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Current Understanding of Ethylene and Fruit Ripening
Abstract
In fleshy fruits such as bananas, apples, peaches, strawberries, melons, squash, and tomatoes, the process of ripening is a synchronized array of developmental and biochemical events that induce changes in color, aroma, and nutritional quality. Ripening and senescence are quite similar, but according to the metabolic activities, ripening is a different phase for fleshy fruit that precedes and may predispose the fruit to senescence. Ethylene is a gaseous phytohormone that controls fruit ripening and plant growth. Extensive experiments on fruit ripening, induced by ethylene through its process of perception, signaling, and gene control, have studied the function of ethylene in fruit ripening, which has been observed in fruit crops similar to that in Arabidopsis . The analysis of fruit ripening–deficient mutant tomatoes has provided an essential breakthrough in discovering signaling components and transcription factors in ethylene involvement in ripening. This chapter discusses the details of fruit maturation and its control by the synthesis, signaling, and response of ethylene.
... ET is an essential phytohormone implicated in several physiological routes, with seed germination, fruit ripening, senescence, and responses to biotic and abiotic stresses Sharma et al. 2019a). Ripening is a coordinated array of developmental and physiological activities that create changes in color, fragrance, and nutritional content in fleshy fruits such as bananas, apples, peaches, strawberries, melons, squash, and tomatoes Gambhir et al. 2023;Gupta et al. 2022;Liu et al. 2023;Verde et al. 2023;Wei et al. 2022). Ripening and senescence are quite similar; however, ripening is a separate phase for fleshy fruit that precedes and may predispose the fruit to senescence, according to metabolic activity. ...
... Ethylene is a gaseous phytohormone that regulates plant development and fruit ripening. Extensive research on fruit ripening triggered by ethylene through its sensing, signaling, and gene regulation processes has investigated the role of ethylene in fruit ripening, which has been observed in fruit crops (Gupta et al. 2022). ...
... Melatonin has the ability to affect ET biosynthesis and signaling pathways (Verde et al. 2023). ET is produced from methionine via enzymatic processes, with 1-aminocyclopropane-1-carboxylic acid (ACC) serving as an intermediary (Fatma et al. 2022;Gupta et al. 2022). Melatonin has been demonstrated to influence the activity of the major enzymes in ET production, ACC synthase (ACS) and ACC oxidase (ACO) ). ...
Melatonin was found in plants in the late 1990s, but its function, signaling, and interaction with other phytohormones still unclear. Melatonin research in plants has increased substantially in recent years, including reports on the impact of this putative plant hormone under biotic and abiotic stress situations. Temperature extremes, salt, drought, hypoxia or anoxia, nutrient deficiency, herbicides, UV radiation stress, and heavy metal toxicity are all important obstacles to horticulture crop production worldwide. To deal with these environmental challenges, plants have evolved complex signaling networks. Phy-tohormones are essential for controlling plant growth, development, and stress responses. Melatonin, a pleiotropic chemical present in a variety of species, has recently emerged as a powerful regulator of plant abiotic stress tolerance. The purpose of this review is to investigate the interplay between melatonin and phytohormones in the control of abiotic stress responses in horticultural crops. We explore the interactions of melatonin with several phytohormones under various abiotic stresses.
Abscisic acid (ABA) is a plant growth regulator known for its functions, especially in seed maturation, seed dormancy, adaptive responses to biotic and abiotic stresses, and leaf and bud abscission. ABA activity is governed by multiple regulatory pathways that control ABA biosynthesis, signal transduction, and transport. The transport of the ABA signaling molecule occurs from the shoot (site of synthesis) to the fruit (site of action), where ABA receptors decode information as fruit maturation begins and is significantly promoted. The maximum amount of ABA is exported by the phloem from developing fruits during seed formation and initiation of fruit expansion. In the later stages of fruit ripening, ABA export from the phloem decreases significantly, leading to an accumulation of ABA in ripening fruit. Fruit growth, ripening, and senescence are under the control of ABA, and the mechanisms governing these processes are still unfolding. During the fruit ripening phase, interactions between ABA and ethylene are found in both climacteric and non-climacteric fruits. It is clear that ABA regulates ethylene biosynthesis and signaling during fruit ripening, but the molecular mechanism controlling the interaction between ABA and ethylene has not yet been discovered. The effects of ABA and ethylene on fruit ripening are synergistic, and the interaction of ABA with other plant hormones is an essential determinant of fruit growth and ripening. Reaction and biosynthetic mechanisms, signal transduction, and recognition of ABA receptors in fruits need to be elucidated by a more thorough study to understand the role of ABA in fruit ripening. Genetic modifications of ABA signaling can be used in commercial applications to increase fruit yield and quality. This review discusses the mechanism of ABA biosynthesis, its translocation, and signaling pathways, as well as the recent findings on ABA function in fruit development and ripening.
Climacteric fruits are characterized by a dramatic increase in autocatalytic ethylene production that is accompanied by a spike in respiration at the onset of ripening. The change in the mode of ethylene production from autoinhibitory to autostimulatory is known as the System 1 (S1) to System 2 (S2) transition. Existing physiological models explain the basic and overarching genetic, hormonal, and transcriptional regulatory mechanisms governing the S1 to S2 transition of climacteric fruit. However, the links between ethylene and respiration, the two main factors that characterize the respiratory climacteric, have not been examined in detail at the molecular level. Results of recent studies indicate that the alternative oxidase (AOX) respiratory pathway may play an essential role in mediating cross-talk between ethylene response, carbon metabolism, ATP production, and ROS signaling during climacteric ripening. New genomic, metabolic, and epigenetic information sheds light on the interconnectedness of ripening metabolic pathways, necessitating an expansion of the current, ethylene-centric physiological models. Understanding points at which ripening responses can be manipulated may reveal key, species- and cultivar-specific targets for regulation of ripening, enabling superior strategies for reducing postharvest wastage.
