Participation of ornithine decarboxylase in early stages of tomato fruit development

The apparent association of ornithine decarboxylase (ODC) with rapid cell proliferation in developing tomato (Lycopersicon esculentum Mill. cv. Pearson ms-35) fruits has been previously described. Further evidence is provided by the use of two ODC inhibitors, α-difluoromethylornithine (α-DFMO) and α-methylornithine (α-MO). Fruit development was inhibited by these inhibitors if applied during the period of intensive cell division. When applied in vitro, the two inhibitors were shown to inhibit the activity of ODC but not that of arginine decarboxylase (ADC). When applied in vivo, α-DFMO, a catalytic irreversible inhibitor, caused 97.1% reduction of ODC activity in the dialyzed extract from the treated ovaries, while it had no effect on ADC. On the other hand, α-MO, a reversible inhibitor, did not reduce the activity of these two enzymes in the dialyzed extracts when applied in vivo. The dialysis procedure probably removed α-MO from the enzyme fraction. Putrescine, the product of both ODC and ADC, alleviated the inhibition of fruit development but did not restore ODC activity to the control level. These results suggest that in the young developing tomato fruit, ODC is the enzyme responsible for the synthesis of putrescine, which is essential for the early stages of fruit development. The reduced activity of ODC elicited by putrescine suggests a mechanism of feedback regulation by enzyme repression or release of an ODC anti-enzyme.     

The Solanum melongena COP1LIKE manipulates fruit ripening and flowering time in tomato (Solanum lycopersicum)

Plant Growth Regulation - The CONSTITUTIVE PHOTOMORPHOGENIC (COP) 1LIKE is a regulatory protein and repressor of photomorphogenesis; which control many processes of development in plants. Here, the...     

Novel promoters that induce specific transgene expression during the green to ripening stages of tomato fruit development

Fruit-specific promoters have been used as genetic engineering tools for studies on molecular mechanism of fruit development and advance in fruit quality and additional value by increasing functional component. Especially fruit-ripening specific promoters have been well utilized and studied in tomato; however, few studies have reported the development of promoters that act at fruit developing stages such as immature green and mature green periods. In this study, we report novel promoters for gene expression during the green to ripening stages of tomato fruit development. Genes specifically expressed at tomato fruit were selected using microarray data. Subsequent to confirmation of the expression of the selected 12 genes, upstream DNA fragments of the genes LA22CD07, Les.3122.2.A1_a_at and LesAffx.6852.1.S1_at which specifically expressed at fruit were isolated from tomato genomic DNA as promoter regions. Isolated promoter regions were fused with the GUS gene and the resultant constructs were introduced into tomato by agrobacterium-mediated transformation for evaluation of promoter activity in tomato fruit. The two promoters of LA22CD07, and LesAffx.6852.1.S1_at showed strong activity in the fruit, weak activity in the flower and undetectable activity in other tissues. Unlike well-known fruit-ripening specific promoters, such as the E8 promoter, these promoters exhibited strong activity in green fruit in addition to red-ripening fruit, indicating that the promoters are suitable for transgene expression during green to ripening stages of tomato fruit development. KEY MESSAGE: Novel fruit-specific promoters have been identified and are suitable for transgene expression during green to ripening stages of tomato fruit development.     

Identification and molecular mapping of loci controlling fruit ripening time in tomato

Using RAPD marker analysis, two quantitative trait loci (QTLs) associated with earliness due to reduced fruit-ripening time (days from anthesis to ripening = DTR) were identified and mapped in an F₂ population derived from a cross between Lycopersicon esculentum’E6203’ (normal ripening) and Lycopersicon esculentum’Early Cherry’ (early ripening). One QTL, on chromosome 5, was associated with a reduction in both ripening time (5 days) and fruit weight (29.3%) and explained 15.8 and 13% of the total phenotypic variation for DTR and fruit weight, respectively. The other QTL, on chromosome 12, was primarily associated with a reduction only in ripening time (7 days) and explained 12.3% of the total phenotypic variation for DTR. The gene action at this QTL was found to be partially dominant (d/a=0.41). Together, these two QTLs explained 25.1% of the total phenotypic variation for DTR. Additionally, two QTLs associated with fruit weight were identified in the same F₂ population and mapped to chromosomes 4 and 6, respectively. Together, these two QTLs explained 30.9% of the total phenotypc variation for fruit weight. For all QTLs, the ’Early Cherry’ alleles caused reductions in both ripening time and fruit weight. The polymorphic band for the most significant RAPD marker (OPAB-06), linked to the reduced ripening time QTL on chromosome 12, was converted to a cleaved amplified polymorphism (CAP) assay for marker-aided selection and further introgression of early ripening time (DTR) into cultivated tomato. Received: 15 March 1999 / Accepted: 29 April 1999     

