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Background: During maturation seeds acquire several physiological traits to enable them to survive drying and disseminate the species. Few studies have addressed the regulatory networks controlling acquisition of these traits at the tissue level particularly in endospermic seeds such as tomato, which matures in a fully hydrated environment and does not undergo maturation drying. Using temporal RNA-seq analyses of the different seed tissues during maturation, gene network and trait-based correlations were used to explore the transcriptome signatures associated with desiccation tolerance, longevity, germination under water stress and dormancy., Results: During maturation, 15,173 differentially expressed genes were detected, forming a gene network representing 21 expression modules, with 3 being specific to seed coat and embryo and 5 to the endosperm. A gene-trait significance measure identified a common gene module between endosperm and embryo associated with desiccation tolerance and conserved with non-endospermic seeds. In addition to genes involved in protection such LEA and HSP and ABA response, the module included antioxidant and repair genes. Dormancy was released concomitantly with the increase in longevity throughout fruit ripening until 14 days after the red fruit stage. This was paralleled by an increase in SlDOG1-2 and PROCERA transcripts. The progressive increase in seed vigour was captured by three gene modules, one in common between embryo and endosperm and two tissue-specific. The common module was enriched with genes associated with mRNA processing in chloroplast and mitochondria (including penta- and tetratricopeptide repeat-containing proteins) and post-transcriptional regulation, as well several flowering genes. The embryo-specific module contained homologues of ABI4 and CHOTTO1 as hub genes associated with seed vigour, whereas the endosperm-specific module revealed a diverse set of processes that were related to genome stability, defence against pathogens and ABA/GA response genes., Conclusion: The spatio-temporal co-expression atlas of tomato seed maturation will serve as a valuable resource for the in-depth understanding of the dynamics of gene expression associated with the acquisition of seed vigour at the tissue level.

This study was conducted to investigate the involvement of antifreeze proteins (AFPs; type I and III) in the germination mechanism of tomato seeds under low temperature stress. Germination of the seeds grown at a room temperature (25°C) was observed on 5 days after sowing (DAS), while all seeds exposed to a low temperature started to germinate at 16 days after sowing (DAS). However, in comparison with control seeds (0 µg/l), seeds treated with AFP I (100, 300, or 500 µg/l) germinated earlier and at a higher percentage until 20 DAS, and seeds treated with 100 µg/l AFP I showed the highest percentage of germination. Surprisingly, AFP III did not significantly increase germination, and the rate was lower among 500 µg/l AFP III-treated seeds compared with control seeds (0 µg/l). The transcription levels of the plasma membrane-associated H + -ATPase gene and antioxidant-related superoxide dismutase ( SOD ) and catalase 1 ( CAT1) genes were analyzed, and the transcription levels of the genes in the seeds grown at 25°C were relatively low. For low temperature-treated seeds, H + -ATPase in control seeds (0 µg/l) was higher compared with that in AFP I-treated seeds and was lower compared with that in AFP III-treated seeds. The expression levels of the antioxidant-related genes ( SOD and CAT1 ) were lower in AFP I-treated seeds than in control seeds (0 µg/l); however, they were higher in AFP III-treated seeds than in control seeds (0 µg/l). Overall, compared with AFP III, AFP I may potentially function as a cold-protective agent by modulating the genes associated with seed germination.

The plant hormone auxin is a major regulator of plant development and response to environmental cues. Auxin plays a particularly central role in flower development, but the knowledge of its role of flower development in crop plants with fleshy fruits, such as tomato, is still scarce. Mutations in the Aux/IAA gene ENTIRE/Indole Acetic Acid 9 (E/IAA9) lead to the precocious development of young gynoecia into parthenocarpic fruits. Here, we compared the distribution of the auxin response sensor DR5::VENUS and the auxin efflux transporter PIN1 between the wild type and entire during successive stages of flower and fruit development. Up-regulation of the DR5::VENUS signal in the shoot apical meristem (SAM) was observed upon the transition to flowering, implicating a possible role for auxin in the transition from a vegetative SAM into an inflorescence meristem. DR5::VENUS was expressed in all initiating floral organs. Additionally, DR5::VENUS was highly expressed during gametogenesis, in both male and female organs, and in the developing seeds during embryogenesis. DR5::VENUS is expressed in functional cell layers such as the anther stomium and tapetum, suggesting that auxin plays a role in flower organ development and function. The entire mutation affected DR5::VENUS expression patterns during inflorescence formation and flower organ development, which correlated with phenotypic alterations. We also show dynamic distribution and localization of the auxin transporter PIN1 during flower and fruit organ development. These results emphasize the dynamic auxin response in inflorescence and flower development and suggest multiple roles of auxin in these processes., (© The Author 2017. Published by Oxford University Press on behalf of Japanese Society of Plant Physiologists. All rights reserved. For permissions, please email: journals.permissions@oup.com.)

Main Conclusions: Exogenously applied GR24 affected somatic embryo formation and morphogenesis of strigolactone-deficient tomato mutant through cross-talk with auxins and cytokinins indicating involvement of SLs in the embryogenic process. Strigolactones (SLs) mediate the regulation of plant responses to the environment through cross-talk with other plant hormones, especially auxins. Auxins play a crucial role in coordinating the morphogenesis and development of plant reproductive organs, including the signal-transduction cascade leading to the reprogramming of gene-expression patterns before embryo formation. SLs' role in these processes is unknown, in contrast to their proven involvement in auxin transport and distribution. We used tomato cv. M82 and its SL-deficient mutant SL-ORT1 to study the influence of SLs on hormone profile in tomato roots and shoots, and their involvement in somatic embryogenesis (SE) and morphogenesis (adventitious root formation). The synthetic SL GR24 had different effects on SE of M82 and SL-ORT1, indicating that SLs influence the cytokinin-to-auxin ratio in tomato SE.

