Climacteric Ripening Research Articles

Expression of auxin-binding protein1 during plum fruit ontogeny supports the potential role of auxin in initiating and enhancing climacteric ripening

Auxin-binding protein1 (ABP1) is an active element involved in auxin signaling and plays critical roles in auxin-mediated plant development. Here, we report the isolation and characterization of a putative sequence from Prunus salicina L., designated PslABP1. The expected protein exhibits a similar molecular structure to that of well-characterized maize-ABP1; however, PslABP1 displays more sequence polarity in the active-binding site due to substitution of some crucial amino-acid residues predicted to be involved in auxin-binding. Further, PslABP1 expression was assessed throughout fruit ontogeny to determine its role in fruit development. Comparing the expression data with the physiological aspects that characterize fruit-development stages indicates that PslABP1 up-regulation is usually associated with the signature events that are triggered in an auxin-dependent manner such as floral induction, fruit initiation, embryogenesis, and cell division and elongation. However, the diversity in PslABP1 expression profile during the ripening process of early and late plum cultivars seems to be due to the variability of endogenous auxin levels among the two cultivars, which consequently can change the levels of autocatalytic ethylene available for the fruit to co-ordinate ripening. The effect of auxin on stimulating ethylene production and in regulating PslABP1 was investigated. Our data suggest that auxin is involved in the transition of the mature green fruit into the ripening phase and in enhancing the ripening process in both auxin- and ethylene-dependent manners thereafter.

ACC-OXIDASE IS THE RATE LIMITING STEP IN ETHYLENE BIOSYNTHESIS DURING POSTHARVEST STORAGE OF TOMATO

It has been well documented in literature that ACC-synthase is the rate limiting step in ethylene biosynthesis during climacteric ripening of tomato. This manuscript investigates how the ethylene metabolism further evolves during post harvest storage of tomato. The current data show how ethylene production declines after tomato fruit reaches the red stage. Metabolite screening revealed that ACC and MACC are highly abundant in red ripe and stored fruits. Surprisingly, although ACC-oxidase in-vitro activity was declining, ACC-synthase in vitro activity was increasing again during postharvest storage. These data suggest that the decline in ethylene production during storage must be regulated at the level of ACC-oxidase. The exact reason why ACC-synthase is still highly active during these stages remains peculiar and needs further investigation.

Role of Brassinosteroids, Ethylene, Abscisic Acid, and Indole-3-Acetic Acid in Mango Fruit Ripening

Rapid ripening of mango fruit limits its distribution to distant markets. To better understand and perhaps manipulate this process, we investigated the role of plant hormones in modulating climacteric ripening of ‘Kensington Pride’ mango fruits. Changes in endogenous levels of brassinosteroids (BRs), abscisic acid (ABA), indole-3-acetic acid (IAA), and ethylene and the respiration rate, pulp firmness, and skin color were determined at 2-day intervals during an 8-day ripening period at ambient temperature (21 ± 1°C). We also investigated the effects of exogenously applied epibrassinolide (Epi-BL), (+)-cis, trans-abscisic acid (ABA), and an inhibitor of ABA biosynthesis, nordihydroguaiaretic acid (NDGA), on fruit-ripening parameters such as respiration, ethylene production, fruit softening, and color. Climacteric ethylene production and the respiration peak occurred on the fourth day of ripening. Castasterone and brassinolide were present in only trace amounts in fruit pulp throughout the ripening period. However, the exogenous application of Epi-BL (45 and 60 ng g−1 FW) advanced the onset of the climacteric peaks of ethylene production and respiration rate by 2 and 1 day, respectively, and accelerated fruit color development and softening during the fruit-ripening period. The endogenous level of ABA rose during the climacteric rise stage on the second day of ripening and peaked on the fourth day of ripening. Exogenous ABA promoted fruit color development and softening during ripening compared with the control and the trend was reversed in NDGA-treated fruit. The endogenous IAA level in the fruit pulp was higher during the preclimacteric minimum stage and declined during the climacteric and postclimacteric stages. We speculate that higher levels of endogenous IAA in fruit pulp during the preclimacteric stage and the accumulation of ABA prior to the climacteric stage might switch on ethylene production that triggers fruit ripening. Whilst exogenous Epi-BL promoted fruit ripening, endogenous measurements suggest that changes in BRs levels are unlikely to modulate mango fruit ripening.

