Cell wall metabolism during maturation, ripening and senescence of peach fruit

Cell wall changes were examined in fruit of a melting flesh peach (Prunus persica L.) allowed to ripen on the tree. Three phases to softening were noted, the first of which began prior to the completion of flesh colour change and an increase in ethylene evolution. Softening in young mature fruit, prior to ripening, was associated with a depolymerization of matrix glycans both loosely and tightly attached to cellulose and a loss of Gal from all cell wall fractions. After the initiation of ripening, but before the melting stage, softening was associated with continuing, progressive depolymerization of matrix glycans. A massive loss of Ara from the loosely bound matrix glycan fraction was observed, probably from side chains of glucuronoarabinoxylan, pectin, or possibly arabinogalactan protein firmly bound into the wall and solubilized in this extract. An increase in the solubilization of polyuronides also occurred during this period, when softening was already well advanced. The extensive softening of the melting period was marked by substantial depolymerization of both loosely and tightly bound matrix glycans, including a loss of Ara from the latter, an increase in matrix glycan extractability, and a dramatic depolymerization of chelator-soluble polyuronides which continued during senescence. Depolymerization of chelator-soluble polyuronides thus occurred substantially after the increase in their solubilization. Ripening-related increases were observed in the activities of exo- and endo-polygalacturonase (EC 3.2.1.67; EC 3.2.1.15), pectin methylesterase (EC 3.1.1.11), endo-1,4-beta-glucanase (EC 3.2.1.4), endo-1,4-beta-mannanase (EC 3.2.1.78), alpha-arabinosidase (EC 3.2.1.55), and beta-galactosidase (EC 3.2.1.23), but the timing and extent of the increases differed between enzymes and was not necessarily related to ethylene evolution. Fruit softening in peach is a continuous process and correlated closely with the depolymerization of matrix glycans, which proceeded throughout development. However, numerous other cell wall changes also took place, such as the deglycosylation of particular polymers and the solubilization and depolymerization of chelator-soluble polyuronides, but these were transient and occurred only at specific phases of the softening process. Fruit softening and other textural changes in peach appear to have a number of stages, each involving a different set of cell wall modifications.     

Exopolygalacturonase protein accumulates late in peach fruit ripening

Two exo-acting polygalacturonase enzymes (exoPG, EC 3.2.1.67) increase in activity as peach ( Prunu persica L. Batsch cvs Coronet and Flavorcrest) fruits ripen. By examining populations of fruit, we show that the increase in activity occurs late in ripening when the fruit are very soft (below 2 kgf). The more abundant form of the enzyme, exoPG 2, was extensively purified and analyzed for its amino acid content and N-terminal amino acid sequence. ExoPG 2 is a polypeptide of M, 66 000 and has a substantial excess of basic over acidic amino acids. Polyclonal antisera to exoPG 2 were raised in mice. The antisera inhibited the enzyme activity and recognized a M_r 66 000 polypeptide in Western blots. Western blot analyses of extracts of fruit ranked for softness revealed a M_r 66 000 polypeptide only in the softest fruit (less than 2.5 kgf). We conclude that the increased in exopolygalacturonase activity that occurs in very soft fruit is due to an increase in the amount of enzyme protein.     