Contrasted responses to carbohydrate limitation in tomato fruit at two stages of development

The metabolic consequences of long‐term carbohydrate depletion have been well documented in many sink organs but not extensively in fruit. Therefore, in the present study the response to sugar limitation in tomato fruit (Lycopersicon esculentum Mill.) was investigated at two developmental stages; during the cell division and cell expansion phases. First, the response in excised fruit cultured in vitro was characterized. Sugar depletion caused an arrest of growth and an exhaustion of carbon reserves. The proteins that were degraded and the nitrogen released was transiently stored as asparagine and glutamine in both developmental stages and also as γ ‐aminobutyric acid (GABA) in expanding fruit. Fruit at the cell division stage appeared to be more sensitive to sugar limitation. The response to sugar depletion was then characterized in fruit from plants submitted to extended darkness. In planta, the effects of sugar‐limitation were similar to those described in vitro but much more attenuated, especially in expanding fruit, which still accumulated dry matter. The expression of cell cycle genes, sugar‐ and nitrogen‐related genes was reduced by darkness. Only asparagine synthetase gene expression was induced in both dark‐treated fruit. Together the present data revealed that the effects of the carbon limitation are more pronounced in the youngest fruits as it is probably controlled by the relative sink strength of the fruit.     

Physiological and molecular analyses of early and late Coffea arabica cultivars at different stages of fruit ripening

Coffee quality is strongly influenced by a great number of factors, among which the fruit ripening stage at harvest time has a major influence on this feature. Studies comprising ethylene production and the regulation of ethylene biosynthesis genes during the ripening process indicate that ethylene plays an important role on coffee fruit ripening. Coffee early cultivars usually show a more uniform ripening process although little is known about the genetic factors that promote the earliness of ripening. Thus, in order to better understand the physiological and genetic factors involved in the regulation of ripening time, and consequently ripening uniformity, this study aimed to analyze ethylene and respiration patterns during coffee ripening, as well as to analyze ACC oxidase, an ethylene biosynthesis enzyme, gene expression, in fruits of early (Catucaí 785-15) and late (Acauã) coffee cultivars. Coffee fruits were harvested monthly from 124 days after flowering (end of February), until complete maturation (end of June). Dry matter, moisture content, color, respiratory rate and ethylene production analysis were performed. In silico analysis identified a coffee ACC oxidase gene (CaACO-like) and its expression was analyzed by real-time PCR. Dry matter and relative water content constantly increased and gradually decreased, respectively, during fruit ripening, and the color analysis enabled the observation of the earliness in the ripening process displayed by Catucaí 785-15 and its higher fruit ripening uniformity. The results obtained from the CaACO-like expression analysis and respiration and ethylene analysis suggest that the differences in ripening behavior between the two coffee cultivars analyzed in this study may be related to the differences in their capacity to produce ethylene, with fruits of Catucaí 785-15 and Acauã showing a typical and an attenuated climacteric phase, respectively, which may have lead to differences in their ripening time and uniformity.     

Function of the polygalacturonase convertor in ripening tomato fruit

In extracts from pericarp tissue of ripening tomato ( Lycopersicon esculentum Mill. cv, Sonato) fruits, two isoenzymes of polygalacturonase (E.C. 3.2.1.15), PG1 and PG2, are usually found. Also in such extracts, or as part of PG1, a convertor (CV) occurs. Incubation of PG2 with this CV gives rise to PG1 or a different isoenzyme, PGx, that is also stable at 65°C but differs in _pH optimum and size from PG1. It appears that CV has two affinity sites that can bind with PG2 or with a polydextran. PG1 is an extraction artifact, consisting of one molecule of CV and two molecules of PG2. PGx is made up of one molecule of CV and one molecule of PG2. It is the CV part of PGx that binds to polydextrans such as Blue Dextran 2000, Sephadex G-100, and cell wall preparations. In this last form PGx is the physiologically active form of the enzyme, solubilizing demethylated pectin.
On Sephacryl S-300, CV appears to have a molecular weight of 81 kDa, but because of its heat stability and partial leakage through a 10 kDa cut-off membrane, it might be a much smaller, rod-like molecule. The polygalacturonase convertor might be a lectin without intrinsic enzyme activity, with a function to immobilize, stabilize and activate enzymic proteins in the cell wall.     