This study aimed to identify the aluminum (Al)-induced proteomes in tomato (Solanum lycopersicum, "Micro-Tom") after long-term exposure to the stress factor. Plants were treated in Magnavaca's solution (pH 4.5) supplemented with 7.5 μM Al(3+) ion activity over a 4 month period beginning at the emergence of flower buds and ending when the lower mature leaves started to turn yellow. Proteomes were identified using a 8-plex isobaric tags for relative and absolute quantification (iTRAQ) labeling strategy followed by a two-dimensional (high- and low-pH) chromatographic separation and final generation of tandem mass spectrometry (MS/MS) spectra of tryptic peptides on an LTQ-Orbitrap Elite mass spectrometer. Principal component analysis revealed that the Al-treatment had induced systemic alterations in the proteomes from roots and leaves but not seed tissues. The significantly changed root proteins were shown to have putative functions in Al(3+) ion uptake and transportation, root development, and a multitude of other cellular processes. Changes in the leaf proteome indicate that the light reaction centers of photosynthetic machinery are the primary targets of Al-induced stress. Embryo and seed-coat tissues derived from Al-treated plants were enriched with stress proteins. The biological processes involving these Al-induced proteins concur with the physiological and morphological changes, such as the disturbance of mineral homeostasis (higher contents of Al, P, and Fe and reduced contents of S, Zn, and Mn in Al-treated compared to nontreated plants) in roots and smaller sizes of roots and the whole plants. More importantly, the identified significant proteins might represent a molecular mechanism for plants to develop toward establishing the Al tolerance and adaptation mechanism over a long period of stress treatment.

Antimicrobial photodynamic treatment (APDT) is largely used in medical domain and could be envisaged as a farming practice against crop pathogens such as bacteria and fungi that generate drops in agricultural yields. Thus, as a prerequisite for this potential application, we studied the effect of water-soluble anionic (TPPS and Zn-TPPS) and cationic (TMPyP and Zn-TMPyP) porphyrins tested on tomato (Solanum lycopersicum) plantlets grown in vitro under a 16 h photoperiod. First of all, under dark conditions, none of the four porphyrins inhibited germination and induced cytotoxic effects on tomato plantlets as etiolated development was not altered. The consequences of porphyrin long-term photoactivation (14 days) were thus studied on in vitro-grown tomato plantlets at phenotypic and molecular levels. Cationic porphyrins especially Zn-TMPyP were the most efficient photosensitizers and dramatically altered growth without killing plantlets. Indeed, tomato plantlets were rescued after cationic porphyrins treatment. To gain insight, the different molecular ways implied in the plantlet tolerance to photoactivated Zn-TMPyP, lipid peroxidation, antioxidative molecules (total thiols, proline, ascorbate), and ROS detoxification enzymes were evaluated. In parallel to an increase in lipid peroxidation and hydrogen peroxide production, antioxidative molecules and enzymes (guaiacol peroxidase, catalase, and superoxide dismutase) were up-regulated in root apparatus in response to photoactivated Zn-TMPyP. This study showed that tomato plantlets could overcome the pressure triggered by photoactivated cationic porphyrin by activating antioxidative molecule and enzyme arsenal and confining Zn-TMPyP into cell wall and/or apoplasm, suggesting that APDT directed against tomato pathogens could be envisaged in the future., (Copyright © 2016 Elsevier B.V. All rights reserved.)

The spatial arrangement of leaves and flowers around the stem, known as phyllotaxis, is controlled by an auxin-dependent reiterative mechanism that leads to regular spacing of the organs and thereby to remarkably precise phyllotactic patterns. The mechanism is based on the active cellular transport of the phytohormone auxin by cellular influx and efflux carriers, such as AUX1 and PIN1. Their important role in phyllotaxis is evident from mutant phenotypes, but their exact roles in space and time are difficult to address due to the strong pleiotropic phenotypes of most mutants in phyllotaxis. Models of phyllotaxis invoke the accumulation of auxin at leaf initials and removal of auxin through their developing vascular strand, the midvein. We have developed a precise microsurgical tool to ablate the midvein at high spatial and temporal resolution in order to test its function in leaf formation and phyllotaxis. Using amplified femtosecond laser pulses, we ablated the internal tissues in young leaf primordia of tomato (Solanum lycopersicum) without damaging the overlying L1 and L2 layers. Our results show that ablation of the future midvein leads to a transient accumulation of auxin in the primordia and to an increase in their width. Phyllotaxis was transiently affected after midvein ablations, but readjusted after two plastochrons. These results indicate that the developing midvein is involved in the basipetal transport of auxin through young primordia, which contributes to phyllotactic spacing and stability., (© 2015. Published by The Company of Biologists Ltd.)

