A Role for Differential Glycoconjugation in the Emission of Phenylpropanoid Volatiles from Tomato Fruit Discovered Using a Metabolic Data Fusion Approach

A role for differential glycoconjugation in the emission of phenylpropanoid volatiles from ripening tomato fruit (Solanum lycopersicum) upon fruit tissue disruption has been discovered in this study. Application of a multiinstrumental analytical platform for metabolic profiling of fruits from a diverse collection of tomato cultivars revealed that emission of three discriminatory phenylpropanoid volatiles, namely methyl salicylate, guaiacol, and eugenol, took place upon disruption of fruit tissue through cleavage of the corresponding glycoconjugates, identified putatively as hexose-pentosides. However, in certain genotypes, phenylpropanoid volatile emission was arrested due to the corresponding hexose-pentoside precursors having been converted into glycoconjugate species of a higher complexity: dihexose-pentosides and malonyl-dihexose-pentosides. This glycoside conversion was established to occur in tomato fruit during the later phases of fruit ripening and has consequently led to the inability of red fruits of these genotypes to emit key phenylpropanoid volatiles upon fruit tissue disruption. This principle of volatile emission regulation can pave the way to new strategies for controlling tomato fruit flavor and taste.More than 7,000 metabolites, including volatiles, have already been identified in plant-based foods and beverages (Goff and Klee, 2006). Volatile organic compounds constitute a significant part of the plant metabolome, and the number of individual volatiles already described for various plants is approaching 2,000 (Dudareva et al., 2006). Significant progress has been made on the functional characterization of these plant volatiles over the past decades. For example, volatiles have been shown to play an important role in the interaction between plants and their environment. They are involved in the defense of plants against pathogens, where they serve as airborne signaling molecules to induce a defense response in other plant parts or neighboring plants (Shulaev et al., 1997). They also act as direct repellents of herbivorous pests or as attractants of the predators of these pests as part of the “cry for help” response (Dudareva et al., 2004; Kappers et al., 2005; Baldwin et al., 2006). In addition, flower volatiles are important for the attraction of pollinators (Dudareva et al., 2004), while fruit volatiles may have a role in attracting seed dispersers (Goff and Klee, 2006; Schwab et al., 2008). Besides their physiological and ecological functions, plant volatiles are also important determinants of consumer quality traits in flowers, fruits, and vegetables as well as the processed products derived from them.Tomato (Solanum lycopersicum) is one of the most important vegetable crops worldwide, and its fresh fruits and processed products are consumed and appreciated in every society. Volatiles are considered as major determinants of tomato fruit flavor (Buttery et al., 1987; Buttery and Ling, 1993; Baldwin et al., 1998, 2000; Tandon et al., 2000; Krumbein et al., 2004; Ruiz et al., 2005; Tieman et al., 2006; Kovács et al., 2009; Zanor et al., 2009). Several hundred tomato fruit volatile compounds have been described in the literature (Petro-Turza, 1987), but only a small part of this diversity is believed to have an impact on tomato fruit organoleptic properties (Buttery and Ling, 1993; Baldwin et al., 2000). We have previously screened red-ripe fruits for variation in their volatile metabolome using a collection of 94 tomato cultivars representing the current diversity within commercial germplasm (Tikunov et al., 2005). In that study, three phenylpropanoid (PhP) volatiles, methyl salicylate (MeSA), guaiacol, and eugenol, were found to be discriminatory within this germplasm collection and roughly divided the cultivars into two groups. Fruits from one group had the capacity to emit significant amounts of these three PhP volatiles upon fruit tissue disruption (blending), while fruits from the other group emitted none or hardly any.The considered relevance of these findings relates to their potential importance in consumer perception of fruit taste differences. It has been proposed previously that PhP volatiles likely have an impact on tomato fruit aroma. MeSA, the methyl ester of salicylic acid, is a potent odor component of wintergreen (Gaultheria procumbens). MeSA content has been shown to be negatively correlated with typical tomato flavor (Krumbein and Auerswald, 1998). Guaiacol is also a well-known flavoring compound