Manipulation of the blue light photoreceptor cryptochrome 2 in tomato affects vegetative development, flowering time, and fruit antioxidant content

Cryptochromes are blue light photoreceptors found in plants, bacteria, and animals. In Arabidopsis, cryptochrome 2 (cry2) is involved primarily in the control of flowering time and in photomorphogenesis under low-fluence light. No data on the function of cry2 are available in plants, apart from Arabidopsis (Arabidopsis thaliana). Expression of the tomato (Solanum lycopersicum) CRY2 gene was altered through a combination of transgenic overexpression and virus-induced gene silencing. Tomato CRY2 overexpressors show phenotypes similar to but distinct from their Arabidopsis counterparts (hypocotyl and internode shortening under both low- and high-fluence blue light), but also several novel ones, including a high-pigment phenotype, resulting in overproduction of anthocyanins and chlorophyll in leaves and of flavonoids and lycopene in fruits. The accumulation of lycopene in fruits is accompanied by the decreased expression of lycopene beta-cyclase genes. CRY2 overexpression causes an unexpected delay in flowering, observed under both short- and long-day conditions, and an increased outgrowth of axillary branches. Virus-induced gene silencing of CRY2 results in a reversion of leaf anthocyanin accumulation, of internode shortening, and of late flowering in CRY2-overexpressing plants, whereas in wild-type plants it causes a minor internode elongation.     

Flavor volatilcs, sugars and color development in ripening in vitro-cultured tomato fruit and calyx

Previous studies showed that the developmental program of calyces of a tomato cultivar ( Lycopersicon esculentum , cv. VFNT Cherry) changed in many aspects to that of fruit when cultured in vitro. The calyces turned red, produced ethylene, had increased tissue content of 1-aminocyclopropane-1-carboxylic acid, had increased levels of the mRNA of polygalacturonase and developed ultrastructural changes in their cell walls that were indistinguishable from those of ripe tomato fruit tissue. We report in the present study the synthesis of volatile flavor compounds, changes in sugar concentrations and color development in cultured calyces that are characteristic of ripening tomato fruit. These ripening parameters of in vitro-cultured tomato fruit were also compared to those of fruit grown in the greenhouse.     

Altered chloroplast development and delayed fruit ripening caused by mutations in a zinc metalloprotease at the lutescent2 locus of tomato

The chloroplast is the site of photosynthesis in higher plants but also functions as the center of synthesis for primary and specialized metabolites including amino acids, fatty acids, starch, and diverse isoprenoids. Mutants that disrupt aspects of chloroplast function represent valuable tools for defining structural and biochemical regulation of the chloroplast and its interplay with whole-plant structure and function. The lutescent1 (l1) and l2 mutants of tomato (Solanum lycopersicum) possess a range of chlorophyll-deficient phenotypes including reduced rates of chlorophyll synthesis during deetiolation and enhanced rates of chlorophyll loss in leaves and fruits as they age, particularly in response to high-light stress and darkness. In addition, the onset of fruit ripening is delayed in lutescent mutants by approximately 1 week although once ripening is initiated they ripen at a normal rate and accumulation of carotenoids is not impaired. The l2 locus was mapped to the long arm of chromosome 10 and positional cloning revealed the existence of a premature stop codon in a chloroplast-targeted zinc metalloprotease of the M50 family that is homologous to the Arabidopsis (Arabidopsis thaliana) gene ETHYLENE-DEPENDENT GRAVITROPISM DEFICIENT AND YELLOW-GREEN1. Screening of tomato germplasm identified two additional l2 mutant alleles. This study suggests a role for the chloroplast in mediating the onset of fruit ripening in tomato and indicates that chromoplast development in fruit does not depend on functional chloroplasts.     