Background: Cytokinins (CKs) are involved in response to various environmental cues, including salinity. It has been previously reported that enhancing CK contents improved salt stress tolerance in tomato. However, the underlying mechanisms of CK metabolism and signaling under salt stress conditions remain to be deciphered., Results: Two tomato isopentenyltransferases, SlIPT3 and SlIPT4, were characterized in tomato and Arabidopsis. Both proteins displayed isopentenyltransferase (IPT) activity in vitro, while their encoding genes exhibited different spatio-temporal expression patterns during tomato plant development. SlIPT3 and SlIPT4 were affected by the endogenous CK status, tightly connected with CKs feedback regulation, as revealed by hormonal treatements. In response to salt stress, SlIPT3 and SlIPT4 were strongly repressed in tomato roots, and differently affected in young and old leaves. SlIPT3 overexpression in tomato resulted in high accumulation of different CK metabolites, following modifications of CK biosynthesis-, signaling- and degradation-gene expression. In addition, 35S::SlIPT3 tomato plants displayed improved tolerance to salinity consecutive to photosynthetic pigments and K(+)/Na(+) ratio retention. Involvement of SlIPT3 and SlIPT4 in salt stress response was also observed in Arabidopsis ipt3 knock-out complemented plants, through maintenance of CK homeostasis., Conclusions: SlIPT3 and SlIPT4 are functional IPTs encoded by differently expressed genes, distinctively taking part in the salinity response. The substantial participation of SlIPT3 in CK metabolism during salt stress has been determined in 35S::SlIPT3 tomato transformants, where enhancement of CKs accumulation significantly improved plant tolerance to salinity, underlining the importance of this phytohormone in stress response.

The modern concept of the hormonal regulation of fruit set, growth, maturation, and ripening is considered. Pollination and fertilization induce ovule activation by surmounting the blocking action of ethylene and ABA to be manifested in auxin accumulation. Active fruit growth by pericarp cell division and elongation is due to the syntheses ofauxin in the developing seed and ofgibberellins in the pericarp. In climacteric fleshy fruits, the maturation is controlled by ethylene via so-called System 1 combining the possibilities of autoinhibition and autocatalysis by ethylene of its own biosynthesis. Transition of tomato fruits from maturation to ripening is characterized by highly active synthesis of ethylene and its receptors due to the functioning of regulatory System 2 resulting in the up-regulation of much greater number of ethylene-inducible genes. In peach fruits, the hormonal regulation of ripening includes also an active auxin involvement in the ethylene biosynthesis, which is combined with the ethylene-induced expression of genes encoding both auxin biosynthesis and the response to auxin. Ethylene induces the expression of genes responsible for the fruit softening, its taste, color, and flavor. Nonclimacteric fleshy fruits produce very small amounts ofethylene; its evolution increases only by the very end of ripening and can be described by a reduced System 1. The ripening of nonclimacteric fruits only weakly depends on ethylene but is stimulated by abscisic acid.

Jasmonates (JAs) are ubiquitously occurring signaling compounds in plants formed in response to biotic and abiotic stress as well as in development. (+)-7-iso-jasmonoyl isoleucine, the bioactive JA, is involved in most JA-dependent processes mediated by the F-box protein COI1 in a proteasome-dependent manner. However, there is an increasing number of examples, where the precursor of JA biosynthesis, cis-(+)-12-oxophytodienoic acid (OPDA) is active in a JA/COI1-independent manner. Here, we discuss those OPDA-dependent processes, thereby giving emphasis on tomato embryo development. Recent data on seed coat-generated OPDA and its role in embryo development is discussed based on biochemical and genetic evidences.

Oxylipins including jasmonates are signaling compounds in plant growth, development, and responses to biotic and abiotic stresses. In Arabidopsis (Arabidopsis thaliana) most mutants affected in jasmonic acid (JA) biosynthesis and signaling are male sterile, whereas the JA-insensitive tomato (Solanum lycopersicum) mutant jai1 is female sterile. The diminished seed formation in jai1 together with the ovule-specific accumulation of the JA biosynthesis enzyme allene oxide cyclase (AOC), which correlates with elevated levels of JAs, suggest a role of oxylipins in tomato flower/seed development. Here, we show that 35S::SlAOC-RNAi lines with strongly reduced AOC in ovules exhibited reduced seed set similarly to the jai1 plants. Investigation of embryo development of wild-type tomato plants showed preferential occurrence of AOC promoter activity and AOC protein accumulation in the developing seed coat and the embryo, whereas 12-oxo-phytodienoic acid (OPDA) was the dominant oxylipin occurring nearly exclusively in the seed coat tissues. The OPDA- and JA-deficient mutant spr2 was delayed in embryo development and showed an increased programmed cell death in the developing seed coat and endosperm. In contrast, the mutant acx1a, which accumulates preferentially OPDA and residual amount of JA, developed embryos similar to the wild type, suggesting a role of OPDA in embryo development. Activity of the residual amount of JA in the acx1a mutant is highly improbable since the known reproductive phenotype of the JA-insensitive mutant jai1 could be rescued by wound-induced formation of OPDA. These data suggest a role of OPDA or an OPDA-related compound for proper embryo development possibly by regulating carbohydrate supply and detoxification.