and has been associated with a so-called “pharmaceutical” aroma in tomato fruits (Causse et al., 2002). Likewise, eugenol is a well-known odorant that gives the distinctive, pungent flavor to cloves (Syzygium aromaticum) and significantly contributes to the aroma of cinnamon (Cinnamomum verum). Although a potential physiological role for these PhP volatiles in tomato fruits remains unclear, they have been implicated to have a signaling and/or defense function (Shulaev et al., 1997; Koeduka et al., 2006; Sasso et al., 2007). Therefore, there is clear potential for the release of these compounds to influence, either negatively or positively, tomato flavor.In plants, PhP volatiles are primarily derived from Phe (Dudareva and Pichersky, 2000). Cinnamic acid, directly derived from Phe by a deamination catalyzed by Phe ammonia lyase, can either be β-oxidatively or nonoxidatively converted into benzoic acid. This can be further hydroxylated into salicylic acid by benzoic acid 2-hydroxylase. Recently, genetic studies in Arabidopsis (Arabidopsis thaliana) revealed the existence of an alternative pathway for the production of salicylic acid from isochorismate, thus bypassing Phe and its derivatives (Wildermuth et al., 2001). Salicylic acid is the immediate precursor of the volatile MeSA, through the action of salicylic acid methyl transferase (Boatright et al., 2004). Cinnamic acid can also be converted to other phenolic acids: p-coumaric acid, caffeic acid, and ferulic acid. Ferulic acid can be converted into coniferyl alcohol and further to eugenol (Gang, 2005). The biochemical origin of guaiacol in plants is not completely known. However, its chemical structure clearly points to the same PhP origin, as has already been demonstrated for bacteria (Chang and Kang, 2004).Glycosylation is a common means to conjugate plant secondary metabolites, in order to facilitate their transport and storage and to reduce their reactivity by blocking reactive hydroxyl groups. In tomato fruit, many volatile compounds, including PhP volatiles, are bound as glycosides, thus representing an aroma reserve (Buttery et al., 1990; Marlatt et al., 1992; Ortiz-Serrano and Gil, 2007). Such glycosidically bound volatiles can be liberated when cell compartmentation is destroyed, as happens on consumption of fresh fruits or industrial processing or as may happen during late ripening stages. As a consequence of this disruption, the contents of different cell compartments can mix and stored volatile glycosides become exposed to endogenous or exogenous cleavage enzymes, such as glycosyl hydrolases (glycosidases), which leads to glycoside cleavage and volatile emission. Thus, understanding the biochemical processes leading to the formation and/or cleavage of volatile glycoconjugates may provide tools to exploit more efficiently the aroma reserve present in tomato fruit in order to improve tomato fruit flavor.In order to gain greater insight into the volatile compound variation in tomato fruit, we previously analyzed the volatile metabolites using solid-phase microextraction-gas chromatography-mass spectrometry (SPME-GC-MS) in a broad screening of 94 contrasting tomato genotypes representing the variation present in the germplasm of commercial tomato varieties (Tikunov et al., 2005; Van Berloo et al., 2008). This nontargeted metabolomics approach enabled the detection and putative identification of 322 volatiles. Subsequent multivariate analysis revealed (1) differences between tomato types (cherry versus round tomatoes) driven by the accumulation of phenolic-derived volatiles such as phenylethanol and phenylacetaldehyde, and (2) that the PhP-derived volatiles MeSA, guaiacol, and eugenol roughly split the set of genotypes into two distinct groups, independent of tomato fruit (pheno)type, where fruits of one of these groups emitted considerable amounts of these three PhP volatiles but fruits of the other emitted little or none.In this paper, we describe the investigation into the biochemical basis underlying this difference in capacity to emit PhP volatiles. A multiinstrumental metabolomics platform was used to profile fruits of the same broad tomato germplasm collection for both volatile and nonvolatile metabolites. Metabolic data fusion of both liquid chromatography (LC)-MS and GC-MS data sets, followed by multivariate analyses, suggested a principle for the regulation of PhP volatile emission in tomato fruit through differential volatile-sugar conjugation patterns. Subsequent series of quantitative biochemical experiments proved an important role of this process in regulating the emission of PhP volatiles from tomato fruit.     