膨大期果面喷施IAA对桃成熟期果实性状和相关基因表达的影响北大核心CSCD

为明确桃果实发育后期外源生长素对果实成熟的影响及更深入的认识生长素调控果实发育与成熟的机制,本研究以桃品种‘小白凤’为试材,采用不同浓度的外源IAA(200,10,0.1μmol/L)喷施第2次快速膨大期的桃果实,在处理后10,20,30 d分别取样,分析桃果实的硬度,糖组分(蔗糖、葡萄糖、果糖、山梨醇),果胶,纤维素含量以及乙烯释放量的变化,并对200μmol/LIAA处理后30 d及对照组的果实进行了转录组测序分析,深入认识生长素调控桃果实成熟的作用和机理。结果表明,(1)IAA处理后30 d时,200μmol/L处理组桃果实果肉硬度与对照组相比显著增加,成熟期平均延迟5 d,而10,0.1μmol/LIAA处理组与同期对照组间无显著差异。(2)果肉蔗糖含量在0.1,200μmol/LIAA处理后30 d显著低于同期对照组,在10μmol/LIAA处理后30 d时与对照组无显著差异;葡萄糖、果糖和山梨醇含量在各浓度IAA处理后30 d时多与对照组间无明显差异。(3)在处理30 d时,果肉中可溶性果胶含量在0.1μmol/L IAA处理组与对照组无显著差异,在10,200μmol/LIAA处理组均显著低于对照组,此时不同浓度IAA处理桃果肉中不可溶果胶和纤维素含量均与对照组之间无显著差异。(4)在IAA处理后10,20,30 d时,果实乙烯释放量均表现为0.1,10μmol/L处理与对照组无显著差异,200μmol/L处理组显著低于对照组。(5)转录组数据显示,200μmol/L IAA处理与对照组中共存在86个差异表达基因,KEGG分析发现其中有6个与果实发育成熟密切相关的代谢途径中多个基因的表达谱发生显著变化,且变化趋势与已测定的生理指标数据吻合。综合分析表明200μmol/L的外源IAA处理第2次快速膨大期的桃果实能够延缓果实成熟的进程。     

Effect of seed on ripening control components during avocado fruit development

Seedless avocado fruit are produced alongside seeded fruit in the cultivar Arad, and both reach maturity at the same time. Using this system, it was possible to show that avocado seed inhibits the ripening process: seedless fruits exhibited higher response to exogenous ethylene already at the fruitlet stage, and also at the immature and mature fruit stages. They produced higher CO₂ levels, and the ethylene peak was apparent at the fruitlet stage of seedless fruit, but not of seeded ones. The expression levels of PaETR, PaERS1 and PaCTR1 on the day of harvest at all developmental stages were very similar between seeded and seedless fruit, except that PaCTR1 was higher in seedless fruit only at very early stages. This expression pattern suggests that the seed does not have an effect on components of the ethylene response pathway when fruits are just picked. The expression of MADS-box genes, PaAG1 and PaAGL9, preceded the increase in ethylene production of mature seeded fruit, but not at earlier stages. However, only PaAGL9 was induced in seedless fruit at early stages of development. Taken together, these data suggest that these genes are perhaps involved in climacteric response in seeded fruit, and the seed is responsible for their induction at normal fruit ripening.     

Effect of salinity on tomato fruit ripening

Tomato (Lycopersicon esculentum Mill) plants from various cultivars growing on half-strength Hoagland solution were exposed at anthesis to 3 or 6 grams per liter NaCl. Salinity shortened the time of fruit development by 4 to 15%. Fruits of salt-treated plants were smaller and tasted better than did fruits of control plants. This result was obtained both for ripe fruits tested on the day of picking and for those picked at 100% development and allowed to ripen at room temperature for 9 days. Percentage of dry weight, total soluble solids, and titratable acidity; content of reducing sugars, Cl⁻, Na⁺, and various pericarp pigments; and electrical conductivity of the juice were higher in fruits of saline-treated plants than they were in those of control plants, while the pH was lower. Ethylene and CO₂ evolution rates during ripening; as well as the activities of pectin methyl esterase, polymethylgalacturonase, and polygalacturonase; were also higher in fruits of the saline-treated plants. The treatment with 6 grams per liter NaCl shortened the fruit shelf life considerably.     

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).     

Effect of pre-storage treatments on mango fruit ripening

The effectiveness of pre-storage treatments of nitrogen (low oxygen), heat and ethanol and acetaldehyde vapours were examined for their potential for improving mango storage. Mature green mango fruit (Mangifera indica L. cv. Keitt) were treated with low oxygen (< 3% oxygen, 97% nitrogen) for 72 h, acetaldehyde (0.12%) and ethanol (1%) vapours for 24 h or heat (38 ± 2°C) for 48 h prior to storage at 14°C. The nitrogen and ethanol treatments induced substantial levels of acetaldehyde and ethanol in the fruit. Initially the firmness of the nitrogen treated fruit remained higher than the control although later in storage this effect was lost. Differences in ripening were reflected in the total soluble solids and acidity levels, nitrogen maintaining a higher acidity and lower total soluble solids (less mature) whereas the heat treated fruit had lower acidity and higher total soluble solids (more mature). Ethanol and acetaldehyde treatments showed no effect. The use of a pre-storage treatment of nitrogen therefore had a beneficial effect on retarding ripening, although as storage progressed this effect was lost.     