Transcriptome and Metabolite Profiling Show That APETALA2a Is a Major Regulator of Tomato Fruit Ripening

Fruit ripening in tomato (Solanum lycopersicum) requires the coordination of both developmental cues as well as the plant hormone ethylene. Although the role of ethylene in mediating climacteric ripening has been established, knowledge regarding the developmental regulators that modulate the involvement of ethylene in tomato fruit ripening is still lacking. Here, we show that the tomato APETALA2a (AP2a) transcription factor regulates fruit ripening via regulation of ethylene biosynthesis and signaling. RNA interference (RNAi)-mediated repression of AP2a resulted in alterations in fruit shape, orange ripe fruits, and altered carotenoid accumulation. Microarray expression analyses of the ripe AP2 RNAi fruits showed altered expression of genes involved in various metabolic pathways, such as the phenylpropanoid and carotenoid pathways, as well as in hormone synthesis and perception. Genes involved in chromoplast differentiation and other ripening-associated processes were also differentially expressed, but softening and ethylene biosynthesis occurred in the transgenic plants. Ripening regulators RIPENING-INHIBITOR, NON-RIPENING, and COLORLESS NON-RIPENING (CNR) function upstream of AP2a and positively regulate its expression. In the pericarp of AP2 RNAi fruits, mRNA levels of CNR were elevated, indicating that AP2a and CNR are part of a negative feedback loop in the regulation of ripening. Moreover, we demonstrated that CNR binds to the promoter of AP2a in vitro.

INDUCTION OF SMALL HEAT-SHOCK PROTEINS IN MESOCARP OF CHERIMOYA FRUIT (ANNONA CHERIMOLAMILL.) PRODUCES CHILLING TOLERANCE

Cherimoya is a major crop in the subtropical areas of the Spanish coast. The fruit are perishable because of their rapid climacteric ripening. Storage between 10 and 12C produces some extension of the shelf life of the fruit as a small delay in senescence is produced. A lower storage temperature produces chilling injury. The aim of this work is to find a way to make the fruit more resistant to low temperatures. In several plant species the synthesis of small molecular size heat-shock proteins (sHSPs) prevents and/or repairs stress induced damage; therefore, we investigated the induction of sHSPs in the mesocarp of cherimoya fruit. Heating the fruit at 55C during 5 h produces alleviation of the chilling-injury symptoms when the fruit are stored at 4C. The thermal stress induces proteins recognized by antibodies against sHSPs of different classes from other plant species. This induction is proportional to the temperature in the experimental range of 46–50C. PRACTICAL APPLICATIONS Refrigeration is the only described way to extent shelf life of cherimoya as senescence is delayed by the decreasing temperature. This is true until chilling injury appears. Senescence of this fruit occurs in 4–5 days when stored at 20C, which is a regular temperature in the production area. Storage at 10–12C allows commercialization during one additional week. Different attempts have been made to extend the period of conservation by using controlled atmosphere and lower temperatures. The problem is that cherimoya fruit are very susceptible to chilling injury. If a short heating treatment induces small molecular size heat-shock proteins (sHSPs), which would further protect the fruit from chilling injury, this would probably allow a lower temperature for transportation and conservation, thus increasing the period for commercialization. Exportation could be done in a safer way.