Ethylene plays a critical regulatory role in climacteric fruit ripening, and its biosynthesis is fine-tuned at the transcriptional and post-translational levels. Nevertheless, the mechanistic link between transcriptional and post-translational regulation of ethylene biosynthesis during fruit ripening is largely unknown. The present study uncovers a coordinated transcriptional and post-translational mechanism of controlling ethylene biosynthesis during banana (Musa acuminata) fruit ripening. NAC (NAM, ATAF and CUC) proteins MaNAC1 and MaNAC2 repress the expression of MaERF11, a protein previously known to negatively regulate ethylene biosynthesis genes MaACS1 and MaACO1. A RING E3 ligase MaXB3 interacts with MaNAC2 to promote its ubiquitination and degradation, leading to the inhibition of MaNAC2-mediated transcriptional repression. In addition, MaXB3 also targets MaACS1 and MaACO1 for proteasome degradation. Further evidence supporting the role of MaXB3 is provided by its transient and ectopic overexpression in banana fruit and tomato (Solanum lycopersicum), respectively, which delays fruit ripening via repressing ethylene biosynthesis and thus ethylene response. Strikingly, MaNAC1 and MaNAC2 directly repress MaXB3 expression, suggesting a feedback regulatory mechanism that maintains a balance of MaNAC2, MaACS1 and MaACO1 levels. Collectively, our findings establish a multilayered regulatory cascade involving MaXB3, MaNACs, MaERF11 and MaACS1/MaACO1 that controls ethylene biosynthesis during climacteric ripening.
DNA methylation is an important epigenetic mark involved in many biological processes. The genome of the climacteric tomato fruit undergoes a global loss of DNA methylation due to active DNA demethylation during the ripening process. It is unclear whether the ripening of other fruits is also associated with global DNA demethylation. We characterized the single-base resolution DNA methylomes of sweet orange fruits. Compared with immature orange fruits, ripe orange fruits gained DNA methylation at over 30,000 genomic regions and lost DNA methylation at about 1,000 genomic regions, suggesting a global increase in DNA methylation during orange fruit ripening. This increase in DNA methylation was correlated with decreased expression of DNA demethylase genes. The application of a DNA methylation inhibitor interfered with ripening, indicating that the DNA hypermethylation is critical for the proper ripening of orange fruits. We found that ripening-associated DNA hypermethylation was associated with the repression of several hundred genes, such as photosynthesis genes, and with the activation of hundreds of genes, including genes involved in abscisic acid responses. Our results suggest important roles of DNA methylation in orange fruit ripening.
The plant hormone ethylene plays a key role in climacteric fruit ripening. Studies on components of ethylene signaling have revealed a linear transduction pathway leading to the activation of ethylene response factors. However, the means by which ethylene selects the ripening-related genes and interacts with other signaling pathways to regulate the ripening process are still to be elucidated. Using tomato (Solanum lycopersicum) as a reference species, the present review aims to revisit the mechanisms by which ethylene regulates fruit ripening by taking advantage of new tools available to perform in silico studies at the genome-wide scale, leading to a global view on the expression pattern of ethylene biosynthesis and response genes throughout ripening. Overall, it provides new insights on the transcriptional network by which this hormone coordinates the ripening process and emphasizes the interplay between ethylene and ripening-associated developmental factors and the link between epigenetic regulation and ethylene during fruit ripening.
Naturally-occurring epimutants are rare and have mainly been described in plants. However how these mutants maintain their epigenetic marks and how they are inherited remain unknown. Here we report that CHROMOMETHYLASE3 (SlCMT3) and other methyltransferases are required for maintenance of a spontaneous epimutation and its cognate Colourless non-ripening (Cnr) phenotype in tomato. We screened a series of DNA methylation-related genes that could rescue the hypermethylated Cnr mutant. Silencing of the developmentally-regulated SlCMT3 gene results in increased expression of LeSPL-CNR, the gene encodes the SBP-box transcription factor residing at the Cnr locus and triggers Cnr fruits to ripen normally. Expression of other key ripening-genes was also up-regulated. Targeted and whole-genome bisulfite sequencing showed that the induced ripening of Cnr fruits is associated with reduction of methylation at CHG sites in a 286-bp region of the LeSPL-CNR promoter, and a decrease of DNA methylation in differentially-methylated regions associated with the LeMADS-RIN binding sites. Our results indicate that there is likely a concerted effect of different methyltransferases at the Cnr locus and the plant-specific SlCMT3 is essential for sustaining Cnr epi-allele. Maintenance of DNA methylation dynamics is critical for the somatic stability of Cnr epimutation and for the inheritance of tomato non-ripening phenotype.
Fleshy fruit undergo a novel developmental program that ends in the irreversible process of ripening and eventual tissue senescence. During this maturation process, fruit undergo numerous physiological, biochemical and structural alterations, making them more attractive to seed dispersal organisms. In addition, advanced or over-ripening and senescence, especially through tissue softening and eventual decay, render fruit susceptible to invasion by opportunistic pathogens. While ripening and senescence are often used interchangeably, the specific metabolic activities of each would suggest that ripening is a distinct process of fleshy fruits that precedes and may predispose the fruit to subsequent senescence.
Ripening of tomato fruits is triggered by the plant hormone ethylene, but its effect is restricted by an unknown developmental cue to mature fruits containing viable seeds. To determine whether this cue involves epigenetic remodeling, we expose tomatoes to the methyltransferase inhibitor 5-azacytidine and find that they ripen prematurely. We performed whole-genome bisulfite sequencing on fruit in four stages of development, from immature to ripe. We identified 52,095 differentially methylated regions (representing 1% of the genome) in the 90% of the genome covered by our analysis. Furthermore, binding sites for RIN, one of the main ripening transcription factors, are frequently localized in the demethylated regions of the promoters of numerous ripening genes, and binding occurs in concert with demethylation. Our data show that the epigenome is not static during development and may have been selected to ensure the fidelity of developmental processes such as ripening. Crop-improvement strategies could benefit by taking into account not only DNA sequence variation among plant lines, but also the information encoded in the epigenome.
