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Ethylene and Fruit Ripening

January 1997
Journal of Plant Growth Regulation 26(2)

Authors:

United States Department of Agriculture

The ripening of fleshy fruits represents the unique coordination of developmental and biochemical pathways leading to changes in color, texture, aroma, and nutritional quality of mature seed-bearing plant organs. The gaseous plant hormone ethylene plays a key regulatory role in ripening of many fruits, including some representing important contributors of nutrition and fiber to the diets of humans. Examples include banana, apple, pear, most stone fruits, melons, squash, and tomato. Molecular exploration of the role of ethylene in fruit ripening has led to the affirmation that mechanisms of ethylene perception and response defined in the model system Arabidopsis thaliana are largely conserved in fruit crop species, although sometimes with modifications in gene family size and regulation. Positional cloning of genes defined by ripening defect mutations in the model fruit system tomato have recently led to the identification of both novel components of ethylene signal transduction and unique transcription factor functions influencing ripening-related ethylene production. Here we summarize recent developments in the regulation of fruit ripening with an emphasis on the regulation of ethylene synthesis, perception, and response.

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Ethylene and Fruit Ripening

Cornelius S. Barry,

and James J. Giovannoni

1,2

Boyce Thompson Institute for Plant Research, Ithaca, New York 14853, USA;

United States Department of Agriculture – Agriculture

Research Service (USDA-ARS), Plant, Soil and Nutrition Laboratory, Cornell University Campus, Tower Road, Ithaca,

New York 14853, USA

BSTRACT

The ripening of ﬂeshy fruits represents the unique

coordination of developmental and biochemical

pathways leading to changes in color, texture, ar-

oma, and nutritional quality of mature seed-bearing

plant organs. The gaseous plant hormone ethylene

plays a key regulatory role in ripening of many

fruits, including some representing important con-

tributors of nutrition and ﬁber to the diets of hu-

mans. Examples include banana, apple, pear, most

stone fruits, melons, squash, and tomato. Molecular

exploration of the role of ethylene in fruit ripening

has led to the afﬁrmation that mechanisms of eth-

ylene perception and response deﬁned in the model

system Arabidopsis thaliana are largely conserved in

fruit crop species, although sometimes with modi-

ﬁcations in gene family size and regulation. Posi-

tional cloning of genes deﬁned by ripening defect

mutations in the model fruit system tomato have

recently led to the identiﬁcation of both novel

components of ethylene signal transduction and

unique transcription factor functions inﬂuencing

ripening-related ethylene production. Here we

summarize recent developments in the regulation of

fruit ripening with an emphasis on the regulation of

ethylene synthesis, perception, and response.

Key words: Ethylene; Fruit; Tomato; Ripening;

Climacteric; Signal transduction

NTRODUCTION

Fruits of different plant species are highly diverse,

ranging from dry seed capsules that burst to allow

seed dispersal, to relatively large complex ﬂeshy fruits

that have evolved bright colors and complex aromas

to attract seed-dispersing birds and animals. Fleshy

fruits in themselves are botanically diverse with some

such as tomato and grape being true berries derived

from the ovary and others such as strawberry, pine-

apple, and apple derived from the receptacle tissues

or from expansion of the sepals. Fleshy fruits also

come in a wide range of sizes, shapes, and colors, and

each species possesses its own very unique ﬂavor

characteristics. Ripening programs can also be di-

verse. For example, avocado do not ripen until after

harvest, whereas the majority of studied fruits ripen

on the plant. Despite this great diversity, aspects of

the ripening of ﬂeshy fruits are often conserved be-

tween species. For example, the onset of ripening is

often associated with color changes, altered sugar

metabolism, fruit softening and alterations in tex-

ture, the synthesis of aroma volatiles, and an in-

creased susceptibility to pathogen infection. These

common events suggest that the underlying genetic

mechanisms that regulate fruit ripening may well be

conserved between fruits of different species (Adams-

Phillips and others 2004a, b; Giovannoni 2004).

Received: 16 January 2007; accepted: 18 January 2007; Online publica-

tion: 6 June 2007

*Corresponding author; e-mail: jjg33@cornell.edu

J Plant Growth Regul (2007) 26:143–159

DOI: 10.1007/s00344-007-9002-y

143

Tomato is the most genetically tractable plant

system for studying fruit ripening because it has

simple diploid genetics and a relatively short gener-

ation time and small habit compared to many other

fruit crop species that are either polyploids or trees.

The ripening phenotype is easy to score and there is a

large collection of germplasm resources, including

monogenic mutants with inhibited or altered ripen-

ing phenotypes (http://www.tgrc.ucdavis.edu/,

http://www.zamir.sgn.cornell.edu/mutants/). There

is also a long history and a wealth of biochemical and

molecular data relative to the processes that are in-

volved during ripening, and a large platform of tools

for functional genomics is continually being devel-

oped, including an emerging genome sequence (Fei

and others 2006; Mueller and others 2005).

Differential screens, candidate gene analysis,

gene expression proﬁling, and digital gene expres-

sion analysis have led to the identiﬁcation of hun-

dreds of genes whose expression proﬁles change

during the course of fruit development and ripening

(Alba and others 2005; Fei and others 2004; Picton

and others 1993a, b; Slater and others 1985; Ze-

gzouti and others 1999). Through a combination of

approaches many of the downstream components

that mediate the biochemical changes associated

with ripening have been deﬁned. For example, cell

wall hydrolases, the enzymes involved in carotenoid

synthesis and sugar metabolism, and some of the

enzymes involved in the generation of ﬂavor and

aroma compounds have been characterized (Chen

and others 2004a, b; Fridman and others 2004;

Hirschberg 2001; Rose and Bennett 1999; Tieman

and others 2006). The pathways that determine the

competency of a fruit to ripen or the signals that

initiate the ripening program are less well deﬁned,

although the molecular identiﬁcation of mutants

that are impaired in fruit ripening are beginning to

yield valuable insight into some of these genetic

pathways, and multiple hormones, including jasm-

onates, auxin, and brassinosteroids, have all been

implicated in the promotion of ripening in various

species (Fan and others 1998; Given and others

1988; Manning and others 2006; Symons and oth-

ers 2006; Vardhini and Rao 2002; Vrebalov and

others 2002). Signaling through the plant hormone

ethylene, however, remains the most well-deﬁned

pathway that mediates the phenotypic changes that

occur during ripening. Treatment of various fruits

with inhibitors that block ethylene synthesis or ac-

tion or the manipulation of these processes by

transgenic or mutant approaches have revealed the

essential role of this hormone in regulating fruit

ripening (Hobson and others 1984; Klee and others

1991; Lanahan and others 1994; Oeller and others

1991; Picton and others 1993a, b; Yang and Hoff-

man 1984). In this review we summarize our cur-

rent understanding of ethylene biosynthesis and

signaling pathways in relation to fruit ripening.

Major emphasis is placed on knowledge obtained

using the tomato model system, although where

appropriate we highlight discoveries and novel

ﬁndings in other fruit crop species.