Trichomes are universal biological structures originating from the aerial epidermis, which serve as an excellent model to study plant differentiation at the cell level. Although the pathway regulating trichome formation in the Rosids has been well characterized, only very recently a few genes were identified for trichome initiation in the Asterids. In this study, we cloned Woolly (Wo), essential for trichome formation in tomato. Transgenic experiments revealed that the woolly phenotype is caused by the mutation in Wo which encodes a homeodomain protein containing a bZIP motif and a START domain. We identified three alleles of Wo and found that each allele contains a missense mutation, which respectively results in an amino acid substitution at the C terminus. Microarray and expression analysis showed that the expression of a B-type cyclin gene, SlCycB2, is possibly regulated by Wo, which also participates in trichome formation. Suppression of Wo or SlCycB2 expression by RNAi decreased the number of type I trichomes, and direct protein-protein interaction was detected between them, implying that both proteins may work together in the regulation of this type of trichome formation. Cytological observation and Wo transcript analysis in the developing seeds showed that embryo development was also correlated with Wo.

Lycopersicon esculentum (tomato) fruit is a widely studied matrix. However, only few works focus their attention on its seeds, which constitute a major byproduct of the tomato processing industry. In this study the antimicrobial potential of ten different tomato seed extracts from "Bull's heart" and "Cherry" varieties were analyzed against Gram-positive (Staphylococcus aureus, Staphylococcus epidermidis, Micrococcus luteus, Enterococcus faecalis and Bacillus cereus) and Gram-negative (Proteus mirabilis, Escherichia coli, Pseudomonas aeruginosa and Salmonella typhimurium) bacteria and fungi (Candida albicans, Aspergillus fumigatus and Trichophyton rubrum). Regarding antibacterial capacity, the different extracts were revealed to be active only against Gram-positive bacteria, E. faecalis being the most susceptible one (MIC: 2.5-10 mg/mL). Concerning antifungal activity, "Bull's heart" extracts were the most active. In a general way C. albicans was the most susceptible species (MIC: 5-10 mg/mL). The chemical composition of the extracts was also pursued, concerning organic acids, phenolics and fatty acids, in order to establish a possible relationship with the observed antimicrobial effect.

In order to compare of different methods and identify the optimum condition for tomato seed extraction, factorial experiments with 3 replications was conducted. In the first experiment, pulp of two tomato cultivars (Faraon, Dominator) were fermented at two temperatures (25, 35 degrees C) and six periods (24, 48, 72, 96, 120, 144 h). The germination of seeds in laboratory as well as seedling emergence and preliminary growth in greenhouse were studied and measured. The results showed that effect of cultivar on traits (except of seedling emergence) was significant. Also the effect of temperature of fermentation, duration of fermentation and also interaction effects of them on seed germination were significant. Totally seed quality decreased with increasing temperature and duration of fermentation and the fermentation duration from 24 to 48 h at temperature 25 degrees C, is recommended. In the second experiment, tomato seeds were extracted by HCL (pH was arranged to 1, 2, 3 for 10, 20, 30 min), H2SO4 (pH was arranged to 1, 2, for 15, 30 min), Sodium carbonate (5, 10% for 24 and 48 h) and fermentation. Percentage germination, germination rate, length of radicle and length of plumule were used for seed quality assessment. The results showed that interaction effect between pH and duration of HCL treatments was significant for seed germination (percentage and rate) and there was an interaction effect between concentration and duration for germination rate in alkali treatments. Different extraction methods had not detrimental effect on percentage germination, but acid treatments produce very bright clean seeds in compare to other treatments.

In this study, seeds of Lycopersicon esculentum Mill. were analyzed by HPLC/UV-PAD/MS(n)-ESI. Fourteen flavonoids were identified, including quercetin, kaempferol, and isorhamnetin derivatives, with 13 of them being reported for the first time in tomato seeds. The major identified compounds were quercetin-3-O-sophoroside, kaempferol-3-O-sophoroside, and isorhamnetin-3-O-sophoroside. A significant cell proliferation inhibition (>80%), against rat basophile leukemia (RBL-2H3) cell line, was observed with this extract (IC(50) = 5980 microg/mL). For acetylcholinesterase inhibitory activity, a concentration-dependent effect was verified (IC(20) = 2400 microg/mL). The same behavior was noted regarding antioxidant capacity, evaluated against DPPH (IC(10) = 284 microg/mL), nitric oxide (IC(25) = 396 microg/L), and superoxide radicals (IC(25) = 3 microg/mL).

In this work, some of the different in vitro developmental pathways into which tomato microspores or microsporocytes can be deviated experimentally were explored. The two principal ones are direct embryogenesis from isolated microspores and callus formation from meiocyte-containing anthers. By means of light and electron microscopy, the process of early embryogenesis from isolated microspores and the disruption of normal meiotic development and change of developmental fate towards callus proliferation, morphogenesis, and plant regeneration have been shown. From microspores isolated at the vacuolate stage, embryos can be directly induced, thus avoiding non-androgenic products. In contrast, several different morphogenic events can be triggered in cultures of microsporocyte-containing anthers under adequate conditions, including indirect embryogenesis, adventitious organogenesis, and plant regeneration. Both callus and regenerated plants may be haploid, diploid, and mostly mixoploid. The results demonstrate that both gametophytic and sporophytic calli occur in cultured tomato anthers, and point to an in vitro-induced disturbance of cytokinesis and subsequent fusion of daughter nuclei as a putative cause for mixoploidy and genome doubling during both tetrad compartmentalization and callus proliferation. The potential implications of the different alternative pathways are discussed in the context of their application to the production of doubled-haploid plants in tomato, which is still very poorly developed.