Regulation of the protein and gene expressions of ethylene biosynthesis enzymes under different temperature during peach fruit ripening

Ethylene has profound effect on fruit development and ripening, and the role of ethylene biosynthesis enzymes involving 1-aminocyclopropane-1-carboxylic acid (ACC) synthase (ACS), ACC oxidase (ACO), and S-Adenosyl-l-methionine synthetase (SAMS) in peach fruit (cv. Xiahui-8) was characterized under 25 and 4 °C, respectively. All these enzymes in ethylene synthesis pathway were identified using 2-DE and real-time PCR. Both protein and gene expressions of ACO and SAMS were much higher at 25 °C than at 4 °C. Among five members of ACS family, PpaACS4 may belong to system II ethylene biosynthesis, while PpaACS3 involved in system I during development stage, and low temperature can induce PpaACS1 expression. The ethylene release and low expressions of proteins and genes of most enzymes indicated that low temperature can effectively postpone ripening stage by reducing ethylene evolution. High gene expression of PpaSAMS did not cause excessive expression of SAMS protein under low temperature, and over-expression of PpaACS1 at low temperature still did not induce increase of ethylene production. The mechanism underlying the phenomenon about how temperature affects ethylene release was also discussed.     

Genome-wide identification and expression analysis of the lipoxygenase gene family during peach fruit ripening under different postharvest treatments

In plants, lipoxygenase (LOX), facilitated by the LOX family genes is closely related to fruit ripening and senescence, but research on LOX in peach fruit is limited. To study the roles of LOX family genes in fruit ripening during storage, a comprehensive overview of the LOX gene family in peach is presented, including their phylogenetic relationships, gene structures and subcellular localizations. Additionally, the fruit quality, including fruit firmness, ethylene production and soluble solids content under different storage conditions, were assessed. Finally, 12 peach genes that encode LOX proteins have been identified, and comparisons of the PpaLOX gene expression levels under different postharvest treatments in peach fruit suggest that PpaLOX2.1, PpaLOX7.1, PpaLOX7.2, and especially PpaLOX2.2, may be required in peach fruit ripening during storage. The results will be useful to further analyze the functions of the LOX family of genes in plants.     

Synthesis of polygalacturonase during tomato fruit ripening

The cell wall degrading enzyme polygalacturonase (E.C. 3.2.1.15) is not detectable in green tomatoes (Lycopersicon esculentum Mill). Activity appears at the onset of ripening and in ripe fruit it is one of the major cell-wall-bound proteins. Radioimmunoassay results, employing an antibody against purified polygalacturonase, suggest that during ripening the enzyme is synthesised de novo. Radioimmunoassay data also show that the low level of polygalacturonase in Never ripe mutants and the lack of activity in ripening inhibitor mutants can be correlated to the levels of immunologically detectable polygalacturonase protein.Abbreviations PG polygalacturonase - Nr Never ripe mutation - rin ripening inhibitor mutation     

Downregulation of RdDM during strawberry fruit ripening

Background

Recently, DNA methylation was proposed to regulate fleshy fruit ripening. Fleshy fruits can be distinguished by their ripening process as climacteric fruits, such as tomatoes, or non-climacteric fruits, such as strawberries. Tomatoes undergo a global decrease in DNA methylation during ripening, due to increased expression of a DNA demethylase gene. The dynamics and biological relevance of DNA methylation during the ripening of non-climacteric fruits are unknown.

Results

Here, we generate single-base resolution maps of the DNA methylome in immature and ripe strawberry. We observe an overall loss of DNA methylation during strawberry fruit ripening. Thus, ripening-induced DNA hypomethylation occurs not only in climacteric fruit, but also in non-climacteric fruit. Application of a DNA methylation inhibitor causes an early ripening phenotype, suggesting that DNA hypomethylation is important for strawberry fruit ripening. The mechanisms underlying DNA hypomethylation during the ripening of tomato and strawberry are distinct. Unlike in tomatoes, DNA demethylase genes are not upregulated during the ripening of strawberries. Instead, genes involved in RNA-directed DNA methylation are downregulated during strawberry ripening. Further, ripening-induced DNA hypomethylation is associated with decreased siRNA levels, consistent with reduced RdDM activity. Therefore, we propose that a downregulation of RdDM contributes to DNA hypomethylation during strawberry ripening.

