two modes assigned as direct and delayed translation. The response of pollen to elevated temperature is mainly regulated at the proteome level, whereby proteins related to synthesis and degradation of proteins are most responsive and might play a central role in the heat stress response of pollen. [ABSTRACT FROM AUTHOR]

Lectin receptor-like kinases (LecRLKs) are crucial factors in response to plant biotrophic pathogens. This study aimed to systemically analyze the effect of tomato LecRLKs on the responses to tomato yellow leaf curl virus (TYLCV) infection. A total of 93 tomato SlyLecRLK genes, including 69 G-type LecRLKs, 23 L-type LecRLKs, and one C-type LecRLK, were identified using genome-wide analysis and were clustered into three subfamilies using phylogenetic analysis. These SlyLecRLK genes were localized on the 12 tomato chromosomes with some gene duplication events. Transcriptomic analysis showed most of SlyLecRLKs were modulated by phytohormones and TYLCV infection. Gene ontology (GO) enrichment analysis showed all SlyLecRLK genes were associated with receptor protein kinase activity and protein modification. Quantitative RT-PCR (qRT-PCR) and gene interference validation showed SlyLecRLKs007, SlyLecRLKs036, and SlyLecRLKs044 genes were inhibited by TYLCV infection, and silencing of them were accompanied by accumulation of TYLCV DNA content. Subcellular localization of LecRLKs023 and LecRLKs044 on cell membranes was determined using a SlyLecRLK-containing A. tumefaciens strain GV3101. These results showed SlyLecRLK genes were key factors for TYLCV resistance, and also provided crucial information uncovering the protective effects of SlyLecRLKs on resisting TYLCV infection in tomato. [ABSTRACT FROM AUTHOR]

Summary: Abiotic stresses are a major cause of crop loss. Ascorbic acid (AsA) promotes stress tolerance by scavenging reactive oxygen species (ROS), which accumulate when plants experience abiotic stress. Although the biosynthesis and metabolism of AsA are well established, the genes that regulate these pathways remain largely unexplored. Here, we report on a novel regulatory gene from tomato (Solanum lycopersicum) named SlZF3 that encodes a Cys2/His2‐type zinc‐finger protein with an EAR repression domain. The expression of SlZF3 was rapidly induced by NaCl treatments. The overexpression of SlZF3 significantly increased the levels of AsA in tomato and Arabidopsis. Consequently, the AsA‐mediated ROS‐scavenging capacity of the SlZF3‐overexpressing plants was increased, which enhanced the salt tolerance of these plants. Protein–protein interaction assays demonstrated that SlZF3 directly binds CSN5B, a key component of the COP9 signalosome. This interaction inhibited the binding of CSN5B to VTC1, a GDP‐mannose pyrophosphorylase that contributes to AsA biosynthesis. We found that the EAR domain promoted the stability of SlZF3 but was not required for the interaction between SlZF3 and CSN5B. Our findings indicate that SlZF3 simultaneously promotes the accumulation of AsA and enhances plant salt‐stress tolerance. [ABSTRACT FROM AUTHOR]

Previous studies suggest that GRAS transcription factors act as essential regulators, not only in plant growth and development but also in response to biotic and abiotic stresses. Recently, 53 GRAS proteins have been identified, but only a few of them have been functionally studied in tomato. Here, we isolated a novel GRAS transcription factor SlGRAS26, its down-regulation generated pleiotropic phenotypes, including reduced plant height with more lateral shoots, internode length, leaf size, even leaflets, accelerated flowering transition and decreased trichome number. Transcription analysis showed that down-regulation of SlGRAS26 altered vegetative growth by suppressing gibberellin (GA) biosynthesis genes and activating the GA inactivating genes, thereby reducing endogenous GA content in transgenic plants. SlGRAS26 may regulate the initiation of lateral buds by regulating the expression of Blind (BL) and BRC1b . The earlier initiation of flower buds in transgenic lines may be controlled by significant up-regulation of SFT , CO1 , SBP3 , SBP13 , and SBP15 genes, related to flowering time. These results demonstrate that SlGRAS26 may play a vital role in the initiation of lateral and inflorescence meristems in tomato. [ABSTRACT FROM AUTHOR]