EGULATION OF

THYLENE

IOSYNTHESIS

URING

RUIT

IPENING

Fruits have classically been categorized based upon

their abilities to undergo a program of enhanced

ethylene production and an associated increase in

respiration rate at the onset of ripening. Fruits that

undergo this transition are referred to as climacteric

and include tomato, apple, peach, and banana,

whereas fruits that do not produce elevated levels of

ethylene are known as nonclimacteric and include

citrus, grape, and strawberry. However, these dis-

tinctions are not absolute, as closely related melon

and capsicum species can be both climacteric and

nonclimacteric (see below for further discussion)

and some so-called nonclimacteric fruits display

enhanced ripening phenotypes in response to

exogenous ethylene (see below for further discus-

sion). Nevertheless, increased ethylene synthesis at

the onset of ripening is required for the normal

ripening of many fruits.

Two systems of ethylene production have been

deﬁned in plants. System 1 functions during normal

growth and development and during stress re-

sponses, whereas system 2 operates during ﬂoral

senescence and fruit ripening. System 1 is autoin-

hibitory, such that exogenous ethylene inhibits

synthesis, and inhibitors of ethylene action can

stimulate ethylene production (Figure 1). In con-

trast, system 2 is stimulated by ethylene and is

therefore autocatalytic, and inhibitors of ethylene

action inhibit ethylene production (McMurchie and

others 1972).

The biochemical features of the ethylene biosyn-

thesis pathway in higher plants are well deﬁned and

have been reviewed previously (Bleecker and Kende

2000). Brieﬂy, ethylene is synthesized from methi-

onine in three steps: (1) conversion of methionine to

S-adenosyl-L-methionine (SAM) catalyzed by the

enzyme SAM synthetase, (2) formation of 1-

aminocyclopropane-1-carboxylic acid (ACC) from

SAM via ACC synthase (ACS) activity, and (3) the

conversion of ACC to ethylene, which is catalyzed

by ACC oxidase (ACO). The formation of ACC also

leads to the production of 5¢-methylthioadenosine

144 Cornelius S. Barry and others

(MTA), which is recycled via the methionine cycle to

yield a new molecule of methionine. Increased res-

piration provides the ATP required for the methio-

nine cycle and can lead to high rates of ethylene

production without high levels of intracellular

methionine. SAM is an important methyl donor and

is involved in multiple aspects of cellular metabo-

lism. Consequently, the two committed steps in the

synthesis of ethylene are the formation of ACC and

its conversion to ethylene. The genes encoding ACS

and ACO have thus been studied in more detail than

other enzymes in the pathway, although there is

evidence that several other genes involved in

methionine synthesis and the methionine salvage

pathway are differentially expressed during ripening

and in response to ethylene (Alba and others 2005;

Zegzouti and others 1999).

ACS and ACO are encoded by multigene families

in higher plants, with tomato possessing at least

nine ACS (LEACS1A,LEACS1B, and LEACS2-8) and

ﬁve ACO (LEACO1-5) genes (Barry and others 1996;

Nakatsuka and others 1998; Oetiker and others

1997; Van-der-Hoeven and others 2002; Zarem-

binski and Theologis 1994). Expression analysis has

revealed that at least four ACS (LEACS1A,LEACS2,

LEACS4,LEACS6) and three ACO (LEACO1,LEACO3,

LEACO4) genes are differentially expressed in to-

mato fruit (Barry and others 1996, 2000; Nakatsuka

and others 1998; Rottmann and others 1991). LE-

ACO1,LEACO3,andLEACO4 are expressed at low

levels in green fruit that are in a system 1 mode of

ethylene synthesis, but the transcripts of each in-

crease at the onset of ripening as the fruit transition

to system 2 ethylene production and response.

During ripening, LEACO1 and LEACO4 are sustained

in expression, whereas the increase in LEACO3

expression is transient (Barry and others 1996; Na-

katsuka and others 1998). In the case of LEACO1

and LEACO4, ripening-related increases in transcript

abundance are largely blocked by 1-MCP treatment,

indicating that these genes are positively regulated

by ethylene. The regulation of ACS gene expression

during fruit ripening has been investigated using a

combination of ethylene and inhibitor studies to-

gether with expression analysis in various ripening

mutants (Barry and others 2000; Nakatsuka and

others 1998). The ripening-inhibitor (rin)andnon-

ripening (nor) mutants fail to undergo the typical

ripening-related increase in ethylene synthesis

(system 2) and respiration that occurs in wild-type

fruit and, as such, maintain a low-level system 1-

type ethylene production as they mature (Tig-

chelaar and others 1978). LEACS6 is expressed in

wild-type green fruit but rapidly declines at the

onset of ripening during the transition to system 2

ethylene synthesis. In contrast, LEACS6 transcripts

persist throughout development and ripening in the

rin mutant (Barry and others 2000). Ethylene and

1-MCP treatments indicated that this ripening-re-

lated decline was mediated by ethylene, suggesting

that LEACS6 is responsible for low-level ethylene

production in preclimacteric fruit (Barry and others

2000; Nakatsuka and others 1998). LEACS1A is also

expressed in preclimacteric fruits and declines upon

ethylene treatment, but transcripts show a transient

increase at the onset of ripening that is rin depen-

dent, suggesting that this gene may be important in

regulating ethylene synthesis during the transition

from system 1 to system 2 ethylene synthesis (Barry

and others 2000). LEACS4 is not expressed in green

fruit but is induced at the onset of ripening. This

induction is dependent on rin and is stimulated by

ethylene. LEACS2 expression is also induced at the

onset of ripening; this induction requires ethylene

but is independent of rin (Barry and others 2000;

Nakatsuka and others 1998). Therefore, it seems

likely that LEACS1A and LEACS4 are responsible for

initiating system 2 ethylene synthesis and that this

is maintained by a combination of LEACS2 and LE-

ACS4. The speciﬁc transcription factors that mediate

the changes in ACS and ACO gene expression at the

onset of ripening remain to be determined. In a

recent study, a 40-bp promoter fragment of LEACS2

was identiﬁed that is required for ethylene-induc-

ing-xylanase (EIX) responsiveness. Both in vitro and

in vivo studies indicated that a novel cysteine pro-

tease, designated LeCP, was bound to this region and

was capable of inducing LEACS2 expression when

Figure 1. Differential expression of ACS and ACO genes

associated with system 1 and system 2 ethylene synthesis

during fruit development and ripening in tomato. Au-

toinhibition of ethylene synthesis during system 1 ethyl-

ene production is mediated by a reduction in LeACS1A and

6expression. Autocatalytic ethylene synthesis at the onset

of fruit ripening is mediated through ethylene-stimulated

expression of LeACS2 and 4and LeACO1 and 4(see text for

details).

Ethylene and Fruit Ripening 145

overexpressed in tomato leaves, suggesting that this

protein may possibly have dual functions as a pro-

tease and a transcriptional regulator (Matarasso and

others 2005). However, it remains to be determined

whether the same promoter region of LEACS2 and

LeCP are required for ethylene-regulated ripening

induction. Although regulation of ACS at the level

of expression and transcript abundance is clearly

important for ripening-related ethylene synthesis,

there is considerable evidence that regulation of

ACS activity, through protein phosphorylation and

turnover, also plays a critical role in the function of

this enzyme (for review, see Argueso and others,

this issue).