Fruit development is a complex yet tightly regulated process. The developing fruit undergoes phases of cell division and expansion
followed by numerous metabolic changes leading to ripening. Plant hormones are known to affect many aspects of fruit growth
and development. In addition to the five classic hormones (auxins, gibberellins, cytokinins, abscisic acid and ethylene) a
few other growth regulators that play roles in fruit development are now gaining recognition. Exogenous application of various
hormones to different stages of developing fruits and endogenous quantifications have highlighted their importance during
fruit development. Information acquired through biochemical, genetic and molecular studies is now beginning to reveal the
possible mode of hormonal regulation of fruit development at molecular levels. In the present article, we have reviewed studies
revealing hormonal control of fruit development using tomato as a model system with emphasis on molecular genetics.
Important traits for complete ripening and consumer fruit quality preferences include development of aroma, flavor, color, texture, and nutritional quality. These attributes are influenced by the endogenously produced hormone ethylene in many fleshy fruits such as apple, avocado, banana, mango, pear and tomato. Even in species where endogenous ethylene seems to play little if any role as an endogenous regulator, exogenous ethylene will often promote ripening characteristics and can be the target of post-harvest strategies designed to accelerate, synchronize or delay ripening. In recent decades the YANG cycle for ethylene biosynthesis has been revealed and characterized at the molecular level with much of this important work done via the analysis of fruit systems. However, the genetic regulation that controls ethylene production at different developmental stages of fruits has only recently begun to be studied. Tomato has emerged as the primary model plant to further understand the molecular biology that controls ethylene synthesis and additional ripening regulators during fruit development. Here we summarize data pertaining to ethylene biology specifically as related to fruit maturation and including recent insights into genetic control of the ripening process prior to and controlling ethylene.
Tomato ripening is a highly coordinated developmental process that coincides with seed maturation. Regulated expression of thousands of genes controls fruit softening as well as accumulation of pigments, sugars, acids, and volatile compounds that increase attraction to animals. A combination of molecular tools and ripening-affected mutants has permitted researchers to establish a framework for the control of ripening. Tomato is a climacteric fruit, with an absolute requirement for the phytohormone ethylene to ripen. This dependence upon ethylene has established tomato fruit ripening as a model system for study of regulation of its synthesis and perception. In addition, several important ripening mutants, including rin, nor, and Cnr, have provided novel insights into the control of ripening processes. Here, we describe how ethylene and the transcription factors associated with the ripening process fit together into a network controlling ripening.
To investigate the regulatory mechanism(s) of ethylene biosynthesis in fruit, transgenic tomatoes with all known LeEIL genes suppressed were produced by RNA interference engineering. The transgenic tomato exhibited ethylene insensitivity phenotypes
such as non-ripening and the lack of the triple response and petiole epinasty of seedlings even in the presence of exogenous
ethylene. Transgenic fruit exhibited a low but consistent increase in ethylene production beyond 40 days after anthesis (DAA),
with limited LeACS2 and LeACS4 expression. 1-Methylcyclopropene (1-MCP), a potent inhibitor of ethylene perception, failed to inhibit the limited increase
in ethylene production and expression of the two 1-aminocyclopropane-1-carboxylic acid (ACC) synthase (ACS) genes in the transgenic fruit. These results suggest that ripening-associated ethylene (system 2) in wild-type tomato fruit
consists of two parts: a small part regulated by a developmental factor through the ethylene-independent expression of LeACS2 and LeACS4 and a large part regulated by an autocatalytic system due to the ethylene-dependent expression of the same genes. The results
further suggest that basal ethylene (system 1) is less likely to be involved in the transition to system 2. Even if the effect
of system 1 ethylene is eliminated, fruit can show a small increase in ethylene production due to unknown developmental factors.
This increase would be enough for the stimulation of autocatalytic ethylene production, leading to fruit ripening.
Ethylene regulates many aspects of the plant life cycle, including seed germination, root initiation, flower development, fruit ripening, senescence, and responses to biotic and abiotic stresses. It thus plays a key role in responses to the environment that have a direct bearing on a plant's fitness for adaptation and reproduction. In recent years, there have been major advances in our understanding of the molecular mechanisms regulating ethylene synthesis and action. Screening for mutants of the triple response phenotype of etiolated Arabidopsis seedlings, together with map-based cloning and candidate gene characterization of natural mutants from other plant species, has led to the identification of many new genes for ethylene biosynthesis, signal transduction, and response pathways. The simple chemical nature of ethylene contrasts with its regulatory complexity. This is illustrated by the multiplicity of genes encoding the key ethylene biosynthesis enzymes 1-aminocyclopropane-1-carboxylic acid (ACC) synthase and ACC oxidase, multiple ethylene receptors and signal transduction components, and the complexity of regulatory steps involving signalling relays and control of mRNA and protein synthesis and turnover. In addition, there are extensive interactions with other hormones. This review integrates knowledge from the model plant Arabidopsis and other plant species and focuses on key aspects of recent research on regulatory networks controlling ethylene synthesis and its role in flower development and fruit ripening.