The effects of pre-sowing magnetic treatments on growth and yield of tomato (cv Campbell-28) were investigated under field conditions. Tomato seeds were exposed to full-wave rectified sinusoidal non-uniform magnetic fields (MFs) induced by an electromagnet at 100 mT (rms) for 10 min and at 170 mT (rms) for 3 min. Non-treated seeds were considered as controls. Plants were grown in experimental plots (30.2 m(2)) and were cultivated according to standard agricultural practices. During the vegetative and generative growth stages, samples were collected at regular intervals for growth rate analyses, and the resistance of plants to geminivirus and early blight was evaluated. At physiological maturity, the plants were harvested from each plot and the yield and yield parameters were determined. In the vegetative stage, the treatments led to a significant increase in leaf area, leaf dry weight, and specific leaf area (SLA) per plant. Also, the leaf, stem, and root relative growth rates of plants derived from magnetically treated seeds were greater than those shown by the control plants. In the generative stage, leaf area per plant and relative growth rates of fruits from plants from magnetically exposed seeds were greater than those of the control plant fruits. At fruit maturity stage, all magnetic treatments increased significantly (P < .05) the mean fruit weight, the fruit yield per plant, the fruit yield per area, and the equatorial diameter of fruits in comparison with the controls. At the end of the experiment, total dry matter was significantly higher for plants from magnetically treated seeds than that of the controls. A significant delay in the appearance of first symptoms of geminivirus and early blight and a reduced infection rate of early blight were observed in the plants from exposed seeds to MFs. Pre-sowing magnetic treatments would enhance the growth and yield of tomato crop., (Copyright 2006 Wiley-Liss, Inc.)

The vacuole is a large, multifunctional organelle related to the processes of cell expansion, solute accumulation, regulation of cytoplasmic ion concentrations, pH homeostasis and osmoregulation, which are directly or indirectly achieved by vacuolar H+-pumps: vacuolar H+-ATPase (V-ATPase; EC 3.6.1.3) and vacuolar H+-pyrophosphatase (V-PPase; EC 3.6.1.1). In this study, we produced antisense-transgenic tomatoes (Lycopersicon esculentum L.) of the V-ATPase A subunit, which is under the control of the fruit-specific 2A11 promoter. One beta-glucuronidase (GUS)-transgenic line (GUS control) and seven A subunit antisense-transgenic lines were obtained. The amount of V-ATPase A subunit mRNA in fruit decreased in all antisense-transgenic lines, but in leaves showed no difference compared with the GUS control line and the nontransformant, suggesting that suppression of the V-ATPase A subunit by a 2A11 promoter is limited to fruit. The antisense-transgenic plants had smaller fruits compared with the GUS control line and the nontransformant. Surprisingly, fruits from the antisense-transgenic plants, except the fruit that still had relatively high expression of A subunit mRNA, had few seeds. Sucrose concentration in fruits from the antisense-transgenic plants increased, but glucose and fructose concentrations did not change. These results show the importance of V-ATPase, not only in fruit growth, but also in seed formation and in sugar composition of tomato fruit.

Polyphenol oxidase (PPO) activity increased markedly in the micropylar region of the endosperm of tomato (Lycopersicon esculentum) seeds after radicle protrusion. Tissue-printing analyses demonstrated that the majority of the activity is localized in the micropylar area. The increase in the activity was due to the increase in the amounts of enzyme. Within the micropylar endosperm region, two PPO isozymes were immunologically detected whose apparent molecular masses were estimated to be approximately 58 and 59 kDa, respectively, by SDS-PAGE. Although PPO activity also developed in the lateral portion of the endosperm, the level of this activity was much lower as compared with that in the micropylar region. Furthermore, the isozyme pattern in the lateral portion differed from that in the micropylar portion. The 58 kDa isozyme that was detected in the latter portion was absent, and only 59 kDa PPO was detectable in the former one. When the endosperm tissues were wounded, an enhancement of the enzyme activity was observed in the wounded region. The wound-induced development of the enzyme activity was associated with the induction of 58 kDa isozyme.

Proteome analysis of embryo and endosperm tissues from germinating tomato seed was conducted using 1-DE, 2-DE, and MS. Mobilization of the most abundant proteins, which showed similar profiles in the two tissues, occurred first in the endosperm. CBB R-250 staining of 2-DE gels revealed 352 and 369 major protein spots in the embryo and endosperm, respectively, at 0 h. Of these, 75 major spots were selected, excised, in-gel digested with trypsin, and analyzed by MALDI-TOF-MS and/or LC-ESI-Q/TOF-MS/MS. Peptide MS and MS/MS data were searched against publicly available protein and EST databases, and 47 proteins identified. Embryo-specific proteins included a BAC19.13 homologue, whereas four proteins specific to the endosperm were tomato mosaic virus coat proteins related to defense mechanisms. The most abundant proteins both in the embryo and endosperm were seed storage proteins, i.e., legumins (11 spots), vicilins (11 spots), albumin (2 spots). Housekeeping enzymes, actin-binding profilin, defense-related protein kinases, nonspecific lipid transfer protein, and proteins involved in general metabolism were also identified. The roles of some of the proteins identified in the embryo and endosperm are discussed in relation to seed germination in tomato.