Tissue dependent variations of DNA methylation and endoreduplication levels during tomato fruit development and ripening

Tomato fruit cells are characterized by a strong increase in nuclear ploidy during fruit development. Average ploidy levels increased to similar levels (above 50C) in two distinct fruit tissues, pericarp and locular tissue. However, ploidy profiles differed significantly between these two tissues suggesting a tissue-specific control of endoreduplication in tomato fruit. To determine possible relationships between endoreduplication and epigenetic mechanisms, the methylation status of genomic DNA from pericarp and locular tissue of tomato fruit was analysed. Pericarp genomic DNA was characterized by an increase of CG and/or CNG methylation at the 5S and 18S rDNA loci and at gyspsy-like retrotransposon sequences during fruit growth. A sharp decrease of the global DNA methylation level together with a reduction of methylation at the rDNA loci was also observed in pericarp during fruit ripening. Inversely, no major variation of DNA methylation either global or locus-specific, was observed in locular tissue. Thus, tissue-specific variations of DNA methylation are unlikely to be triggered by the induction of endoreduplication in fruit tissues, but may reflect tissue-specific ploidy profiles. Expression analysis of eight putative tomato DNA methyltransferases encoding genes showed that one chromomethylase (CMT) and two rearranged methyltransferases (DRMs) are preferentially expressed in the pericarp during fruit growth and could be involved in the locus-specific increase of methylation observed at this developmental phase in the pericarp.     

基于生理发育时间的加工番茄生育期模拟模型

综合考虑温度和光照对加工番茄生理发育效应的影响,引入了品种基本发育因子（IDF）,通过分析不同类型加工番茄的生育期与环境因素的动态关系,建立了基于生理发育时间（PDTv）的加工番茄生育期模拟模型,并利用不同年份、生态区、品种、种植方式的试验资料对模型进行了检验.结果表明：所建模型对加工番茄从播种到各个发育阶段（出苗、开花、坐果、红熟和拉秧）天数的模拟值与观测值的回归估计标准误差（RMSE）分别为1.09、2.03、2.05、2.77和2.53 d,该模型的预测精度明显高于基于有效积温的发育模型（其RMSE分别为1.90、6.63、6.33、9.36 和6.84 d）.     

Ethylene receptor degradation controls the timing of ripening in tomato fruit

Fruit ripening in tomato requires the coordination of both developmental cues and the phytohormone ethylene. The multigene ethylene receptor family has been shown to negatively regulate ethylene signal transduction and suppress ethylene responses. Here we demonstrate that reduction in the levels of either of two family members, LeETR4 or LeETR6, causes an early-ripening phenotype. We provide evidence that the receptors are rapidly degraded in the presence of ethylene, and that degradation probably occurs through the 26S proteasome-dependent pathway. Ethylene exposure of immature fruits causes a reduction in the amount of receptor protein and earlier ripening. The results are consistent with a model in which receptor levels modulate timing of the onset of fruit ripening by measuring cumulative ethylene exposure.     

Effect of the Cnr mutation on carotenoid formation during tomato fruit ripening

The characteristic pigmentation of ripe tomato fruit is due to the deposition of carotenoid pigments. In tomato, numerous colour mutants exist. The Cnr tomato mutant has a colourless, non-ripening phenotype. In this work, carotenoid formation in the Cnr mutant has been studied at the biochemical level. The carotenoid composition of Ailsa Craig (AC) and Cnr leaves was qualitatively and quantitatively similar. However, Cnr fruits had low levels of total carotenoids and lacked detectable levels of phytoene and lycopene. The presence of normal tocopherols and ubiquinone-9 levels in the ripe Cnr fruits suggested that other biosynthetically related isoprenoids were unaffected by the alterations to carotenoid biosynthesis. In vitro assays confirmed the virtual absence of phytoene synthesis in the ripe Cnr fruit. Extracts from ripe fruit of the Cnr mutant also revealed a reduced ability to synthesise the carotenoid precursor geranylgeranyl diphosphate (GGPP). These results suggest that besides affecting the first committed step in carotenoid biosynthesis (phytoene synthase) the Cnr mutation also affects the formation of the isoprenoid precursor (GGPP).