TOMATO AGAMOUS‐LIKE 1 is a component of the fruit ripening regulatory network

After fertilization, the expanding carpel of fleshy fruit goes through a phase change to ripening. Although the role of ethylene signalling in mediating climacteric ripening has been established, knowledge regarding the regulation of ethylene biosynthesis and its association with fruit developmental programs is still lacking. A functional screen of tomato transcription factors showed that silencing of the TOMATO AGAMOUS-LIKE 1 (TAGL1) MADS box gene results in altered fruit pigmentation. Over-expressing TAGL1 as a chimeric repressor suggested a role in controlling ripening, as transgenic tomato fruit showed reduced carotenoid and ethylene levels, suppressed chlorophyll breakdown, and down-regulation of ripening-associated genes. Moreover, fruits over-expressing TAGL1 accumulated more lycopene, and their sepals were swollen, accumulated high levels of the yellow flavonoid naringenin chalcone and contained lycopene. Transient promoter-binding assays indicated that part of the TAGL1 activity in ripening is executed through direct activation of ACS2, an ethylene biosynthesis gene that has recently been reported to be a target of the RIN MADS box factor. Examination of the TAGL1 transcript and its over-expression in the rin mutant background suggested that RIN does not regulate TAGL1 or vice versa. The results also indicated RIN-dependent and -independent processes that are regulated by TAGL1. We also noted that fruit of TAGL1 loss-of-function lines had a thin pericarp layer, indicating an additional role for TAGL1 in carpel expansion prior to ripening. The results add a new component to the current model of the regulatory network that controls fleshy fruit ripening and its association with the ethylene biosynthesis pathway.

RESPONSE OF 1‐METHYLCYCLOPROPENE TREATED “GRANNY SMITH” APPLE FRUIT TO AIR AND CONTROLLED ATMOSPHERE STORAGE CONDITIONS

ABSTRACT In the study, the efficacy of 1‐Methylcyclopropene (1‐MCP) on the quality of apples cv. “Granny Smith” was examined. Fruits were stored for 120 and 180 days at 0 ± 0.5C and 90 ± 5% relative humidity (RH) under normal (NA) and controlled atmosphere (CA), and samples taken at 120 and 180 days of storage were kept at 20 ± 2C and 60 ± 5% RH for 7 days. Weight loss was higher in fruits stored under NA. The lowest respiration rates were in CA while the highest rates were found in NA‐stored fruits without pretreated. 1‐MCP + CA reduced the internal ethylene concentration from 10.63 µL/L to 0.60 µL. The incidence and severity of superficial scald was 68.10 and 77.00% in fruits stored in NA. 1‐MCP consistently suppressed the expression of superficial scald. 1‐MCP maintained the quality fruit kept in CA and NA environments to a higher degree than nontreated apples over a 180‐day storage period. The results indicated that cv. “Granny Smith” could be stored successfully under 1‐MCP + CA conditions and that 1‐MCP + NA may be a viable alternative to CA for optimal eating quality.PRACTICAL APPLICATIONS1‐Methylcyclopropene (1‐MCP) is an effective inhibitor of ethylene action and it binds irreversibly to ethylene receptors. 1‐MCP delays the onset of the climacteric ripening of fruits. Recently research has been conducted on the effects of 1‐MCP on postharvest behavior of fruits and vegetables with the objective of reducing physiological disorders and quality losses to prolong their commercial life. The use of 1‐MCP is easy. It is important that the fruit treated with 1‐MCP reach a good sensory quality at the commercial level. Superficial scald of apples is a physiological disorder, which following long‐term storage has the potential to destroy the market value and utility of millions of tons of fresh apples annually. The present work evaluated a prestorage conditioning treatment to protect against this physiological disorder. Results will allow the long storage of apple cv. “Granny Smith,” benefiting growers, exporters, shippers and distributors of this horticultural product.

Candidate genes and QTLs for fruit ripening and softening in melon

Different factors affect the quality of melon fruit and among them long shelf life is critical from the consumer's point of view. In melon, cultivars showing both climacteric and non-climacteric ripening types are found. In this study we have investigated climacteric ripening and fruit softening using a collection of near-isogenic lines (NILs) derived from the non-climacteric melon parental lines PI 161375 (SC) and "Piel de Sapo" (PS). Surprisingly, we found that QTL eth3.5 in NIL SC3-5b induced a climacteric-ripening phenotype with increased respiration and ethylene levels. Data suggest that the non-climacteric phenotypes from PI 161375 and "Piel de Sapo" may be the result of mutations in different genes. Several QTLs for fruit flesh firmness were also detected. Candidate genes putatively involved in ethylene regulation, biosynthesis and perception and cell wall degradation were mapped and some colocations with QTLs were observed. These results may provide additional data towards understanding of non-climacteric ripening in melon.