An Ecological Role of Volatiles Produced by Lasiodiplodia theobromae

One function of the volatiles produced by the fungus Lasiodiplodia theobromae was supposed to be ecological, as an insect attractant, which could get the attracted insects to be a vector of the fungal spores. L. theobromae, a mellein-producing fungus, was isolated from the outer surface of a danaid butterfly, Idea leuconoe. Ingestion by the butterfly of mellein on some plant tissues that contained mellein from 0.3 to 11 ppm was observed in the field and also in an insectarium. All these plant tissues were infected with L. theobromae, therefore, the mellein was concluded to be produced by the fungi. These observation suggests the presence of a kind of mutualistic relation between Idea leuconoe and L. theobromae associated with mellein, which is a suspected sex pheromone of the butterfly, accumulated in its hairpencil.     

The Photoprotective Role of Spermidine in Tomato Seedlings under Salinity-Alkalinity Stress

Polyamines are small, ubiquitous, nitrogenous compounds that scavenge reactive oxygen species and stabilize the structure and function of the photosynthetic apparatus in response to abiotic stresses. Molecular details underlying polyamine-mediated photoprotective mechanisms are not completely resolved. This study investigated the role of spermidine (Spd) in the structure and function of the photosynthetic apparatus. Tomato seedlings were subjected to salinity-alkalinity stress with and without foliar application of Spd, and photosynthetic and morphological parameters were analyzed. Leaf dry weight and net photosynthetic rate were reduced by salinity-alkalinity stress. Salinity-alkalinity stress reduced photochemical quenching parameters, including maximum photochemistry efficiency of photosystem II, quantum yield of linear electron flux, and coefficient of photochemical quenching (qP). Salinity-alkalinity stress elevated nonphotochemical quenching parameters, including the de-epoxidation state of the xanthophyll cycle and nonphotochemical quenching (NPQ). Microscopic analysis revealed that salinity-alkalinity stress disrupted the internal lamellar system of granal and stromal thylakoids. Exogenous Spd alleviated the stress-induced reduction of leaf dry weight, net photosynthetic rate, and qP parameters. The NPQ parameters increased by salinity-alkalinity stress were also alleviated by Spd. Seedlings treated with exogenous Spd had higher zeaxanthin (Z) contents than those without Spd under salinity-alkalinity stress. The chloroplast ultrastructure had a more ordered arrangement in seedlings treated with exogenous Spd than in those without Spd under salinity-alkalinity stress. These results indicate that exogenous Spd can alleviate the growth inhibition and thylakoid membrane photodamage caused by salinity-alkalinity stress. The Spd-induced accumulation of Z also may have an important role in stabilizing the photosynthetic apparatus.     

Agrobacterium tumefaciens Interaction with Suspension-Cultured Tomato Cells

Adherence of Agrobacterium tumefaciens to suspension-cultured tomato cells has been characterized using a quantitative binding assay. Saturable binding of radiolabeled A. tumefaciens to plant cells resulted in 100 to 300 bacteria bound per cell. Specificity of A. tumefaciens binding was also inferred from two additional results: (a) an initial incubation of plant cells with A. tumefaciens reduced subsequent binding of radiolabeled A. tumefaciens by 60% to 75%; (b) tomato cells bound less than three E. coli per cell. Protease treatment of plant cells had no effect on subsequent bacterial binding, but prior treatment of plant cells with pectinolytic enzymes increased binding 2- to 3-fold. Pectin-enriched and neutral polymer-enriched fractions were obtained from tomato cell walls. The soluble pectin-enriched fraction inhibited binding of bacteria to plant cells by 85% to 95%, whereas the neutral polymer fraction only partially inhibited binding. Preliminary characterization of the activity showed it is heat stable, partially inactivated by protease treatment, and substantially inactivated by acid hydrolysis.     

The Role of Viral Infection in Inducing Variability in Virus-Free Progeny in Tomato

The effect of virus-host interactions on subsequent generations is poorly understood. The evaluation of the effects of viral infection on inheritance of quantitative traits in the progeny of infected plants and elucidation of a possible relationship between chiasma frequency in the infected plants and variability of traits in the progeny were investigated. The current study involved genotypes of four intraspecific hybrids of tomato (Solanum lycopersicum L.), their parental forms and two additional cultivars. Used as infection were the tobacco mosaic virus (TMV) and potato virus X (PVX). The consequences of the effect of viral infection were evaluated based on chromosome pairing in diakinesis and/or by examining quantitative and qualitative traits in the progeny of the infected tomato plants. Tomato plants infected with TMV + PVX were found to differ in chiasma frequency per pollen mother cell or per bivalent. Deviations have been observed for genotypes of both F1 hybrids and cultivars. At the same time, differences in mean values of the traits under study have only been found for progeny populations (F2-F4) derived from virus-infected F1 hybrids, but not in the case of progeny of the infected cultivars. The rate of recombinants combining traits of both parents increased significantly (2.22-8.24 times) in progeny populations of hybrids infected with TMV + PVX. The above suggests that the observed effects could be the result of modification of recombination frequencies that can be manifested in heterozygous hybrids and make small contributions to variability in cases of 'homozygous' tomato genotypes (I.e. Cultivars).     