Potatoes and tomatoes, members of the Solanaceae plant family, serve as major, inexpensive low-fat food sources providing for energy, high-quality protein, fiber, vitamins, pigments, as well as other nutrients. These crops also produce biologically active secondary metabolites, which may have both adverse and beneficial effects in the diet. This limited overview, based largely on our studies with the aid of HPLC, TLC, ELISA, GC-MS, and UV spectroscopy, covers analytical aspects of two major potato trisaccharide glycoalkaloids, alpha-chaconine and alpha-solanine, and their hydrolysis products (metabolites) with two, one, and zero carbohydrate groups; the potato water-soluble nortropane alkaloids calystegine A3 and B2; the principal potato polyphenolic compound chlorogenic acid; potato inhibitors of digestive enzymes; the tomato tetrasaccharide glycoalkaloids dehydrotomatine and alpha-tomatine and hydrolysis products; the tomato pigments beta-carotene, lycopene, and chlorophyll; and the anticholinergic alkaloids atropine and scopolamine present in Datura stramonium (jimson weed) seeds that contaminate grain and animal feed. Related studies by other investigators are also mentioned. Accurate analytical methods for these food ingredients help assure the consumer of eating a good-quality and safe diet.

The parthenocarpic fruit ( pat) gene of tomato is a recessive mutation conferring parthenocarpy, which is the capability of a plant to set seedless fruits in the absence of pollination and fertilization. Parthenocarpic mutants offer a useful method to regulate fruit production and a suitable experimental system to study ovary and fruit development. In order to map the Pat locus, two populations segregating from the interspecific cross Lycopersicon esculentum x Lycopersicon pennellii were grown, and progeny plants were classified as parthenocarpic or wild-type by taking into account some characteristic aberrations affecting mutant anthers and ovules. Through bulk segregant analysis, we searched for both random and mapped AFLPs linked to the target gene. In this way, the Pat locus was assigned to the long arm of chromosome 3, as also confirmed by the analysis of a set of L. pennellii substitution and introgression lines. Afterwards, the Pat position was refined by using simple sequence repeats (SSRs) and conserved ortholog set (COS) markers mapping in the target region. The tightest COSs were converted into CAPS or SCAR markers. At present, two co-dominant SCAR markers encompassing a genetic window of 1.2 cM flank the Pat locus. Considering that these markers are orthologous to Arabidopsis genes, a positional cloning exploiting the tomato- Arabidopsis microsynteny seems to be a short-term objective.

Two samples of tomato seeds, a by-product of the tomato canning industry were evaluated to determine proximate analysis, amino acid content, and digestibility, TMEn, and protein efficiency ratio. Tomato seeds were also used to replace corn and soybean meal (SBM) in a chick diet on an equal true amino acid digestibility and TMEn basis. Tomato seeds were found to contain 8.5% moisture, 25% CP, 20.0% fat, 3.1% ash, 35.1% total dietary fiber, 0.12% Ca, 0.58% P, and 3,204 kcal/kg of TMEn. The total amounts of methionine, cystine, and lysine in the tomato seeds were 0.39, 0.40, and 1.34%, respectively, and their true digestibility coefficients, determined in cecectomized roosters, were 75, 70, and 54%, respectively. The protein efficiency ratio (weight gain per unit of protein intake) value when fed to chicks at 9% CP was 2.5 compared to 3.6 for SBM (P < or = 0.05). When corn-SBM diets were formulated on an equal true amino acid digestibility and TMEn basis, up to 15% tomato seeds could replace corn and SBM without any adverse affects on chick weight gain, feed intake, or gain:feed ratio from 8 to 21 d posthatch. Tomato seeds at any level in the diet did not significantly affect skin pigmentation. Although the protein quality of tomato seeds may not be as high as SBM, tomato seeds do contain substantial amounts of digestible amino acids and TMEn. When formulating diets on a true digestible amino acid and TMEn basis, tomato seeds can be supplemented into chick rations at up to 15% without any adverse affects on growth performance.

Although tomato has been the subject of extensive quantitative trait loci (QTLs) mapping experiments, most of this work has been conducted on transient populations (e.g., F2 or backcross) and few homozygous, permanent mapping populations are available. To help remedy this situation, we have developed a set of inbred backcross lines (IBLs) from the interspecific cross between Lycopersicon esculentum cv. E6203 and L. pimpinellifolium (LA1589). A total of 170 BC2F1 plants were selfed for five generations to create a set of homozygous BC2F6 lines by single-seed descent. These lines were then genotyped for 127 marker loci covering the entire tomato genome. These IBLs were evaluated for 22 quantitative traits. In all, 71 significant QTLs were identified, 15% (11/71) of which mapped to the same chromosomal positions as QTLs identified in earlier studies using the same cross. For 48% (34/71) of the detected QTLs, the wild allele was associated with improved agronomic performance. A number of new QTLs were identified including several of significant agronomic importance for tomato production: fruit shape, firmness, fruit color, scar size, seed and flower number, leaf curliness, plant growth, fertility, and flowering time. To improve the utility of the IBL population, a subset of 100 lines giving the most uniform genome coverage and map resolution was selected using a randomized greedy algorithm as implemented in the software package MapPop (http://www.bio.unc.edu/faculty/vision/lab/ mappop/). The map, phenotypic data, and seeds for the IBL population are publicly available (http://soldb.cit.cornell.edu) and will provide tomato geneticists and breeders with a genetic resource for mapping, gene discovery, and breeding.