The gaseous hormone ethylene is perceived by a family of ethylene receptors which interact with the Raf-like kinase CTR1. SlTPR1 encodes a novel TPR (tetratricopeptide repeat) protein from tomato that interacts with the ethylene receptors NR and LeETR1 in yeast two-hybrid and in vitro protein interaction assays. SlTPR1 protein with a GFP fluorescent tag was localized in the plasmalemma and nuclear membrane in Arabidopsis, and SlTPR1-CFP and NR-YFP fusion proteins were co-localized in the plasmalemma and nuclear membrane following co-bombardment of onion cells. Overexpression of SlTPR1 in tomato resulted in ethylene-related pleiotropic effects including reduced stature, delayed and reduced production of inflorescences, abnormal and infertile flowers with degenerate styles and pollen, epinasty, reduced apical dominance, inhibition of abscission, altered leaf morphology, and parthenocarpic fruit. Similar phenotypes were seen in Arabidopsis overexpressing SlTPR1. SlTPR1 overexpression did not increase ethylene production but caused enhanced accumulation of mRNA from the ethylene responsive gene ChitB and the auxin-responsive gene SlSAUR1-like, and reduced expression of the auxin early responsive gene LeIAA9, which is known to be inhibited by ethylene and to be associated with parthenocarpy. Cuttings from the SlTPR1-overexpressors produced fewer adventitious roots and were less responsive to indole butyric acid. It is suggested that SlTPR1 overexpression enhances a subset of ethylene and auxin responses by interacting with specific ethylene receptors. SlTPR1 shares features with human TTC1, which interacts with heterotrimeric G-proteins and Ras, and competes with Raf-1 for Ras binding. Models for SlTPR1 action are proposed involving modulation of ethylene signalling or receptor levels.
Considerable progress in tomato molecular biology has been made over the past five years. At least 19 different mRNAs which increase in amount during tomato fruit ripening have been cloned and genes for enzymes involved in cell wall degradation (polygalacturonase and pectinesterase) and ethylene synthesis (ACC synthase) have been identified by conventional procedures. Transgenic plants have been used to identify regions of DNA flanking fruit-specific, ripening-related and ethylene-regulated genes and trans-acting factors which bind to these promoters have also been identified.
Antisense genes expressed in transgenic plants have proved to be highly effective for inhibiting the specific expression of ripening-related genes. These experiments have changed our understanding of how softening occurs in tomato fruit. Antisense techniques have also been used to identify genes encoding enzymes for carotenoid biosynthesis (phytoene synthase) and ethylene biosynthesis (the ethylene-forming enzyme). The altered characteristics of fruit transformed with specific antisense genes, such as retarded ripening and resistance to splitting, may prove to be of value to fruit growers, processors and ultimately the consumer.
Ethylene controls fruit ripening. Expression of antisense RNA to the rate-limiting enzyme in the biosynthetic pathway of ethylene, 1-aminocyclopropane-1-carboxylate synthase, inhibits fruit ripening in tomato plants. Administration of exogenous ethylene or propylene reverses the inhibitory effect. This result demonstrates that ethylene is the trigger and not the by-product of ripening and raises the prospect that the life-span of plant tissues can be extended, thereby preventing spoilage.
Seedlings of tomato fruit ripening mutants were screened for their ability to respond to ethylene. Ethylene induced the triple response in etiolated hypocotyls of all tomato ripening mutants tested except for one, Never ripe (Nr). Our results indicated that the lack of ripening in this mutant is caused by ethylene insensitivity. Segregation analysis indicated that Nr-associated ethylene insensitivity is a single codominant trait and is pleiotropic, blocking senescence and abscission of flowers and the epinastic response of petioles. In normal tomato flowers, petal abscission and senescence occur 4 to 5 days after the flower opens and precede fruit expansion. If fertilization does not occur, pedicel abscission occurs 5 to 8 days after petal senescence. If unfertilized, Nr flowers remained attached to the plant indefinitely, and petals remained viable and turgid more than four times longer than their normal counterparts. Fruit development in Nr plants was not preceded by petal senescence; petals and anthers remained attached until they were physically displaced by the expanding ovary. Analysis of engineered 1-aminocyclopropane-1-carboxylate (ACC) synthase-overexpressing plants indicated that they are phenotypic opposites of Nr plants. Constitutive expression of ACC synthase in tomato plants resulted in high rates of ethylene production by many tissues of the plant and induced petiole epinasty and premature senescence and abscission of flowers, usually before anthesis. There were no obvious effects on senescence in leaves of ACC synthase overexpressers, suggesting that although ethylene may be important, it is not sufficient to cause tomato leaf senescence; other signals are clearly involved.
Transcription of the E4 gene is controlled by an increase in ethylene concentration during tomato fruit ripening. To investigate the molecular basis for ethylene regulation, we have examined the E4 promoter to identify cis elements and trans-acting factors that are involved in E4 gene expression. In transgenic tomato plants a chimeric gene construct containing a 1.4-kilobase E4 promoter fused to a beta-glucuronidase reporter gene is rapidly induced by ethylene in ripening fruit. Deletion of E4 promoter sequences to 193 base pairs reduces the level of GUS activity but does not affect ethylene induction. Transient expression of E4 promoter-luciferase chimeric gene constructs containing various deletions, introduced into tomato fruit pericarp by particle bombardment, indicates that a positive ethylene-responsive region is localized between nucleotides -161 and -85 relative to the transcription start site. DNase I footprint analysis shows that a nuclear factor in unripe fruit interacts specifically with sequences in this element, from -142 to -110, which are required for the ethylene response. The DNase I footprint of this factor is reduced in ethylene-treated unripe fruit and undetectable in ripe fruit. Based on the correlation of a nuclear factor binding site with promoter sequences required for ethylene induction, we propose that this in vitro DNA-binding activity may represent a factor that is involved in ethylene-regulated E4 gene expression.
We isolated a recessive Arabidopsis mutant, ctr1, that constitutively exhibits seedling and adult phenotypes observed in plants treated with the plant hormone ethylene. The ctr1 adult morphology can be phenocopied by treatment of wild-type plants with exogenous ethylene and is due, at least in part, to inhibition of cell elongation. Seedlings and adult ctr1 plants show constitutive expression of ethylene-regulated genes. The epistasis of ctr1 and other ethylene response mutants has defined the position of CTR1 in the ethylene signal transduction pathway. The CTR1 gene has been cloned, and the DNA sequences of four mutant alleles were determined. The gene encodes a putative serine/threonine protein kinase that is most closely related to the Raf protein kinase family.