The Role of Viral Infection in Inducing Variability in Virus-Free Progeny in Tomato

The effect of virus-host interactions on subsequent generations is poorly understood. The evaluation of the effects of viral infection on inheritance of quantitative traits in the progeny of infected plants and elucidation of a possible relationship between chiasma frequency in the infected plants and variability of traits in the progeny were investigated. The current study involved genotypes of four intraspecific hybrids of tomato （Solanum lycopersicum L.）, their parental forms and two additional cultivars. Used as infection were the tobacco mosaic virus （TMV） and potato virus X （PVX）. The consequences of the effect of viral infection were evaluated based on chromosome pairing in diakinesis and/or by examining quantitative and qualitative traits in the progeny of the infected tomato plants. Tomato plants infected with TMV ＋ PVX were found to differ in chiasma frequency per pollen mother cell or per bivalent. Deviations have been observed for genotypes of both F1 hybrids and cultivars. At the same time, differences in mean values of the traits under study have only been found for progeny populations （F2-F4） derived from virus-infected F1 hybrids, but not in the case of progeny of the infected cultivars. The rate of recombinants combining traits of both parents increased significantly （2.22-8.24 times） in progeny populations of hybrids infected with TMV＋PVX. The above suggests that the observed effects could be the result of modification of recombination frequencies that can be manifested in heterozygous hybrids and make small contributions to variability in cases of ＇homozygous＇ tomato genotypes （i.e. cultivars）.     

Role of Ethylene in the Biosynthetic Pathways of Aroma Volatiles in Ripening Fruit

During the past decade, fruit aroma biosynthetic pathways were established in some climacteric fruits, such as tomato, apple, and melon. Inhibition of ethylene biosynthesis or its action in these fruits can reduce the production of fruit volatiles. Furthermore, ethylene partially regulates expression of a few important enzyme genes in fruit volatile biosynthetic pathways. The aim of this review is to bring together recent advances for understanding the regulatory role of ethylene in the biosynthesis of aroma volatiles in some fruits.     

Influence of Sink-Source Interaction on Dry Matter Production in Tomato

Sink-source ratio in tomato was manipulated, in six glasshouseexperiments, by fruit pruning (trusses pruned to two to sevenfruits immediately after fruit set of each truss), truss pruning(removal of every other truss at anthesis) and truss pruningin plants with two shoots. Periodic destructive harvest wereconducted for about 100 d after flowering of the first truss.Dry matter production was not influenced by sink-source ratio,whereas dry matter distribution between fruits and vegetativeparts was greatly affected. The fraction of dry matter distributedto the fruits at the end of the fruit pruning experiments (F_fruits)could be described accurately as a saturation-type functionof number of fruits retained per truss (N_f): F_fruits = 0.660(l-e^-0.341N_f). Specific leaf area and internode length decreasedand plant leaf area increased when sink-source ratio was reduced.Removal of every other truss at anthesis did reduce dry matterpartitioning into the fruits, but it did not influence internodelength. Plant development (number of visible leaves at the endof the experiments) was not influenced by sink-source ratio.In four experiments some plants were pruned to one fruit pertruss. Final dry matter production was 8-24% lower for theseplants, compared with plants with more than one fruit per truss.This was, at least party, the result of less light interceptionby these plants, which had strongly curled leaves pointing downwards. Results indicate that effects of sink demand on dry matter productionper unit of intercepted radiation and probably on leaf photosyntheticrate in commercial tomato production can be ignored.Copyright1995, 1999 Academic Press Dry matter production, feedback control, glasshouse, growth analysis, Lycopersicon esculentum, pruning, sink demand, sink-source ratio, tomato     

黑芥子酶基因pyk10根特异启动子在番茄中的验证

黑芥子酶是一类催化芥子油苷水解的同工酶,pyk10是一个在拟南芥根和下胚轴中特异表达的黑芥子酶基因.从拟南芥Columbia生态型基因组中克隆的长度为1 450 bp的pykl0基因启动子片段,以gus基因为报告基因构建了植物表达载体pPykG.通过农杆菌介导法,将pykl0的根特异表达启动子以及gus基因转入了番茄中蔬6号,经PCR检测,转化植株中扩增出pyk10启动子特异性条带.组织化学法检测及定量分析,显示pyk10启动子驱动gus基因在番茄的根部特定表达.