Conclusions

Our findings provide new insight into the DNA methylation dynamics during the ripening of non-climacteric fruit and suggest a novel function of RdDM in regulating an important process in plant development.

     

Mitochondrial enzymes in mango fruit during ripening

Mitochondria isolated from immature (developing), mature (unripe), and ripe mango pulp actively oxidized the intermediates of the Krebs cycle. The oxidation of citrate, oxoglutarate, succinate and malate by both unripe and ripe fruit mitochondria was several fold greater than that by mitochondria from immature fruit. The levels of malic dehydrogenase and succinic dehydrogenase increased with the onset of ripening, whereas the level of citrate synthase increased several fold on maturation but decreased six-fold on ripening. Isocitrate dehydrogenase and malic enzyme were very high in the immature fruit but after a sudden decrease in the matured fruit showed a considerable rise thereafter. The ratio of the activities of isocitrate lyase to isocitrate dehydrogenase is considerably higher in the immature fruit and greatest in the unripe (mature) fruit. This, together with a higher concentration of glyoxylate at these stages, indicate the operation of the glyoxylate bypass. Oxidized and reduced forms of pyridine nucleotides were estimated.     

Gene expression during fruit ripening in avocado

The poly(A) ⁺RNA populations from avocado fruit (Persea americana Mill cv. Hass) at four stages of ripening were isolated by two cycles of oligo-dT-cellulose chromatography and examined by invitro translation, using the rabbit reticulocyte lysate system, followed by two-dimensional gel electrophoresis (isoelectric focusing followed by sodium dodecyl sulfate polyacrylamide gel electrophoresis) of the resulting translation products. Three mRNAs increased dramatically with the climacteric rise in respiration and ethylene production. The molecular weights of the corresponding translation products from the ripening-related mRNAs are 80,000, 36,000, and 16,500. These results indicate that ripening may be linked to the expression of specific genes.     

Polygalacturonase activity and variation in ripening of papaya fruit with tissue depth and heat treatment

Effects of tissue position (viz. outer vs inner mesocarp) and heat treatment (48°C, 20 min) on variations in polygalacturonase (EC 3.2.1.15 and EC 3.2.1.67) activity and ripening of fruits of Carica papaya L. cv. Backcross Solo were investigated. Polygalacturonase activity increased during ripening concomitantly with an increase in tissue softness and soluble polyuronide level. Throughout ripening, inner mesocarp tissue was softer and contained higher polygalacturonase activity than outer mesocarp tissue. Titratable acidity as well as ß-galactosidase (EC 3.2.1.23) activity also increased during ripening; however, unlike polygalacturonase, their level or activity was lower in inner than in outer mesocarp. Ascorbic acid could partially account for the increase in titratable acidity during ripening but contributed very little to the differences in titratable acid levels between outer and inner mesocarp. Heat treatment had no effect on either fruit softness or titratable acidity, but it markedly reduced the increase in ascorbic acid and polygalacturonase activity during ripening. Ripening, as reflected by changes in tissue softness and polygalacturonase activity, progressed outwardly from the interior towards the exterior of the fruit. The effect of heat treatment in suppressing polygalacturonase activity was relatively greater in inner than in outer mesocarp, suggesting that sensitivity of the enzyme to heat treatment may vary with stage of ripeness of the tissue.     

Pectate lyase activity during ripening of banana fruit

Pectate lyase (PEL) activity was demonstrated in ripe banana fruits on supplementing the homogenizing medium with cysteine and Triton X-100. The enzyme was characterized on the basis of alkaline pH optimum, elimination of the activity by EDTA and activation by Ca(2+). PEL activity was not detected in preclimacteric banana fruits. PEL activity increased progressively from early climacteric and reached maximum level at climacteric peak and declined in post climacteric and over ripened fruits. Replacing pectate with pectin in PEL assay manifested enzyme activity even in preclimacteric fruits. In contrast to PEL, polygalacturonase activity progressively increased during fruit ripening even in postclimacteric fruits.