Expansins are proteins that induce extension in isolated plant cell walls in vitro and have been proposed to disrupt noncovalent interactions between hemicellulose and cellulose microfibrils. Because the plant primary cell wall acts as a constraint to cell enlargement, this process may be integral to plant cell expansion, and studies of expansins have focused on their role in growth. We report the identification of an expansin (LeExp1) from tomato that exhibits high levels of mRNA abundance and is specifically expressed in ripening fruit, a developmental period when growth has ceased but when selective disassembly of cell wall components is pronounced. cDNAs closely related to LeExp1 were also identified in ripening melons and strawberries, suggesting that they are a common feature of fruit undergoing rapid softening. Furthermore, the sequence of LeExp1 and its homologs from other ripening fruit define a subclass of expansin genes. Expression of LeExp1 is regulated by ethylene, a hormone known to coordinate and induce ripening in many species. LeExp1 is differentially expressed in the ripening-impaired tomato mutants Nr, rin, and nor, and mRNA abundance appears to be influenced directly by ethylene and by a developmentally modulated transduction pathway. The identification of a ripening-regulated expansin gene in tomato and other fruit suggests that, in addition to their role in facilitating the expansion of plant cells, expansins may also contribute to cell wall disassembly in nongrowing tissues, possibly by enhancing the accessibility of noncovalently bound polymers to endogenous enzymic action.
This book covers the biochemistry and molecular biology of ripening of the world's major fruit types.
Ethylene plays an essential role in climacteric fruit ripening via the ethylene signaling pathway. Ethylene response factor (ERF) is a critical downstream component of the ethylene signaling pathway. However, the transcriptional regulatory mechanism underlying ERF in persimmon fruit ripening remains poorly understood. Here, we explored the role of DkERF8/16/18 in regulating persimmon fruit ripening. Transmission electron microscopy showed that persimmon fruit softening was associated with middle lamella degradation and cell wall swelling and distortion. The expression of five ERF genes (DkERF8/16/18/19/24), twelve cell-wall-modifying genes (DkPG1, DkPL1, DkPE1/2, Dkβ-GAL1, DkEGase1, DkXTH2/9/10/11, DkMAN1, DkEXP4) and four ethylene biosynthesis genes (DkACS1/2, DkACO1/2) was induced by ethylene and suppressed by 1-MCP during persimmon fruit storage. Dual luciferase assays, site mutations and electrophoretic mobility shift assays indicated that DkERF8 and DkERF16 activate DkXTH11 and DkEXP4, respectively, by binding to their promoters. DkERF18 binds to the DkACS2 promoter, increasing its activity. Transient overexpression of DkERF8 promotes the conversion of acid soluble pectin to water soluble pectin in persimmon fruit. Moreover, transient overexpression of DkERF18 resulted in increased ethylene production. These results suggest that DkERF8/16/18 may be involved in persimmon fruit ripening by promoting cell wall modification and ethylene biosynthesis.
‘Mopan’ persimmon (Diospyros kaki L. cv. Mopan) is an economically important persimmon cultivar that is widely grown in northern China areas. Expanded understanding of postharvest fruit physiological changes and the role that ethylene plays will enhance the efficiency of fruit storage and marketing. Treatments with ethylene and 1-MCP on ‘Mopan’ persimmons showed opposite effects on softening, color, total soluble solid (TSS), total and soluble tannin contents, ethylene production and respiration rate during fruit storage. Ethylene and 1-MCP treatments also affected the expression of five ethylene signaling genes DkCTR1, DkETR1, DkETR2, DkERF22 and DkERF19, three de-astringent related genes DkADH1, DkPDC1 and DkPDC2, and four cell wall-hydrolyzing enzyme genes DkPG1, DkXTH2, DkPME1 and Dkβ-GAL1 during fruit storage. High correlations were observed between ethylene signal pathway genes, physiological characters and ripening related gene expression. The results indicate that ethylene greatly accelerate fruit ripening, and the ripening process can be inhibited by 1-MCP treatment both on physiological and molecular levels. The application of 1-MCP delayed seven to ten days on fruit ripening compared to controls. The overall information obtained from this study demonstrates that ethylene plays the critical roles in postharvest ripening, gene expressions and physiological property changes of ‘Mopan’ persimmon fruit during storage. The suppression of ethylene activity by 1-MCP provides the industry with an applicable technology to improve postharvest processing and ‘Mopan’ persimmon fruit value.
Fruit has evolved myriad forms that facilitate seed dispersal in varied environmental and ecological contexts. Because fleshy fruits become attractive and nutritious to seed-dispersing animals, the transition from unripe to ripe fruit represents a dramatic shift in survival strategy-from protecting unripe fruit against damaging animals to making it appealing to those same animals once ripened. For optimal fitness, ripening therefore must be tightly controlled and coordinated with seed development. Fruits, like many vegetative tissues of plants that contribute to human diets, are also subject to decay, which is enhanced as a consequence of the ripening transition. As such, ripening control has enormous relevance for both plant biology and food security. Here, we review the complex interactions of hormones and transcription factors during fleshy-fruit ripening, with an emphasis on the recent discovery that epigenome dynamics are a critical and early regulator of the cascade of molecular events that ultimately contribute to fruit maturation and ripening. Expected final online publication date for the Annual Review of Plant Biology Volume 68 is April 29, 2017. Please see http://www.annualreviews.org/page/journal/pubdates for revised estimates.
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It is estimated that as many as 100 million people subsist on bananas and plantains as their main energy source (Rowe, 1981). Also the international trade in bananas is of considerable economic importance. The estimated world production of banana and plantain fruits in 1990 was about 71 million tonnes; bananas accounted for 46 million tonnes of which, in 1989, 8.1 million tonnes were exported (FAO, 1989; FAO, 1990).