Regulatory Role of Cystathionine-γ-Synthase and de novo Synthesis of Methionine in Ethylene Production during tomato Fruit Ripening

The essential amino acid methionine is a substrate for the synthesis of S-adenosyl-methionine (SAM), that donates its methyl group to numerous methylation reactions, and from which polyamines and ethylene are generated. To study the regulatory role of methionine synthesis in tomato fruit ripening, which requires a sharp increase in ethylene production, we cloned a cDNA encoding cystathionine gamma-synthase (CGS) from tomato and analysed its mRNA and protein levels during tomato fruit ripening. CGS mRNA and protein levels peaked at the "turning" stage and declined as the fruit ripened. Notably, the tomato CGS mRNA level in both leaves and fruit was negatively affected by methionine feeding, a regulation that Arabidopsis, but not potato CGS mRNA is subject to. A positive correlation was found between elevated ethylene production and increased CGS mRNA levels during the ethylene burst of the climacteric ripening of tomato fruit. In addition, wounding of pericarp from tomato fruit at the mature green stage stimulated both ethylene production and CGS mRNA level. Application of exogenous methionine to pericarp of mature green fruit increased ethylene evolution, suggesting that soluble methionine may be a rate limiting metabolite for ethylene synthesis. Moreover, treatment of mature green tomato fruit with the ethylene-releasing reagent Ethephon caused an induction of CGS mRNA level, indicating that CGS gene expression is regulated by ethylene. Taken together, these results imply that in addition to recycling of the methionine moieties via the Yang pathway, operating during synthesis of ethylene, de novo synthesis of methionine may be required when high rates of ethylene production are induced.

Monitoring gene expression along pear fruit development, ripening and senescence using cDNA microarrays

In order to elucidate the molecular events associated with pear ( Pyrus communis L. cv. Rocha) fruit development and climacteric ripening, high-density cDNA microarrays were constructed. cDNA clones (1364 total) were isolated from a fruit cDNA library and from a subtractive library and arrayed. The expression of these ESTs was monitored from early fruit development, through ripening, until complete fruit senescence, allowing a global coverage of the entire fruit life. A comparison was made with fruit that failed to ripe (FR) due to precocious harvesting. Based on the similarities in transcript expression profiles ESTs were grouped in different clusters. Transcripts encoding kinases and phosphatases were induced specifically during early developmental stages of pear fruit. Another set of genes was activated at the onset of the climacteric period when fruit softening rates also increased. Among these were transcripts encoding for cell wall modifications and pigment and aroma biosynthesis. Transcripts putatively involved in defence response, oxidative stress, primary and secondary metabolism, signalling and transcription regulation were also isolated. According to our results main changes in expression profiles occur in two phases: the cessation of growth and entrance in the initial maturation stages (pre-climacteric), and the entrance into the climacteric period.

HARVEST MATURITY RELATIONSHIP TO FLAVOR REGENERATION AFTER CA-STORAGE OF 'DELICIOUS' APPLES

Red Delicious producers in the USA are under increasing pressure to produce fruit with optimum “on-shelf” dessert quality following storage. In addition to traditional quality measures like firmness and soluble solids, we are studying the ripening-related events that affect flavor perception. Ester production is always closely linked to the onset of climacteric ripening, while prolonged low-oxygen storage is usually detrimental to volatile production. Experiments focused on optimum harvest time for maintaining sweetness, sourness and aroma generating capacity during CA storage. In multiple-harvest experiments with ‘Redchief Delicious’ apples, we investigated the relationship between the internal ethylene maturity indicator and flavor retention and regeneration after storage for different lengths of time. Using a cohort of untrained panelists, it was found that overall flavor perception and perceived fruit ripeness begins to increase at the onset of the climacteric. CA conditions reduce post-storage volatile production when compared to those stored in refrigerated air although not to a level below those displayed in “over-mature” fruit at harvest after 3 months in storage. As harvest maturity advanced, the time required to regenerate aroma volatiles to an “optimum” level after removal from CA storage decreased markedly. Apparently, there is a linear relationship between attainment of optimal eating quality and time out of storage. Firmness, soluble solids, and titratable acidity of fruit from all harvest dates remained at acceptable levels throughout the post-storage ripening period.