The physiologic and molecular pathways that act

to initiate the transition from a system 1 to a system

2 mode of ethylene synthesis at the onset of rip-

ening remain undeﬁned. However, a recent study

performed on detached persimmon (Diospyros kaki

Thunb.) fruit indicated that ripening-related ethyl-

ene synthesis in the fruit was initiated by a burst of

drought-induced ethylene synthesis from the fruit

calyx following harvest (Nakano and others 2003).

Detached persimmon fruit initiated ethylene pro-

duction and associated loss of ﬁrmness within two

days after harvest. Treatment of fruit parts with the

ethylene action inhibitor 1-methylcyclopropene (1-

MCP) inhibited ethylene synthesis in all tissues

except the calyx, indicating that the ethylene pro-

duced from the calyx was independent of ethylene

itself. Induction of an ACC synthase gene, DkACS2,

correlated with ethylene synthesis and was unaf-

fected by 1-MCP. The calyx of fruits stored at high

humidity initiated ethylene production and DkACS2

expression four days later than fruits stored at low

humidity. Similar delays were also observed in the

pulp under high-humidity conditions. These data

clearly show a role for water loss in regulating the

onset of ethylene synthesis in detached persimmon

fruit. Although this mechanism has been described

only in persimmon fruit to date, certain fruits,

including avocado and the wild species of tomato

Solanum chilense and Solanum peruvianum, are

known to initiate ripening once abscission has oc-

curred from the parent plant. It is possible that

water loss from these fruits following detachment

could be a possible mechanism to initiate ethylene

synthesis and ripening (Grumet and others 1981;

Dopico and others 1993).

ISTINCT

RANSCRIPTION

ACTORS

PSTREAM OF

THYLENE

YNTHESIS TO

EGULATE

RUIT

IPENING

With the exception of ethylene, the signaling

pathways that regulate fruit ripening remain largely

undeﬁned. In tomato, three pleiotropic nonripening

mutants, ripening-inhibitor (rin), non-ripening (nor),

and Colorless non-ripening (Cnr), have been described

in which virtually all aspects of the ripening process

are inhibited, including ethylene synthesis, in-

creased respiration, carotenoid accumulation, soft-

ening, and aroma production (Thompson and

others 1999; Tigchelaar and others 1978) (Figure 2).

In these three mutants, the typical ripening-associ-

ated rise in autocatalytic ethylene synthesis is

blocked due to abnormal regulation of ACS expres-

sion (see above). Although ethylene synthesis is

blocked in these mutants, studies using rin and nor

fruits have indicated that they retain the capacity to

synthesize wound ethylene, indicating that the

mutations are not simply the result of a general

block in ethylene synthesis (Lincoln and Fischer

1988; Yokotani and others 2004). Similarly, exog-

enous ethylene does not restore ripening in these

mutants, although ethylene-regulated gene

expression can be partially restored, indicating that

rin,nor, and Cnr fruits do not display ethylene

insensitivity (Barry and others 2000; Grifﬁths and

Figure 2. Fruit ripening mutants of tomato. From left to right, ripe fruit of wild type (cultivar Ailsa Craig) and near

isogenic lines homozygous for the ripening-inhibitor (rin), non-ripening (nor), Never-ripe (Nr), and Green-ripe (Gr) loci. Note

association of the macrocalyx (mc) (large sepal) phenotype with the rin mutation. The rin and nor loci act upstream in the

ripening regulatory pathway and are required for system 2 ethylene synthesis during fruit ripening. The nonripening

phenotypes of Nr and Gr are caused by reduced ethylene responsiveness (see text for details).

146 Cornelius S. Barry and others

others 1999; Thompson and others 1999; Yen and

others 1995; Yokotani and others 2004). Together

these data suggest that rin,nor, and Cnr act up-

stream of ethylene in the ripening cascade and

determine the competency of the fruit to ripen.

The molecular identities of the rin and Cnr loci

have been determined using positional cloning

strategies, and both encode different classes of

transcription factor (Manning and others 2006;

Vrebalov and others 2002). The rin locus harbors a

deletion occurring between two adjacent MADS-

box genes. Genetic complementation and antisense

experiments conﬁrmed that one of these genes,

termed LEMADS-RIN, was responsible for conferring

the nonripening phenotype of the rin mutant,

whereas the associated macrocalyx (mc) phenotype

was the result of a promoter deletion in a second

gene, termed LEMADS-MC (Vrebalov and others

2002). RIN is a member of the SEPALLATA sub-

family of MADS-box genes, whereas MC is a mem-

ber of the APETALA 1 subfamily (Litt and Irish 2003;

Malcomber and Kellogg 2005). As MADS-box pro-

teins have been shown to act together in multimeric

complexes, it is possible that other MADS-box genes

act together with RIN to regulate fruit ripening in

tomato. Indeed, expression analysis and data min-

ing of EST collections have revealed several possible

candidates to fulﬁll this role (Fei and others 2004;

Giovannoni 2004). The Cnr mutation is the result of

an epigenetic mutation that causes hypermethyla-

tion and reduced expression of a SQUAMOSA PRO-

MOTER BINDING PROTEIN (SBP) gene (Manning

and others 2006). SBP-box proteins have been

shown to directly regulate the expression of MADS-

box genes, raising the possibility that CNR may act

to directly inﬂuence the expression of RIN or other

MADS-box genes during fruit ripening. The char-

acterization of the RIN and CNR transcription factors

is currently the subject of intense research. It will be

particularly interesting to identify the direct targets

of these proteins and determine how they are able

to regulate ethylene synthesis during ripening.

THYLENE

IGNALING IN

OMATO

ONSERVATION AND

IVERSITY

Much of our knowledge concerning the mode of

action of ethylene in plants has been generated

from the use of the triple-response screen in Ara-

bidopsis to identify mutants that are either insensi-

tive to ethylene or show enhanced ethylene

responses in the absence of exogenous ethylene.

The power of Arabidopsis molecular genetics has

facilitated the rapid identiﬁcation of many compo-

nents of the signaling pathway from an initial mu-

tant phenotype. The components of ethylene

signaling and their mechanisms of action in Ara-

bidopsis are the subject of two additional reviews by

Hall and others and Li and Guo, in this special issue.

We focus our discussions on ethylene signaling re-

search in fruit crop species, primarily reviewing re-

search on tomato and how ﬁndings differ from the

Arabidopsis model.

THYLENE

ECEPTORS

The development of the triple-response screen in

Arabidopsis (Bleecker and others 1988; Guzman and

Ecker 1990), together with the identiﬁcation of

ETR1 as an ethylene receptor (Chang and others

1993), led directly to the identiﬁcation of an eth-

ylene-insensitive mutant of tomato and the cloning

of the family of ethylene receptors to which it be-

longed. The Never-ripe (Nr) mutant was initially

described as a nonripening mutant 50 years ago

(Rick 1956). Although Nr clearly displayed a dra-

matic inhibition of fruit ripening (Figure 2), other

phenotypes associated with Nr had been over-

looked. However, in light of the ﬁndings from Ara-

bidopsis, Lanahan and coworkers (1994) showed

that Nr displayed a range of phenotypes that could

be directly attributed to reduced ethylene sensitiv-

ity. The semidominant ethylene-insensitive pheno-

type of Nr is reminiscent of the etr1 ethylene

receptor mutant, and with the availability of the

ETR1 gene for use as a heterologous probe, several

ethylene receptor homologs were identiﬁed in to-

mato, one of which was found to cosegregate with

the Nr phenotype on chromosome 9 (Yen and

others 1995). Subsequent molecular analysis re-

vealed a single C > T base change that results in

conversion of a conserved proline residue into leu-

cine (Wilkinson and others 1995). This proline

residue lies within the N-terminal ethylene binding

domain of the NR protein at a similar location to

dominant ethylene-insensitive alleles of ETR1 that

disrupt ethylene binding (Chang and others 1993;

Hall and others 1999).