Key message: In vitro tomato pollen tubes show a cytoplasmic calcium gradient that oscillates with the same period as growth.Abstract: Pollen tube growth requires coordination between the tip-focused cytoplasmic calcium concentration ([Ca2+]cyt) gradient and the actin cytoskeleton. This [Ca2+]cyt gradient is necessary for exocytosis of small vesicles, which contributes to the delivery of new membrane and cell wall at the pollen tube tip. The mechanisms that generate and maintain this [Ca2+]cyt gradient are not completely understood. Here, we studied calcium dynamics in tomato (Solanum lycopersicum) pollen tubes using transgenic tomato plants expressing the Yellow Cameleon 3.6 gene under the pollen-specific promoter LAT52. We use tomato as an experimental model because tomato is a Solanaceous plant that is easy to transform, and has an excellent genomic database and genetic stock center, and unlike Arabidopsis, tomato pollen is a good system to do biochemistry. We found that tomato pollen tubes showed an oscillating tip-focused [Ca2+]cyt gradient with the same period as growth. Then, we used a pharmacological approach to disturb the intracellular Ca2+ homeostasis, evaluating how the [Ca2+]cyt gradient, pollen germination and in vitro pollen tube growth were affected. We found that cyclopiazonic acid (CPA), a drug that inhibits plant PIIA-type Ca2+-ATPases, increased [Ca2+]cyt in the subapical zone, leading to the disappearance of the Ca2+ oscillations and inhibition of pollen tube growth. In contrast, 2-aminoethoxydiphenyl borate (2-APB), an inhibitor of Ca2+ released from the endoplasmic reticulum to the cytoplasm in animals cells, completely reduced [Ca2+]cyt at the tip of the tube, blocked the gradient and arrested pollen tube growth. Although both drugs have antagonistic effects on [Ca2+]cyt, both inhibited pollen tube growth triggering the disappearance of the [Ca2+]cyt gradient. When CPA and 2-APB were combined, their individual inhibitory effects on pollen tube growth were partially compensated. Finally, we found that GsMTx-4, a peptide from spider venom that blocks stretch-activated Ca2+ channels, inhibited tomato pollen germination and had a heterogeneous effect on pollen tube growth, suggesting that these channels are also involved in the maintenance of the [Ca2+]cyt gradient. All these results indicate that tomato pollen tube is an excellent model to study calcium dynamics. [ABSTRACT FROM AUTHOR]

Key message: An NB-LRR gene, TYNBS1, was isolated from Begomovirus-resistance locus Ty-2. Transgenic plant analysis revealed that TYNBS1 is a functional resistance gene. TYNBS1 is considered to be synonymous with Ty-2.Abstract: Tomato yellow leaf curl disease caused by Tomato yellow leaf curl virus (TYLCV) is a serious threat to tomato (Solanum lycopersicum L.) production worldwide. A Begomovirus resistance gene, Ty-2, was introduced into cultivated tomato from Solanum habrochaites by interspecific crossing. To identify the Ty-2 gene, we performed genetic analysis. Identification of recombinant line 3701 confirmed the occurrence of a chromosome inversion in the Ty-2 region of the resistant haplotype. Genetic analysis revealed that the Ty-2 gene is linked to an introgression encompassing two markers, SL11_25_54277 and repeat A (approximately 200 kb). Genomic sequences of the upper and lower border of the inversion section of susceptible and resistant haplotypes were determined. Two nucleotide-binding domain and leucine-rich repeat-containing (NB-LRR) genes, TYNBS1 and TYNBS2, were identified around the upper and lower ends of the inversion section, respectively. TYNBS1 strictly co-segregated with TYLCV resistance, whereas TYNBS2 did not. Genetic introduction of genomic fragments containing the TYNBS1 gene into susceptible tomato plants conferred TYLCV resistance. These results demonstrate that TYNBS1 is a functional resistance gene for TYLCV, and is synonymous with the Ty-2 gene. [ABSTRACT FROM AUTHOR]

Abstract: GRAS family transcription factors are involved in multiple biological processes in plants. Here, we report that GRAS2 plays a vital role in regulating fruit weight in tomato (Solanum lycopersicum). We establish that the expression of GRAS2 was elevated in ovaries and maintained at a constant level in fertilized ovules. Reduction of GRAS2 expression in transgenic plants reduced fruit weight through modulating ovary growth and cell size. At the metabolic level, downregulation of GRAS2 decreased activities of the gibberellic acid biosynthesis and signal transduction pathways, leading to insufficient levels of active gibberellic acid during the initial ovary development of tomato. Moreover, genotypic diversity of GRAS2 was consistent with the molecular basis of fruit weight evolution, suggesting that GRAS2 contributes to the molecular basis of the evolution of fruit weight in tomato. Collectively, these findings enhance our understanding of GRAS2 functions, in fruit development of tomato, and demonstrate a strong association between the GRAS gene family and fruit development. [ABSTRACT FROM AUTHOR]