Ethylene in Plant Biology, Second Edition provides a definitive survey of what is currently known about this structurally simplest of all plant growth regulators. This volume contains all new material plus a bibliographic guide to the complete literature of this field. Progress in molecular biology and biotechnology as well as biochemistry, plant physiology, development, regulation, and environmental aspects is covered in nine chapters co-authored by three eminent authorities in plant ethylene research. This volume is the modern text reference for all researchers and students of ethylene in plant and agricultural science. Key Features * Completely updated * Concise, readable style for students and professional * Contains an extensive bibliographic guide to the original literature * Well illustrated with diagrams and photographs * Thorough coverage of: * ethylene and ethephon roles and effects * stress ethylene * biosynthesis of ethylene * molecular biology of ethylene * action of ethylene * agricultural uses of ethylene.
Plants as immobile organisms need to constantly monitor the changes in the environment to modify and adjust developmental and metabolic pathways accordingly. The responses to these environmental cues require an integrative mechanism where external and internal signals are detected and processed to trigger an appropriate 'reaction' in the plant. Hormones play a key role in mediating some of these integrative processes and in generating the response reactions. The identification and characterization of the basic hormone signalling components and their interactions represent the first step towards comprehensive understanding of plant responses to intrinsic and extrinsic cues. A relatively well-characterized ethylene signalling and response pathway, together with numerous evidences of its interactions with other signalling/response pathways, provide an excellent example to illustrate our current knowledge and perspective on how signal integration occurs in plants.
Plants respond in complex ways to their environment, to their internal physiological status, and to the activity of other plants, pathogens, herbivores, and organisms. Plant Signaling 2000, a symposium sponsored by the Penn State Intercollege Graduate Program in Plant Physiology (May 18–20, 2000), explored the machinery underlying these responses and their potential for cross talk. We recount here some of the major themes emerging from this interdisciplinary symposium, which ranged from genetic and biochemical analyses of signaling pathways in Arabidopsis and other model plants to field studies of plants responding to insect damage.
1-Aminocyclopropane-1-carboxylic acid synthase (ACS) is one of the key regulatory enzymes involved in the synthesis of the hormone ethylene and is encoded by a multigene family containing at least eight members in tomato (Lycopersicon esculentum). Increased ethylene production accompanies ripening in tomato, and this coincides with a change in the regulation of ethylene synthesis from auto-inhibitory to autostimulatory. The signaling pathways that operate to bring about this transition from so-called system-1 to system-2 ethylene production are unknown, and we have begun to address these by investigating the regulation of ACS expression during ripening. Transcripts corresponding to four ACS genes,LEACS1A, LEACS2, LEACS4, and LEACS6, were detected in tomato fruit, and expression analysis using the ripening inhibitor(rin) mutant in combination with ethylene treatments and the Never-ripe (Nr) mutant has demonstrated that each is regulated in a unique way. A proposed model suggests that system-1 ethylene is regulated by the expression ofLEACS1A and LEACS6. In fruit a transition period occurs in which the RIN gene plays a pivotal role leading to increased expression of LEACS1A and induction of LEACS4. System-2 ethylene synthesis is subsequently initiated and maintained by ethylene-dependent induction ofLEACS2.
To investigate the transcriptional regulation of two 1-aminocyclopropane-1-carboxylate (ACC) oxidase (ACO) genes of peach, chimeric fusions between β-glucuronidase (GUS) reporter gene, and Pp-ACO1 and Pp-ACO2 promoters have been constructed and introduced in tomato (cv Microtom). Pp-ACO1 promoter is able to induce in transgenic tomato plants the same pattern of expression observed in peach. In fact, Pp-ACO1–GUS activity was localized in leaf blade, ovary, leaf and fruit abscission zones and pericarp, and it is up-regulated by propylene and wounding. A gradient of transcript accumulation has been observed in ripening fruit tissues: in tomato it decreased moving from the inner to the outer pericarp, while in peach the opposite occurred. This different behavior could be related to the fruit type (berry vs. drupe). In order to identify cis-acting element involved in ethylene induction of Pp-ACO1, portions of its promoter fused with the GUS gene have been constructed and used in peach fruit transient activity assay. The deletion analysis has shown that a region located between −716 and −346 bp, containing an ethylene-responsive element (ERE), is responsible for the higher stimulation by the gas. In addition, two auxin-responsive elements (AUXre), probably responsible for the auxin suppression of the propylene induction of Pp-ACO1 gene expression, are present upstream from EREs. Pp-ACO2 promoter is able to drive the expression in vascular bundles of immature and ripe fruit, senescent leaf blade, and in fruit and leaf abscission zones. In peach, Pp-ACO2 mRNA is detected only in immature fruit, epicotyl and root of seedling. This discrepancy might be imputed to a lesser stability and translation of Pp-ACO2 mRNA in comparison to that of chimeric one.
Plants as immobile organisms need to constantly monitor the changes in the environment to modify and adjust developmental and metabolic pathways accordingly. The responses to these environmental cues require an integrative mechanism where external and internal signals are detected and processed to trigger an appropriate ‘reaction’ in the plant. Hormones play a key role in mediating some of these integrative processes and in generating the response reactions. The identification and characterization of the basic hormone signalling components and their interactions represent the first step towards comprehensive understanding of plant responses to intrinsic and extrinsic cues. A relatively well-characterized ethylene signalling and response pathway, together with numerous evidences of its interactions with other signalling/response pathways, provide an excellent example to illustrate our current knowledge and perspective on how signal integration occurs in plants.
Loss-of-function ethylene insensitive 2 (EIN2) mutations showed ethylene insensitivity in Arabidopsis, which indicated an essential role of EIN2 in ethylene signaling. However, the function of EIN2 in fruit ripening has not been investigated. To gain a better understanding of EIN2, the temporal regulation of LeEIN2 expression during tomato fruit development was analyzed. The expression of LeEIN2 was constant at different stages of fruit development, and was not regulated by ethylene. Moreover, LeEIN2-silenced tomato fruits were developed using a virus-induced gene silencing fruit system to study the role of LeEIN2 in tomato fruit ripening. Silenced fruits had a delay in fruit development and ripening, related to greatly descended expression of ethylene-related and ripening-related genes in comparison with those of control fruits. These results suggested LeEIN2 positively mediated ethylene signals during tomato development. In addition, there were fewer seeds and locules in the silenced fruit than those in the control fruit, like the phenotype of parthenocarpic tomato fruit. The content of auxin and the expression of auxin-regulated gene were declined in silenced fruit, which indicated that EIN2 might be important for crosstalk between ethylene and auxin hormones.