To date, a total of six ethylene receptors have

been identiﬁed in tomato: LeETR1,LeETR2,NR (also

referred to as LeETR3), and LeETR4,5, and 6(Klee

2004; Lashbrook and others 1998; Payton and others

1996; Tieman and Klee 1999; Wilkinson and others

1995; Zhou and others 1996). Based on structural

similarity, the Arabidopsis ethylene receptors have

been classiﬁed into two subfamilies (Guo and Ecker

2004). Subfamily-1 consists of ETR1 and ERS1

that share three N-terminal membrane-spanning

Ethylene and Fruit Ripening 147

domains and a conserved carboxy terminus histidine

(His) kinase domain. LeETR1,LeETR2,andNR pos-

sess a structure that is consistent with the subfamily-

1 receptors. Subfamily-2 receptors in Arabidopsis

(ERS2, ETR2, and EIN4) lack a complete His kinase

domain and possess an additional transmembrane-

spanning domain at the N terminus. The tomato

receptors LeETR4,5, and 6can be classiﬁed as sub-

family-2 receptors. In addition, receptor structure

differs with regard to the presence or absence of a

receiver domain at the carboxy terminus. Arabidopsis

ETR1, ETR2, and EIN4 all possess a receiver domain,

as do all of the tomato receptors with the exception

of NR. Thus, the receptor complement varies slightly

between Arabidopsis and tomato. Tomato contains

an additional subfamily-1 receptor compared with

Arabidopsis, but contains only a single receptor

lacking a receiver domain (NR), whereas Arabidopsis

has two receptors (ERS1 and ERS2) that lack this

domain. It is evident that the ethylene receptor

family shows a high degree of structural divergence

in plants, but despite this diversity all of the recep-

tors thus far examined have the capacity to bind

ethylene when expressed in yeast (OÕMalley and

others 2005).

The tomato ethylene receptors are differentially

expressed in organs and tissues at various stages of

development and in response to exogenous stimuli

(Ciardi and others 2001; Lashbrook and others

1998; Moeder and others 2002; Tieman and Klee

1999). The changes in receptor proﬁles appear to be

quantitative rather than qualitative, with expres-

sion of receptors detected in all tissues so far

examined, implying that all tissues have the po-

tential to respond to ethylene. However, speciﬁc

receptors appear to be more prevalent in certain

tissues; for example, NR and LeETR4 are highly ex-

pressed in reproductive tissues and transcript

abundance is enhanced during fruit ripening (Ti-

eman and Klee 1999; Wilkinson and others 1995).

Different expression levels of receptors may poten-

tially lead to different pools of ethylene receptors

that may act to regulate speciﬁc responses.

Characterization of the individual functions of

members of the ethylene receptor gene family is

subject to ongoing investigation. Experiments de-

signed to downregulate speciﬁc receptor isoforms

using antisense suppression have been reported for

LeETR1,NR,andLeETR4 (Hackett and others 2000;

Tieman and others 2000; Whitelaw and others

2002). Downregulation of LeETR1 expression in

transgenic plants did not alter fruit ripening but

resulted in plants with shorter internodes and re-

duced rates of ﬂoral abscission (Whitelaw and oth-

ers 2002). Downregulation of NR expression in a

wild-type background did not result in any dramatic

phenotypes but did result in subtle changes indica-

tive of slightly delayed fruit ripening, that is, re-

duced rates of ethylene synthesis and slower

carotenoid accumulation (Tieman and others 2000).

Elevated expression of LeETR4 was detected in the

NR antisense lines, suggesting that this receptor may

compensate for loss of NR. Reduction of LeETR4

expression using an antisense transgene resulted in

plants with enhanced ethylene sensitivity mani-

fested through extreme epinasty, increased ﬂoral

abscission, enhanced triple response, and acceler-

ated fruit ripening, conﬁrming that LeETR4 acts as a

negative regulator of ethylene responses in tomato

(Tieman and others 2000). Interestingly, these

phenotypes could be complemented by overex-

pression of a NR transgene, indicating that these two

receptors are functionally redundant, a phenome-

non that is unexpected when one considers that

they are extremely divergent (see above). Although

studies of individual receptor function in tomato

requires additional experimentation, an obvious

difference between the tomato and Arabidopsis sys-

tems is evident. Reduction of the subfamily-2

receptor LeETR4 in transgenic plants leads to strong

phenotypic effects throughout the plant, whereas

single loss-of-function mutants in type 2 receptors

of Arabidopsis do not show dramatic phenotypic

changes (Hua and Meyerowitz 1998).

GREEN-RIPE E

NCODES A

OVEL

EGULATOR OF

THYLENE

ESPONSES

The dominant Green-ripe (Gr) and Never-ripe 2 (Nr-2)

mutants of tomato fail to fully ripen and possess a

fruit phenotype very similar to that of Nr (Jarret and

others 1984; Kerr 1958, 1982) (Figure 2). This

similarity prompted an examination of the ethylene

physiology of Gr and Nr-2. Examination of ethylene

synthesis and responses in Gr and Nr-2 fruits indi-

cated that reduced ethylene responsiveness was the

basis for ripening inhibition in these mutants (Barry

and others 2005). However, unlike Nr, which shows

reduced ethylene sensitivity throughout the whole

plant, Gr and Nr-2 show reduced ethylene sensi-

tivity predominantly in fruit and ﬂoral tissues with

weak ethylene insensitivity evident in roots. Dark-

grown Gr and Nr2 hypocotyls and petioles maintain

normal ethylene responsiveness. High-resolution

genetic and physical mapping of the Gr and Nr-2 loci

revealed that they were both linked to a 38-kb

interval of the long arm of chromosome 1, sug-

gesting that they may be allelic. This hypothesis was

conﬁrmed when sequence analysis of this region

148 Cornelius S. Barry and others

identiﬁed a 334-bp deletion in both Gr and Nr-2

mutants compared to wild type. The deletion occurs

at the junction between the 5¢-UTR and the pro-

moter of a gene of unknown function and causes

ectopic expression of the gene in Gr and Nr-2 mu-

tant backgrounds, consistent with a dominant gain-

of-function mutation (Barry and Giovannoni 2006).

To avoid confusion with the NR ethylene receptor,

the gene residing at the Gr and Nr-2 locus was

designated GREEN-RIPE (GR).

Ectopic expression of GR under the control of the

CaMV35S promoter recreated the Gr mutant phe-

notype but did not lead to plants that displayed

whole-plant ethylene insensitivity despite high

levels of transgene expression. This suggests that GR

is able to selectively modify ethylene responsiveness

in a tissue-dependent manner indicating that com-

ponents of the ethylene signaling pathway must be

distinct in different tissues of tomato (Barry and

Giovannoni 2006). These differences have yet to be

deﬁned, but in a separate study a homolog of GR,

REVERSION TO ETHYLENE SENSITIVITY 1 (RTE1),

was identiﬁed as a speciﬁc suppressor of ethylene

insensitivity mediated by the etr1-2 mutant allele of

Arabidopsis, suggesting that RTE1 and, therefore,

possibly GR act at the level of the ethylene receptors

(Resnick and others 2006). It is tempting to specu-

late that the speciﬁcity of GR action in tomato may

be linked to different pools of ethylene receptors

that are present in different tissues in tomato

(Lashbrook and others 1998; Tieman and Klee

1999).