Relationship of harvest maturity to flavor regeneration after CA storage of ‘Delicious’ apples

Red Delicious producers in the USA face increased pressure to produce fruit with optimum ‘on-shelf’ dessert quality following storage. In addition to firmness and soluble solids measurement other ripening-related events affect flavor perception. Ester production is always closely linked to the onset of climacteric ripening, while prolonged low-oxygen storage is usually detrimental to volatile production. Experiments focused on timing of the optimum harvest for maintaining sweetness, sourness and aroma generating capacity during CA storage. In multiple-harvest experiments with ‘Redchief Delicious’ apples, we investigated the relationship between the internal ethylene maturity indicator and flavor retention and regeneration after storage for different lengths of time. Using a cohort of untrained panelists, it was found that overall flavor perception and perceived fruit ripeness begins to increase at the onset of the climacteric. CA conditions reduce post-storage volatile production when compared with those stored in refrigerated air although not to a level below those displayed in ‘over-mature’ fruit at harvest after 3 months in storage. As harvest maturity advanced, the time required to regenerate aroma volatiles to an ‘optimum’ level after removal from CA storage decreased markedly. A linear relationship between attainment of optimal eating quality and time out of storage was established. Firmness, soluble solids, and titratable acidity of fruit from all harvest dates remained at acceptable levels throughout the post-storage ripening period.

The abundant class III chitinase homolog in young developing banana fruits behaves as a transient vegetative storage protein and most probably serves as an important supply of amino acids for the synthesis of ripening-associated proteins.

Analyses of the protein content and composition revealed dramatic changes in gene expression during in situ banana (Musa spp.) fruit formation/ripening. The total banana protein content rapidly increases during the first 60 to 70 d, but remains constant for the rest of fruit formation/ripening. During the phase of rapid protein accumulation, an inactive homolog of class III chitinases accounts for up to 40% (w/v) of the total protein. Concomitant with the arrest of net protein accumulation, the chitinase-related protein (CRP) progressively decreases and several novel proteins appear in the electropherograms. Hence, CRP behaves as a fruit-specific vegetative storage protein that accumulates during early fruit formation and serves as a source of amino acids for the synthesis of ripening-associated proteins. Analyses of individual proteins revealed that a thaumatin-like protein, a beta-1,3-glucanase, a class I chitinase, and a mannose-binding lectin are the most abundant ripening-associated proteins. Because during the ripening of prematurely harvested bananas, similar changes take place as in the in situ ripening bananas, CRP present in immature fruits is a sufficient source of amino acids for a quasi-normal synthesis of ripening-associated proteins. However, it is evident that the conversion of CRP in ripening-associated proteins takes place at an accelerated rate, especially when climacteric ripening is induced by ethylene. The present report also includes a discussion of the accumulation of the major banana allergens and the identification of suitable promoters for the production of vaccines in transgenic bananas.

Worried about your diet?

Fruit is supposed to be enormously important in terms of our (ill‐defined) general health. Although no‐one knows what five portions are, and the data for specific disease risks are few, the public health message to eat fruit and vegetables is clear—rather like ‘Go to work on an Egg’ or ‘Drink a Pint of Milk a Day’. But this sceptical view does not conceal the fact that ripe fruits are an important part of the human diet. The maturation and ripening of fleshy fruits varies considerably in style among plant species and involves changes in colour, texture, flavour and smell, all of which must be co‐ordinated. These changes are also accompanied by changes in nutritional values. Those accustomed to human crises and the use of words in a particular way must adopt a Tweedeldumic point of view here, for there are two types of ripening: climacteric and non‐climacteric. In climacteric ripening, there is a burst of respiration at the beginning of the process …

Content of bioactive compounds in pungent spice red pepper (paprika) as affected by ripening and genotype