beta-1,3-Glucanase (EC 3.2.1.39) and chitinase (EC 3.2.1.14) mRNAs, proteins, and enzyme activities were expressed specifically in the micropylar tissues of imbibed tomato (Lycopersicon esculentum Mill.) seeds prior to radicle emergence. RNA hybridization and immunoblotting demonstrated that both enzymes were class I basic isoforms. beta-1,3-Glucanase was expressed exclusively in the endosperm cap tissue, whereas chitinase localized to both endosperm cap and radicle tip tissues. beta-1,3-Glucanase and chitinase appeared in the micropylar tissues of gibberellin-deficient gib-1 tomato seeds only when supplied with gibberellin. Accumulation of beta-1,3-glucanase mRNA, protein and enzyme activity was reduced by 100 microM abscisic acid, which delayed or prevented radicle emergence but not endosperm cap weakening. In contrast, expression of chitinase mRNA, protein, and enzyme activity was not affected by abscisic acid. Neither of these enzymes significantly hydrolyzed isolated tomato endosperm cap cell walls. Although both beta-1,3-glucanase and chitinase were expressed in tomato endosperm cap tissue prior to radicle emergence, we found no evidence that they were directly involved in cell wall modification or tissue weakening. Possible functions of these hydrolases during tomato seed germination are discussed.

The effect of 2,4,5-trichlorophenoxy acetic acid (2,4,5-T) on the germination and seedling vigor of different crop seeds was tested. Seeds of rice, maize, sorghum, finger millet, and horse gram were comparatively more tolerant to the chemical with no marked effect up to a concentration of 200 mg 2,4,5-T L(-)(1) as tested by the filter paper method. Tomato and brinjal (egg plant) were highly susceptible. Even at 5 and 10 mg 2,4,5-T L(-)(1), marked reduction in the germination and seedling vigor of tomato and egg plant, respectively, was observed. At 20 and 30 mg L(-)(1), the germination of tomato and egg plant seeds, respectively, were completely inhibited on filter paper, whereas the inhibitory concentrations in soil was 40 mg 2,4,5-T kg(-)(1) soil. Several abnormalities were observed in the chemically affected seedlings. Protease activity of the seeds germinating in the presence of the chemical was drastically reduced. Bioremediation of the chemically contaminated soil with Burkholderia cepacia AC1100, by inoculation of the soil 7 days before sowing the seeds, completely protected the seeds, resulting in normal germination and an improved seedling vigor.

Laminarin-hydrolysing activity developed in the endosperm of tomato (Lycopersicon esculentum) seeds following germination. The enzyme was basic (pI>10) and the apparent molecular mass was estimated to be 35 kDa by SDS-PAGE. It was specific for linear beta-1,3-glucan substrates. Laminarin was hydrolysed by the enzyme to yield a mixture of oligoglucosides, indicating that the enzyme had an endo-action pattern. Thus, the enzyme was identified as beta-1,3- endoglucanase (EC 3.2.1.39). The activity of the enzyme developed in the endosperm after radicle protrusion (germination) had occurred and the enzyme activity was localized exclusively in the micropylar region of the endosperm where the radicle had penetrated. When the lateral endosperm region, where no induction of the enzyme occurred, was wounded (cut or punctured), there was a marked enhancement of beta-1,3-glucanase activity. Thus the post-germinative beta-1, 3-glucanase activity in the micropylar endosperm portion might be brought about by wounding resulting from endosperm rupture by radicle penetration.

The role of abscisic acid (ABA) in the weakening of the endosperm cap prior to radicle protrusion in tomato (Lycopersicon esculentum Mill. cv. Moneymaker) seeds was studied. The endosperm cap weakened substantially in both water and ABA during the first 38 h of imbibition. After 38 h the force required for endosperm cap puncturing was arrested at 0.35 N in ABA, whereas in water a further decrease occurred until the radicle protruded. During the first 2 d of imbibition endo-beta-mannanase activity was correlated with the decrease in required puncture force and with the appearance of ice-crystal-induced porosity in the cell walls as observed by scanning electron microscopy. Prolonged incubation in ABA resulted in the loss of endo-beta-mannanase activity and the loss of ice-crystal-induced porosity, but not in a reversion of the required puncture force. ABA also had a distinct but minor effect on the growth potential of the embryo. However, endosperm cap resistance played the limiting role in the completion of germination. It was concluded that (a) endosperm cap weakening is a biphasic process and (b) inhibition of germination by ABA is through the second step in the endosperm cap weakening process.

High frequency of streptomycin resistant variants of Lycopersicon esculentum were isolated on selective shoot regeneration medium supplemented with IAA (0.5 mg/L), zeatin (1.5 mg/L) and streptomycin sulphate (500 mg/L). Nonmutagenized (controls) and NMU treated cotyledons were placed on shoot regeneration medium supplemented with antibiotic streptomycin. Resistant shoots appeared at a high frequency in mutagenized cotyledons, whereas in controls morphogenesis was suppressed, accompanied by bleaching. Shoot regeneration occurred from the nodular tissues developed at the cut ends of cotyledons. Resistant shoots developed into complete plantlets on rooting medium containing selective concentration of antibiotic. Stability of streptomycin resistance was confirmed by leaf assay and reciprocal crosses between streptomycin-resistant and sensitive plants.