GR and RTE1 encode predicted transmembrane

proteins of unknown function that are conserved in

plants, animals, and protists but are not present in

bacterial or fungal genomes. Higher plants typically

contain two or three GR homologs, whereas animal

and protist genomes possess a single copy of this

gene. In plants there appears to be two phyloge-

netically distinct clades. One clade contains GR,

RTE1, and a closely related tomato gene designated

GREEN-RIPE LIKE 1 (GRL1). The second clade

contains distant homologs of GR and RTE1 desig-

nated GRL2 and RTE1 HOMOLOG (RTH). GR and

RTE1 have clear impacts on ethylene responses in

plants, but it remains to be determined if GRL1,

GRL2, and RTH also function in the ethylene re-

sponse pathway or some other aspect of cellular

metabolism. The second scenario seems more likely,

particularly for the more distinct GRL2 and RTH

genes and in light of the fact that animals and

protists are not known to signal using ethylene. The

cloning of GR and RTE1 has identiﬁed new proteins

that can inﬂuence the ethylene response pathway

in plants; however, determining how these proteins

interact and function within the context of other

pathway components is required.

ULTIPLE

CTR K

INASES ARE

RESENT IN

ROP

LANTS

The constitutive triple response (ctr1) mutant of Ara-

bidopsis was identiﬁed in a genetic screen designed

to identify dark-grown seedlings that possess the

triple-response phenotype in the absence of exoge-

nous ethylene (Guzman and Ecker 1990). The CTR1

gene encodes a protein with high similarity to the

mammalian RAF serine/threonine MAP kinase ki-

nase kinase (MAP3K) and acts as a negative regu-

lator of ethylene responses (Kieber and others

1993). CTR1 localizes to the endoplasmic reticulum,

the same location as the ETR1 ethylene receptor,

and interacts preferentially with the type I ethylene

receptors ETR1 and ERS1 (Clark and others 1998;

Gao and others 2003). The identiﬁcation of a

MAP3K that functions within the ethylene signaling

pathway has led to the speculation that a MAP ki-

nase cascade may mediate ethylene responses in

Arabidopsis, although this hypothesis awaits experi-

mental validation.

To date, four CTR1 homologs have been isolated

from tomato: tCTR1 (also known as ER50), tCTR2,

tCTR3, and tCTR4 (Adams-Phillips and others 2004a,

b; Leclercq and others 2002; Lin and others 1998;

Zegzouti and others 1999). These genes were iden-

tiﬁed either using differential display (ER50) or

through heterologous hybridization using the Ara-

bidopsis CTR1 gene as a probe. Phylogenetic analysis

has indicated that tCTR1,tCTR3, and tCTR4 are clo-

sely related to Arabidopsis CTR1. In addition, these

three genes are all able to complement, at least

partially, the weak ctr1-8 allele of Arabidopsis, sug-

gesting that the tomato genes are functionally

equivalent to CTR1. Interestingly, the efﬁcacy of

complementation follows the phylogenetic rela-

tionship of the tomato genes to Arabidopsis CTR1

such that tCTR3 is able to fully complement ctr1-8,

whereas tCTR1 and tCTR4 display only partial com-

plementation (Adams-Phillips and others 2004a, b;

Leclercq and others 2002). This may be indicative of

slightly different signaling speciﬁcities of these pro-

teins. tCTR1,3,and4show differential expression in

various plant tissues, but as in the case of receptor

gene expression, all tissues express CTR-like genes

(Adams-Phillips and others 2004a, b; Leclercq and

others 2002). tCTR2 is more divergent and is the

likely ortholog of the ENHANCED DISEASE RESIS-

TANCE 1 (EDR1) gene of Arabidopsis that has been

implicated in disease resistance, stress responses,

Ethylene and Fruit Ripening 149

and ethylene-induced leaf senescence and may act

at the interface between the ethylene and salicylic

acid signaling pathways (Adams-Phillips and others

2004a, b; Frye and others 2001; Tang and others

2005).

These data indicate that tomato possesses at least

three CTR genes, whereas Arabidopsis possesses only

a single gene. Subsequent analysis of EST and

genomic sequence repositories has uncovered evi-

dence for multiple CTR-like genes in a number of

species, suggesting that the single gene found in

Arabidopsis may be the exception (Adams-Phillips

and others 2004a, b). The presence of multiple CTRs

raises questions as to the individual functions of

these genes and the level of redundancy that

operates within this gene family. These questions

will need to be addressed through the use of RNAi

targeted at individual members of the family and by

gain-of-function analysis. However, virus-induced

gene silencing (VIGS) of a generic tCTR sequence

led to tomato plants that showed severe epinasty

and upregulation of ethylene-induced gene

expression, indicating that silencing of tCTR genes

can mimic the Arabidopsis ctr1 mutant phenotype

(Liu and others 2002). Because CTR1 is able to

interact with at least some of the Arabidopsis ethyl-

ene receptors, it is likely that the tCTR proteins will

also interact with the tomato ethylene receptors.

The presence of larger gene families for both CTRs

and the ethylene receptors in tomato compared to

Arabidopsis indicates that the possible interactions

between these two families is potentially more

complex and that localization experiments will need

to be performed together with interaction studies to

conﬁrm that any interactions are physiologically

meaningful.

RANSCRIPTIONAL

EGULATION OF

THYLENE

ESPONSES

URING

RUIT

IPENING

In Arabidopsis, downstream ethylene responses are

mediated by two classes of transcription factors en-

coded by EIN3 and ERF gene families. ETHYLENE-

INSENSITIVE 3 (EIN3) also includes the EIN3-LIKE

(EIL) family members and the ETHYLENE RE-

SPONSE FACTOR (ERF) family is inclusive of genes

referred to as ETHYLENE-RESPONSIVE ELEMENT

BINDING PROTEIN (EREBP). EIN3 is a positive reg-

ulator of ethylene responses with loss of function

resulting in ethylene insensitivity, whereas over-

expression results in a constitutive triple-response

phenotype (Chao and others 1997). Emerging evi-

dence suggests that the ethylene response pathway

is regulated at least in part by turnover of the EIN3

protein. In the absence of ethylene, two partially

redundant F-box proteins, EIN3-binding F-box 1

and 2 (EBF1 and EBF2), target EIN3 for degrada-

tion. A negative feedback loop exists whereby EIN3

is self-regulating through directly inﬂuencing the

accumulation of EBF1 and EBF2 (Gagne and others

2004; Guo and Ecker 2003; Potuschak and others

2003; Yanagisawa and others 2003). In a recent

study, the EIN5 protein was identiﬁed as the XRN4

5¢ﬁ3¢exoribonuclease (Olmedo and others 2006).