(Managing editor: Li-Hui Zhao)
To study the significance of stress-inducible ethylene in the hypersensitive reaction of tobacco (Nicotiana tabacum) to tobacco mosaic virus (TMV), six types of transgenic plants were generated containing sense or antisense constructs corresponding to the ethylene biosynthesis enzymes ACC-synthase (ACS) and/or ACC-oxidase under the control of the CaMV 35S promoter. In most primary transformants, expression of antisense RNA or incomplete sense RNA resulted in a reduction of inducible ethylene production, while plants overexpressing full-length sense ACS mRNA had higher ethylene levels than control plants. Although efficient overexpression or gene silencing was evident at the transcript level, there was no correlation with the amounts of ethylene produced, supporting the notion that ACS is post-transcriptionally regulated. Transgenic tobacco plants with altered ethylene levels were not impaired in their ability to respond with necrotic local lesions to infection with TMV. However, in the ACS sense and antisense transformants, alterations in ethylene levels did affect stem length and leaf chlorophyll levels in accordance with established ethylene responses.
The ethylene antagonists, 2,5-norbornadiene (NBD) and silver nitrate, were used to probe the involvement of endogenous ethylene in the natural degreening of citrus fruit. Mature-green, detached Shamouti orange (Citrus sinensis L. Osbeck) fruit were treated with NBD vapor or dipped in solutions of silver nitrate. More than 80% of the chlorophyll was lost from control fruit after 8 days. NBD (0.11 mmole/liter) inhibited the loss of chlorophyll by 60%. NBD also antagonized the degreening induced by exogenous ethylene by 50%. Silver nitrate (0.1 mM) inhibited the loss of chlorophyll by 55%. Ethylene evolution of mature, green detached fruit was -1.h-1 (ca. 13.5 nl.Kg-1FW.h-1) and did not change significantly for 7 days after harvest. NBD concentrations up to 0.22 mmole/liter did not enhance ethylene evolution. Not with-standing the extremely low amounts of ethylene evolved, the inhibition of degreening by NBD and silver nitrate suggests that endogenous ethylene is involved in the control of this process in mature citrus fruit.
Expansins are known to participate in several processes during plant growth and development particularly where wall extension and cell expansion are required. These are also believed to prepare cell wall for their subsequent degradation by cell wall hydrolases during ripening particularly in climacteric fruits. The identification and characterization of a fruit-specific expansin, MaExp1, gene is reported here for the first time from banana. The MaExp1 cDNA of 1098 bp encodes a polypeptide of 255 amino acids, which has all the characteristics of an α-expansin. Ethylene exposure to unripe mature banana fruit induces MaExp1 expression, which increases with the progression in ripening and 1-MCP (1-methyl cyclopropene) treatment prior to ethylene exposure inhibits expression. No expression has been detected in any other tissue. The genomic sequence analysis has revealed that MaExp1 contains two short introns of 83 and 79 nucleotides. The 888 bp proximal promoter of the gene shows presence of a putative ethylene and auxin responsive elements and a direct repeat with in the sequence. The IAA treatment enhanced MaExp1 expression only in the presence of ethylene suggesting its effect as synergistic and additive. It is suggested that MaExp1 could be used for manipulating ripening in banana and its promoter could be a suitable candidate for expressing foreign gene (vaccines) in transgenic banana fruit.
While the grape has been classified as a non-climacteric fruit whose ripening is thought to be ethylene independent, we show here that a transient increase of endogenous ethylene production occurs just before veraison (i.e. inception of ripening). We observed that ethylene perception, at this time, is required for at least the increase of berry diameter, the decrease of berry acidity and anthocyanin accumulation in the ripening berries; these latter experiments were performed with 1-methylcyclopropene, a specific inhibitor of ethylene receptors. The potential roles of ethylene in berry development and ripening are discussed.
Ripening in climacteric fruit is triggered by the action of ethylene and results in activation of several cell wall hydrolases. Their action on cell walls results in wall disassembly leading to softening. One of the exotic varieties of mango, ‘Dashehari’ (Mangifera indica cv. Dashehari), grown mainly in Northern India, suffers from rapid and uneven ripening making it unfit for export. Several biochemical and physiological studies have been performed to understand the process of ripening in this mango. However, there have so far been no substantial data on the molecular analysis of genes related to softening, in ‘Dashehari’, and other varieties of mango in general. We report here isolation and characterization of an α-expansin gene, MiExpA1 that is correlated with softening in mango. The expression of this gene is under dual control, being triggered by ethylene treatment within 90 min followed by a ripening associated peak in transcript accumulation on the third day after ethylene treatment. At the protein level, expression of the expansin is detectable from the second day itself and continues throughout the course of softening. Treatment with 1-MCP inhibits both ripening/softening as well as MiExpA1 transcript and protein accumulation. It is suggested that MiExpA1 expression is ethylene dependent and its expression increases with the progression of ripening. This gene could be a good candidate for manipulating ripening in mango.
The ripening-impaired tomato mutant Never-ripe (Nr) is insensitive to the plant hormone ethylene. The gene that cosegregates with the Nr locus encodes a protein with homology to the Arabidopsis ethylene receptor ETR1 but is lacking the response regulator domain found in ETR1 and related prokaryotic two-component signal transducers. A single amino acid change in the sensor domain confers ethylene insensitivity when expressed in transgenic tomato plants. Modulation of NR gene expression during fruit ripening controls response to the hormone ethylene.