AbstractThis work was conducted to investigate changes in the bioactive nutrient content of hot (pungent) paprika as a function of ripening in four of the most important varieties cultivated in Hungary. For the quantitative determination, recent liquid chromatographic (HPLC) methods were applied. Forty carotenoids were separated, with β‐carotene and lutein of interest from the biological point of view. The major component of vitamin E was α‐tocopherol. Capsaicin, dihydrocapsaicin and nordihydrocapsaicin were the dominant pungency principles. As for vitamin C components, only ascorbic acid could be detected and determined. The results indicated that all the examined compounds were at low level in the mature green fruits, and the onset of climacteric ripening caused their content to grow. In accord with the advance in ripening, carotenoids were being formed even at the overripening stage, while tocopherols, capsaicinoids and ascorbic acid reached their maximum level at the colour break or red stage and then declined. The highest values with regard to carotenoids were estimated in SZ‐178 and F0‐3 cultivars. The best retention of ascorbic acid was found in SZ‐178, which contained 4.86 mg g−1 dry matter in the dry pods. On the other hand, retention of α‐tocopherol during ripening and natural drying was found to depend on variety. The highest value for vitamin E was recorded in SZ‐178 followed by K‐V2, which contained the highest level of α‐tocopherol at the red stage of ripening.© 2001 Society of Chemical Industry

Genetic mapping of ripening and ethylene-related loci in tomato

The regulation of tomato fruit development and ripening is influenced by a large number of loci as demonstrated by the number of existing non-allelic fruit development mutations and a multitude of genes showing ripening-related expression patterns. Furthermore, analysis of transgenic and naturally occurring tomato mutants confirms the pivotal role of the gaseous hormone ethylene in the regulation of climacteric ripening. Here we report RFLP mapping of 32 independent tomato loci corresponding to genes known or hypothesized to influence fruit ripening and/or ethylene response. Mapped ethylene-response sequences fall into the categories of genes involved in either hormone biosynthesis or perception, while additional ripening-related genes include those involved in cell-wall metabolism and pigment biosynthesis. The placement of ripening and ethylene-response loci on the tomato RFLP map will facilitate both the identification and exclusion of candidate gene sequences corresponding to identified single gene and quantitative trait loci contributing to fruit development and ethylene response.

Factors that Influence Biosynthesis of Volatile Flavor Compounds in Apple Fruits

Volatile ester molecules are important contributors to the perception of fruit taste. Biosynthesis of volatile compounds occurs via several biochemical pathways. Ongoing studies have concentrated on alcohol acetyl transferase, the terminal step in the acetate ester synthesis pathway. Our studies on volatile biosynthesis in apples have revealed several interesting phenomena. First, the nature and amount of volatile compounds are cultivarand strain-dependent. Studies with `Delicious' show a relationship between amount of peel coloration and flavor volatile content of tissue. Second, it is possible to manipulate the preharvest growing environment to influence the content of some volatiles in the fruit. Third, generation of volatiles is closely linked to the onset of climacteric ripening. Other experiments show the response of apples to different storage conditions with regard to volatile ester synthesis. In some cultivars softening apparently provides ester precursor molecules, leading us to speculate that there are glycosidically bound intermediates that are liberated by the action of cell-wall degradation.

142 ESTER FORMATION IN RELATION TO HARVEST MATURITY AND CONTROLLED-ATMOSPHERE STORAGE OF `GALA' APPLES

Volatile esters from acids and alcohols are important components of flavor and odor perception in apples (Malus domestica Borkh.). We are interested in understanding the biochemical basis for ester synthesis/flavor retention in `Gala' apples held in controlled atmosphere storage. The relationship between acetyl CoA alcohol transferase (AAT) acetate ester-formin activity, non-ethylene volatile emission, and flesh volatile content of `Gala' apples during the maturation period and after removal from CA storage was investigated. At the appropriate times, apples were sampled for volatile compounds in the headspace and flesh using solid sorbent along with purge-and-trap capillary gas chromatography. Subsequently, acetate ester forming activity was assayed on partially-purified extracts of cortical tissue. During storage, the accumulation of the major flavor notes butyl acetate and 2-methyl butyl acetate in the flesh was decreased as oxygen levels in storage atmospheres were lowered. AAT activity is closely linked to the onset of climacteric ripening and is sensitive to atmospheres having low oxygen contents.