Both EBF1 and EBF2 mRNAs are signiﬁcantly more

abundant in the ein5 mutant background than in

wild type, suggesting that the wild-type function of

EIN5 is to regulate the accumulation of EIN3 via

turnover of EBF1/2 transcripts.

EIN3 binds a conserved motif known as the pri-

mary ethylene responsive element (PERE) that is

present within the promoters of ERF1 and several

senescence and ripening-related genes, including

E4,GST1,andLeACO1 (Solano and others 1998).

ERF1 expression is rapidly induced by ethylene, and

overexpression of ERF1 confers a constitutive eth-

ylene response phenotype. ERFs in turn bind to the

GCC-box that is present in the promoters of several

stress- and pathogen-responsive genes, including

chitinases and PDF1.2 (Solano and others 1998).

ERF1 and related genes form a subgroup of the large

APETALA2 (AP2) family of DNA-binding proteins

that consists of 145 members in Arabidopsis (Gutt-

erson and Reuber 2004). These transcription factors

act to both positively and negatively regulate tran-

scription and are involved in a wide range of

developmental processes, responses to environ-

mental challenges, and pathogen infections.

To date, four EIN3-LIKE genes have been de-

scribed in tomato, LeEIL1-4 (Tieman and others

2001; Yokotani and others 2003). LeEIL1-3 are each

able to complement the Arabidopsisein3-1 mutant

allele, indicating that they are able to function in

the ethylene signaling pathway. Antisense sup-

pression of EIL1-3 in tomato revealed that this gene

family is functionally redundant (Tieman and oth-

ers 2001). However, overexpression of a GFP-tagged

EIL1 in the Nr mutant restored normal fruit ripen-

ing and the expression of a subset of ethylene-

inducible genes in transgenic fruit. In addition,

petiole epinasty was restored in the 35S:EIL1:GFP

transgenic lines, but seedling responses remained

unaltered, suggesting that individual members of

the tomato EIL family may perform speciﬁc func-

tions in vivo (Chen and others 2004a, b). The

observation that sequences related to the PERE are

contained within the promoters of the ethylene-

regulated and ripening-related genes E4 and LeACO1

150 Cornelius S. Barry and others

suggests that EIN3 proteins may directly regulate

their transcription and that of other coregulated

genes (Solano and others 1998).

Thirty-six ERF1-like genes have been built into an

Arabidopsis and rice phylogeny and it is likely that as

more ESTs and tomato genomic sequence become

available, this number will increase (Gutterson and

Reuber 2004). Given the high copy number of this

gene family, it is not surprising that several mem-

bers have been isolated from fruit and show differ-

ential expression patterns during fruit development

and ripening (Alba and others 2005; Tournier and

others 2003). Because of the large size of this gene

family, assigning functions to individual family

members will be a daunting task, although the use

of phylogeny coupled to phenotypic information

from Arabidopsis may help guide studies in tomato

(Gutterson and Reuber 2004). Furthermore, the

promoters of many of the genes that have been

associated with ripening do not contain the GCC-

box that forms the binding site of the ERF protein,

suggesting that these factors may have a limited role

in the regulation of ripening. Interestingly, a num-

ber of stress- and defense-associated genes whose

expression can be enhanced by ethylene have been

shown to be induced during fruit ripening (Alba and

others 2005; Picton and others 1993a, b; Zegzouti

and others 1999). Stress-related genes often contain

the GCC-box within their promoter regions that can

be directly targeted by ERF proteins. Therefore, it is

possible that the ERF proteins that are present in

tomato fruit may regulate the expression of this

subset of ripening-related genes, a hypothesis that

will become open to testing as the tomato genome

sequence becomes available in the next few years

(Mueller and others 2005).

Clearly ﬁnding the immediate targets and in vivo

function of both the EIN3-LIKE and ERF protein

families during ripening will be important if a

transcriptional network of ethylene-regulated gene

expression is to be elucidated. A recent study

examining the comparative transcriptome of tomato

fruit during development and ripening in wild type

and the Nr mutant revealed that 37% of the gene

expression changes observed were inﬂuenced by

ethylene (Alba and others 2005). Furthermore,

clustering of gene expression proﬁles revealed that

many of these changes were coordinated, implying

that large groups of genes are coregulated. Although

this approach addressed only steady-state mRNA

levels and therefore did not distinguish between

transcriptional and post-transcriptional events, it is

highly likely that a large proportion of these chan-

ges involve at least partial ethylene-regulated tran-

scriptional control. Dissecting speciﬁc signaling

modules within this vast sea of ethylene-regulated

events will be challenging. One approach that may

be useful in this regard is a scaled-up version of the

experimental system used by Chen and others

(2004a, b) to study EIL1 function in tomato (de-

scribed above). A series of EIL or ERF constructs

under the control of an inducible promoter could be

introduced into the Nr mutant, possibly using

transient expression systems, and early changes in

gene expression could be monitored by microarrays

following induction. This approach could reveal the

identities of subsets of coregulated genes or path-

ways and provide information about potential tar-

gets of EILs and ERFs in tomato that could then be

addressed by in vitro and in vivo DNA-binding

studies.

LIMACTERIC

ONCLIMACTERIC

AND

OMEWHERE IN

ETWEEN

ARIATION IN

THE

THYLENE

HYSIOLOGY OF

IPENING

RUITS

Fruits have been classically categorized into cli-

macteric and nonclimacteric based on increased

ethylene synthesis and a concomitant rise in the

rate of respiration during ripening (Lelievre and

others 1997). The role of ethylene as the ‘‘ripening

hormone’’ in climacteric fruits such as tomato, ap-

ple, and banana has been ﬁrmly established. How-

ever, there is an increasing body of experimental

evidence that implicates ethylene in the ripening of

fruits that have been classically thought of as non-

climacteric. There are also a number of species in

which the fruits of different varieties and cultivars

exhibit both climacteric and nonclimacteric behav-

ior.

A Role for Ethylene in the Ripening of

‘‘Nonclimacteric’’ Fruit

The highly sensitive technique of laser photoacou-

stic spectroscopy coupled with the development of

speciﬁc apparatus to determine in planta ethylene

production in fruits and ﬂowers of strawberry dur-

ing development and ripening has revealed an in-

crease in ethylene production and a concomitant

rise in respiration rate in red ripe strawberry fruits

(Iannetta and others 2006). Furthermore, experi-

ments with the ethylene action inhibitor silver

thiosulfate revealed that this increased ethylene

production was under the control of a positive

feedback mechanism in ripe fruits, suggesting that a

form of autocatalytic ethylene production is opera-

tional during ripening in strawberry. The timing of

Ethylene and Fruit Ripening 151

this ripening-related increase in ethylene produc-

tion is distinct from the patterns of ethylene pro-

duction typically associated with the ripening of

fruits such as tomato. For example, ethylene pro-

duction increases at the onset of ripening in tomato,

and ripening is severely disrupted in transgenic fruit

where this phenomenon is blocked (Oeller and

others 1991). In contrast, the increase observed in

strawberry was not detected until 24 h after the

fruits had developed full red pigmentation. There-

fore, the physiologic role of the increased ethylene

synthesis in strawberry remains to be determined.