We studied cell cycle events in embryos of tomato (Lycopersicon esculentum Mill. cv Moneymaker) seeds during imbibition in water and during osmoconditioning ("priming") using both quantitative and cytological analysis of DNA synthesis and beta-tubulin accumulation. Most embryonic nuclei of dry, untreated control seeds were arrested in the G(1) phase of the cell cycle. This indicated the absence of DNA synthesis (the S-phase), as confirmed by the absence of bromodeoxyuridine incorporation. In addition, beta-tubulin was not detected on western blots and microtubules were not present. During imbibition in water, DNA synthesis was activated in the radicle tip and then spread toward the cotyledons, resulting in an increase in the number of nuclei in G(2). Concomitantly, beta-tubulin accumulated and was assembled into microtubular cytoskeleton networks. Both of these cell cycle events preceded cell expansion and division and subsequent growth of the radicle through the seed coat. The activation of DNA synthesis and the formation of microtubular cytoskeleton networks were also observed throughout the embryo when seeds were osmoconditioned. However, this pre-activation of the cell cycle appeared to become arrested in the G(2) phase since no mitosis was observed. The pre-activation of cell cycle events in osmoconditioned seeds appeared to be correlated with enhanced germination performance during re-imbibition in water.

Experiments with ethylene-insensitive tomato (Lycopersicon esculentum) and petunia (Petunia x hybrida) plants were conducted to determine if normal or adventitious root formation is affected by ethylene insensitivity. Ethylene-insensitive Never ripe (NR) tomato plants produced more below-ground root mass but fewer above-ground adventitious roots than wild-type Pearson plants. Applied auxin (indole-3-butyric acid) increased adventitious root formation on vegetative stem cuttings of wild-type plants but had little or no effect on rooting of NR plants. Reduced adventitious root formation was also observed in ethylene-insensitive transgenic petunia plants. Applied 1-aminocyclopropane-1-carboxylic acid increased adventitious root formation on vegetative stem cuttings from NR and wild-type plants, but NR cuttings produced fewer adventitious roots than wild-type cuttings. These data suggest that the promotive effect of auxin on adventitious rooting is influenced by ethylene responsiveness. Seedling root growth of tomato in response to mechanical impedance was also influenced by ethylene sensitivity. Ninety-six percent of wild-type seedlings germinated and grown on sand for 7 d grew normal roots into the medium, whereas 47% of NR seedlings displayed elongated tap-roots, shortened hypocotyls, and did not penetrate the medium. These data indicate that ethylene has a critical role in various responses of roots to environmental stimuli.

Among the different sources of genetic parthenocarpy described in tomato, the mutation referred to as parthenocarpic fruit (pat) is of particular interest because of its strong expressivity and because it confers earlier ripening, higher fruit set and enhanced fruit quality. As a pleiotropic effect, pat flowers have aberrantly developing androecia and reduced male and female fertility. In this work we extend the early description of the pat phenotype by investigating the expression of parthenocarpy in three different environments and by using light and scanning electron microscopy to analyse the development of male and female floral organs. The degree of parthenocarpy was high in the three experimental environments and was characterised by a precocious initiation of ovary growth to pre-anthesis floral stages. Aberrations in anther development were evident at flower bud stages and resulted in shorter, irregular and teratoid organs. Ectopic production of carpel-like structures bearing external ovules was evident in the most severely altered androecia. Analysis of ovule development revealed that a fraction of pat ovules becomes aberrant from very early stages, having defective integument growth. Meiosis was irregular in aberrant ovules and megaspore or gamete production was severely hampered. The described pat syndrome suggests that parthenocarpy in this mutant could be a secondary effect of a gene controlling, at early stages, organ identity and development.

The identification of plant genes involved in early phases of in vitro morphogenesis can not only contribute to our understanding of the processes underlying growth regulator-controlled determination, but also provide novel markers for evaluating the outcome of in vitro regeneration experiments. To search for such genes and to monitor changes in gene expression accompanying in vitro regeneration, we have adapted the mRNA differential display technique to the comparative analysis of a model system of tomato cotyledons that can be driven selectively toward either shoot or callus formation by means of previously determined growth regulator supplementations. Hormone-independent transcriptional modulation (mainly down-regulation) has been found to be the most common event, indicating that a non-specific reprogramming of gene expression quantitatively predominates during the early phases of in vitro culture. However, cDNA fragments representative of genes that are either down-regulated or induced in a programme-specific manner could also be identified, and two of them (G35, G36) were further characterized. One of these cDNA fragments, G35, corresponds to an mRNA that is down-regulated much earlier in callus- (day 2) than in shoot-determined explants (day 6). The other, G36, identifies an mRNA that is transiently expressed in shoot-determined explants only, well before any macroscopic signs of differentiation become apparent, and thus exhibits typical features of a morphogenetic marker.

The Pto gene encodes a serine/threonine kinase that confers resistance to bacterial speck disease in tomato. Using the yeast two-hybrid system, we identified a second serine/threonine kinase, Pto-interacting 1 (Pti1), that physically interacts with Pto. Cross-phosphorylation assays revealed that Pto specifically phosphorylates Pti1 and that Pti1 does not phosphorylate Pto. Fen, another serine/threonine kinase from tomato that is closely related to Pto, was unable to phosphorylate Pti1 and was not phosphorylated by Pti1. Expression of a Pti1 transgene in tobacco plants enhanced the hypersensitive response to a P. syringae pv. tabaci strain carrying the avirulence gene avrPto. These findings indicate that Pti1 is involved in a Pto-mediated signaling pathway, probably by acting as a component downstream of Pto in a phosphorylation cascade.