The transition of fleshy fruit maturation to ripening is regulated by exogenous and endogenous signals that coordinate the transition of the fruit to a final state of attractiveness to seed dispersing organisms. Tomato is a model for biology and genetics regulating specific ripening pathways including ethylene, carotenoids and cell wall metabolism in addition to upstream signaling and transcriptional regulators. Ripening-associated transcription factors described to date including the RIN-MADS, CLEAR NON-RIPENING, TAGL1 and LeHB-1 genes all encode positive regulators of ripening phenomena. Here we describe an APETALA2 transcription factor (SlAP2a) identified through transcriptional profiling of fruit maturation that is induced during, and which negatively regulates, tomato fruit ripening. RNAi repression of SlAP2a results in fruits that over-produce ethylene, ripen early and modify carotenoid accumulation profiles by altering carotenoid pathway flux. These results suggest that SlAP2a functions during normal tomato fruit ripening as a modulator of ripening activity and acts to balance the activities of positive ripening regulators.
Development and ripening in fruit is a unique phase in the life cycle of higher plants which encompasses several stages progressively such as fruit development, its maturation, ripening and finally senescence. During ripening phase, several physiological and biochemical changes take place through differential expression of various genes that are developmentally regulated. Expression and/or suppression of these genes contribute to various changes in the fruit that make it visually attractive and edible. However, in fleshy fruit massive losses accrue during post harvest handling of the fruit which may run into billions of dollars worldwide. This encouraged scientists to look for various ways to save these losses. Genetic engineering appears to be the most promising and cost effective means to prevent these losses. Most fleshy fruit ripen in the presence of ethylene and once ripening has been initiated proceeds uncontrollably. Ethylene evokes several responses during ripening through a signaling cascade and thousands of genes participate which not only sets in ripening but also responsible for its spoilage. Slowing down post ripening process in fleshy fruit has been the major focus of ripening-related research. In this review article, various developments that have taken place in the last decade with respect to identifying and altering the function of ripening-related genes have been described. Role of ethylene and ethylene-responsive genes in ripening of fleshy fruit is also included. Taking clues from the studies in tomato as a model fruit, few case studies are reviewed.
The gaseous plant hormone ethylene acts as a pivotal mediator to respond to and coordinate internal and external cues in modulating plant growth dynamics and developmental programs. Genetic analysis of Arabidopsis thaliana has been used to identify key components and to build a linear ethylene-signaling pathway from the receptors through to the nuclear transcription factors. Studies applying integrative approaches have revealed new regulators, molecular connections and mechanisms in ethylene signaling and unexpected links to other plant hormones. Here, we review and discuss recent discoveries about the functional mode of ethylene receptor complexes, dual mitogen-activated protein kinase cascade signaling, stability control of the master nuclear transcription activator ETHYLENE INSENSITIVE 3 (EIN3), and the contextual relationships between ethylene and other plant hormones, such as auxin and gibberellins, in organ-specific growth regulation.
FLESHY fruits have been divided into two classes on the basis of their
respiratory behaviour during ripening: climacteric fruit, such as
bananas, which undergo a large increase in respiration (climacteric
rise) accompanied by marked changes in composition and texture, and
non-climacteric fruit such as citrus, which show no changes in
respiration that can be associated with distinct changes in the
composition of the fruit1. An increase in the level of
endogenous ethylene is considered to be the immediate trigger of
ripening in climacteric fruits2. Fruits of this class usually
produce large amounts of ethylene once ripening is under way. They may
also be induced to ripen by treatment with ethylene at concentrations
above about 0.1 p.p.m. for a suitable period3. The ripening
induced by exogenous ethylene has been considered to be qualitatively
identical with that which occurs naturally3. In both cases,
once ripening is induced it has been considered that endogenous ethylene
production rises autocatalytically4. Uninjured citrus fruit
have been shown to produce low amounts of ethylene5. Their
respiration may be increased by treatment with ethylene6 and
disappearance of chlorophyll (colouring) and ageing may be more
rapid18.
Twelve cDNAs corresponding to mRNAs inducible by ethylene were isolated by differential screening of a cDNA library from ethylene-treated Citrus sinensis fruits. Northern analysis of RNA extracted from flavedo of ethylene-treated fruits and from fruits at different maturation stages showed that some of the mRNAs corresponding to these cDNAs were regulated both by ethylene treatment and during fruit maturation. The effect of exogenous ethylene on leaves and of endogenous ethylene on flowers showed that gene induction was not restricted to the flavedo tissue. The possible role of ethylene during maturation of the non-climacteric Citrus fruit is discussed.
The response of Arabidopsis thaliana etiolated seedlings to the plant hormone ethylene is a conspicuous phenotype known as the triple response. We have identified genes that are required for ethylene perception and responses by isolating mutants that fail to display a triple response in the presence of exogenous ethylene. Five new complementation groups have been identified. Four of these loci, designated ein4, ein5, ein6 and ein7, are insensitive to ethylene. The fifth complementation group, eir1, is defined by a novel class of mutants that have agravitropic and ethylene-insensitive roots. Double-mutant phenotypes have allowed the positioning of these loci in a genetic pathway for ethylene signal transduction. The ethylene-response pathway is defined by the following loci: ETR1, EIN4, CTR1, EIN2, EIN3, EIN5, EIN6, EIN7, EIR1, AUX1 and HLS1. ctr1-1 is epistatic to etr1-3 and ein4, indicating that CTR1 acts after both ETR1 and EIN4 in the ethylene-response pathway. Mutations at the EIN2, EIN3, EIN5, EIN6 and EIN7 loci are all epistatic to the ctr1 seedling phenotype. The EIR1 and AUX1 loci define a root-specific ethylene response that does not require EIN3 or EIN5 gene activity. HLS1 appears to be required for differential cell growth in the apical hook. The EIR1, AUX1 and HLS1 genes may function in the interactions between ethylene and other plant hormones that occur late in the signaling pathway of this simple gas.