Nevertheless, in support of a role for ethylene in the

ripening of strawberry, a body of molecular evi-

dence concerning ethylene-related changes in gene

expression is becoming available. For example, in-

creased expression of cystathionine-gamma-syn-

thase (CGS) and ACC oxidase genes has been

reported during ripening of strawberry (Aharoni

and others 2002; Marty and others 2000; Trainotti

and others 2005). Similarly, increased expression of

ethylene receptor homologs in strawberry fruit was

also observed with increasing ripeness. In particular,

increased expression of FaETR2, a type II receptor

homolog that is closely related to the ripening-re-

lated LeETR4, was associated with ripening (Train-

otti and others 2005). In addition, the isolation and

characterization of a peptide methionine sulfoxide

reductase (PMSR) gene that is expressed late in

strawberry ripening were also recently described

(Pedraza-Lopez and others 2006). This gene is

homologous to the tomato ripening-related gene E4,

whose expression is regulated by ethylene in to-

mato and may be involved in the methionine sal-

vage pathway that operates during increased

ethylene synthesis (Lincoln and others 1987). Al-

though currently untested, it may be pertinent to

examine the relationship of ethylene and the

expression of FaPMSR and other genes whose

expression is induced at late stages of ripening of

strawberry fruit. The emerging data could be con-

sistent with either a regulatory role for ethylene in

as-yet deﬁned aspects of ripening in strawberry fruit

or a response to the dramatic cellular changes

associated with ripening and the concomitant loss in

cellular integrity and senescence characteristic of

this developmental process. Either answer would be

interesting because the former would demonstrate a

fundamental role of ethylene in ripening of most if

not all fruits and the other might point to a more

primal developmental response that may indeed

have been recruited through evolution as a signal-

ing system to catalyze the ripening process in a

subset of species. The creation of transgenic straw-

berry with reduced ethylene responsiveness, possi-

bly by expression of a dominant mutant allele of an

ethylene receptor, may help clarify this role.

Small but signiﬁcant increases in ethylene syn-

thesis at the onset of ripening have also been de-

tected in grape berries. Chervin and others (2004)

demonstrated the presence of a transient peak of

ethylene production in grapes just prior to the onset

of ripening, and experiments with 1-MCP indicated

that ethylene was required at this stage for the onset

of anthocyanin accumulation, fruit swelling, and

the decrease in acidity that is associated with rip-

ening. Concomitant with an ethylene-stimulated

rise in anthocyanin production, the abundance of

four transcripts encoding enzymes involved in

anthocyanin synthesis also increased following

ethylene treatment (El-Kereamy and others 2003).

In an additional study, 1-MCP treatment of grape

berries was found to partially repress the ripening-

induced expression of the VvADH2 gene that en-

codes an alcohol dehydrogenase (Tesniere and

others 2004). These studies suggest that ethylene

may inﬂuence multiple aspects of ripening in grape.

A similar strategy as described for strawberry above

could clarify the role of ethylene in grape berry

ripening.

Citrus is also classiﬁed as a nonclimacteric fruit,

but studies with inhibitors of ethylene action re-

vealed that ripening-related color changes in the

ﬂavedo are regulated by endogenous ethylene and

that ethylene treatments can stimulate both chlo-

rophyll breakdown and carotenoid accumulation

(Goldschmidt and others 1993; Purvis and Barmore

1981; Stewart and Wheaton 1972). Furthermore,

studies have shown some genes, including chloro-

phyllase, to be ethylene regulated in citrus fruit

(Alonso and others 1995; Jacob-Wilk and others

1999). Recently, as in the case of strawberry, auto-

catalytic ethylene production has also been de-

scribed in citrus fruit, but again with altered timing

relative to typical ripening climacteric fruit.

Whereas mature fruits exhibited no increased eth-

ylene production associated with ripening, har-

vested immature fruits produce high levels of

ethylene that can be further stimulated by ethylene

and propylene treatments and inhibited by 1-MCP,

indicating the autocatalytic nature of this phenom-

enon (Katz and others 2004). This study clearly

showed that citrus fruit have the capability to pro-

duce autocatalytic ethylene, although there may be

little signiﬁcance of this phenomenon in relation to

the ripening process. Citrus and other fruit trees

undergo a continual process of fruit drop or selective

abscission to ensure that resources are available to

allow the development of mature fruits. It is possible

that the climacteric behavior of young citrus fruitlets

152 Cornelius S. Barry and others

may be linked to this abscission process or may be

the result of stress during detachment as was wit-

nessed in persimmon fruit (see above).

Climacteric and Nonclimacteric:

Ethylene-Mediated Ripening Inhibition

Through Natural Allelic Variation

Apple cultivars are highly heterozygous and their

pedigrees are often poorly deﬁned. In addition, they

possess different ripening rates leading to varied

storage properties ranging from rapid postharvest

deterioration to cultivars that can be stored for up to

a year under optimal conditions. A clear positive

correlation exists between ethylene production

during storage and softening, which in turn is

tightly associated with postharvest deterioration

(Gussman and others 1993). Sunako and others

(1999) identiﬁed two allelic forms of the Malus

domestica ACS1 gene from the Golden Delicious

cultivar, MdACS1-1 and MdACS1-2.MdACS1-1 is

highly expressed at the onset of apple fruit ripening,

whereas MdACS1-2 transcripts are absent from rip-

ening fruits. Sequence analysis of these alleles re-

vealed only seven nucleotide substitutions within

the protein-coding region of MdACS1 and six of

these encoded silent mutations. However, more

sequence divergence was evident in the 5¢-ﬂanking

region, including the presence of a short inter-

spersed DNA element (SINE) in the MdACS1-2 allele.

Thirty-ﬁve apple cultivars were tested for the pres-

ence or the absence of the SINE element and results

indicated that cultivars were either homozygous for

either MdACS1-1 or MdACS1-2 or heterozygous for

each allele. In subsequent studies, cultivars that

were homozygous for the MdACS1-2 allele had sig-

niﬁcantly lower internal ethylene concentrations

than MdACS1-1 homozygotes or MdACS1-1/MdACS1-

2heterozygous individuals and possessed enhanced

storage capabilities and reduced rates of fruit drop

(Harada and others 2000; Oraguzie and others 2004;

Sato and others 2004). These data indicate that the

insertion of a SINE into the promoter of the MdACS1

gene is directly responsible for reduced gene

expression, lower ethylene production, and en-

hanced storage properties of certain apple cultivars.

This discovery provides a useful tool for selecting

new apple varieties with optimal storage properties.

Reduction in ACS gene expression and ethylene

production also seems to be responsible for the

nonripening phenotype in peach cultivars that carry

the recessive stony hard (hd) mutation. hd fruit fail to

soften on the tree or postharvest, although other

ripening traits such as color development, soluble

solids, and ﬂavor characteristics are fairly normal

(Haji and others 2001). Furthermore, the pheno-

type of hd can be reversed by ethylene treatment

(Haji and others 2003). Expression of a ripening-

related ACS gene, PpACS1, is eliminated in hd

backgrounds during ripening, although this gene

remains wound-inducible in both leaves and fruits

(Tatsuki and others 2006). The mechanism of the

reduction of PpACS1 expression in hd is currently

unknown. Southern analysis failed to reveal any

signiﬁcant structural differences in PpACS1 between

hd and a normal-ripening cultivar, indicating that

disruption of the promoter by a transposable ele-

ment, as in the case of the MdACS1-2 allele, is un-

likely. It is possible that a SNP or small deletion or

insertion may disrupt a ripening-speciﬁc transcrip-

tion factor binding site in the PpACS1promoter or

that the hd phenotype may be caused by a mutation

in a ripening-speciﬁc transcription factor.

Melon is also a fruit crop that exhibits great phe-

notypic diversity. Fruits of the cantaloupe type often

have netted skin and orange ﬂesh, are susceptible to

abscission, and produce large quantities of ethylene.

In contrast, melons of the ‘‘honey dew’’ type are of-

ten smooth skinned, have green or yellow ﬂesh, do

not abscise, and produce little or no ethylene during

ripening. Therefore, melons behave as both climac-

teric and nonclimacteric fruit, and the level of eth-

ylene production produced by melon fruit is directly

proportional to postharvest rates of decay (Zheng and

Wolff 2000). Perin and others (2002) examined the

ethylene physiology of a smooth-fruited, nonabscis-

ing melon designated PI161375. They found that

exogenous ethylene failed to induce abscission, fruit

ethylene production, or the expression of ethylene-

regulated genes, suggesting that PI161375 fruit is

ethylene insensitive. However, the seedling triple

response in this line was normal. Genetic analysis of a

recombinant inbred population generated from a

charentais (cantaloupe) ·PI161375 cross indicated

that fruit abscission and ethylene production were

controlled by two independent loci designated

Abscission layer (Al)-3 and Al-4. F

progeny of the

charentais ·PI161375 cross produced fruits that

abscised and produced a climacteric ethylene peak,

indicating that the alleles from the PI161375 parent

are recessive. The fruit and abscission zone-speciﬁc

responses in PI161375 share similarities to, but are

distinct from, tomato mutants that display inhibited

fruit ripening. For example, like the PI161375 line,

the rin,nor, and Cnr mutants can be classiﬁed as

nonclimacteric; however, they retain the capacity to

respond to exogenous ethylene at the level of gene

expression (Tigchelaar and others 1978; Thompson

and others 1999; Yen and others 1995; Yokotani and

others 2004). In contrast, the Gr mutant of tomato

Ethylene and Fruit Ripening 153

displays tissue-speciﬁc ethylene insensitivity associ-

ated with fruit ripening and abscission, but unlike

PI161375, Gr fruit are capable of synthesizing large

quantities of ethylene (Barry and others 2005).

Pepper fruit also exhibits a wide range of ethylene

production rates and respiratory behavior during

ripening (Villavicencio and others 1999). Although

a role for endogenous ethylene in mediating the

phenotypic changes associated with ripening in

pepper has not been determined, exogenous ethyl-

ene treatments can lead to enhanced carotenoid

accumulation (Fox and others 2005). From the data

generated by studies on apple and melon, it seems

that large differences in fruit ethylene production

appear to be controlled by one or two genetic loci.

Although still unproven, it is possible that a similar

situation may be occurring between closely related

pepper species. QTL analysis on segregating popu-

lations generated from crosses between high and

low ethylene-producing parents should be able to

resolve these issues and address whether differing

ripening mechanisms truly exist within these spe-

cies or (as is more likely) nonclimacteric varieties

represent allelic variants in ethylene synthesis, re-

sponse, or more general ripening genes within what

are normally climacteric species.

ONTROLLING

THYLENE

ESPONSES FOR

ORTICULTURAL

ROP

MPROVEMENT

Clearly, ethylene is required for the ripening of many

fruits and in its absence the ripening process fails to

proceed to completion, rendering the product

unpalatable. However, once initiated, ripening is a

one-way process and the beneﬁcial aspects of ethyl-

ene for generating a high-quality product can soon be

outweighed by its propensity to stimulate over-rip-

ening and decay. This is particularly true under

postharvest storage conditions where considerable

effort is expended to control ethylene effects not only

in fruits but also in vegetables and ornamental crops.

Depending on the commodity, specialized harvest-

ing, packaging, shipping, temperature, and con-

trolled atmospheres may be required, adding to the

cost of production through additional labor and en-

ergy use. Control of ethylene responses is therefore

important for the agricultural and food industries.

Chemical Control of Ethylene Responsiveness

Several compounds have been developed that can

block ethylene action in fruits, vegetables, and ﬂoral

crops and these are thought to act through binding

to the ethylene receptors (Sisler 2006). 1-Methyl-

cyclopropene (1-MCP) is a potent inhibitor of eth-

ylene responses and under the tradenames

EthylBloc



and SmartFreshit has been approved

for commercial use on ornamental and edible hor-

ticultural products, respectively. A whole research

ﬁeld has evolved to test the efﬁcacy and physiologic

effects of 1-MCP on fruit crops and a wide range of

effects are observed that vary between species and

even between cultivars (Watkins 2006). It appears

that this compound does have limitations in many

species, but major successes have been reported for

prolonging the storage life of apples leading to

widespread commercial utilization within the apple

industry.

Genetic Control of Ethylene Synthesis and

Responsiveness

The role of ethylene in regulating fruit ripening in

tomato has been unambiguously determined

through the genetic manipulation of ethylene syn-

thesis and perception (Klee and others 1991; Oeller

and others 1991; Picton and others 1993a, b; Wil-

kinson and others 1997). Transgenic manipulation

of ethylene synthesis through downregulation of

ACS and ACO expression has also been achieved in

melon and apple (Ayub and others 1996; Dandekar

and others 2004). Transgenic melon expressing an

ACO antisense gene displayed inhibition of ripening,

including a reduction in external pigmentation and

fruit softening. The transgenic lines also failed to

develop a peduncular abscission zone and therefore

did not abscise from the plant at maturity, leading to

an enhanced accumulation of sugars (Guis and

others 1997). The production of volatile esters that

represent important ﬂavor components in ripe

melon fruit are also greatly inhibited in the ACO

antisense lines (Bauchot and others 1998; Flores

and others 2002). In addition, fruit of the antisense

lines are less susceptible to chilling injury than those

of wild type (Flores and others 2004). Similarly,

ethylene-suppressed apples were ﬁrmer, had an

extended shelf life, altered sugar and organic acid

proﬁles, and reduced volatile ester formation that

was accompanied by ethylene-dependent reduc-

tions in alcohol acyl-CoA transferase (AAT) activity

(Dandekar and others 2004; Deﬁlippi and others

2004, 2005).

The natural variation seen in ethylene production

and responsiveness in apples, peaches, melons, and

peppers indicate that there is considerable scope for

the generation of new cultivars within these species

with differing ethylene physiology and ripening

characteristics. The development of introgression

lines of tomato harboring deﬁned chromosomal

154 Cornelius S. Barry and others

segments of wild species within the cultivated

Solanum lycopersicum genome is providing unprece-

dented quantities of information about loci that

control fruit quality traits (Gur and others 2004; Liu

and others 2003; Schauer and others 2006). Con-

siderable variation in ethylene synthesis and rip-

ening characteristics of wild species of tomato have

been reported (Grumet and others 1981). It may be

possible to harness the power of the introgression

lines for uncovering QTL that alter ethylene-related

phenotypes during fruit development and ripening.

ONCLUSIONS

From utilizing the wealth of information generated

using the Arabidopsis model and from many funda-

mental studies on fruit crop species, we now have a

basic framework of ethylene biosynthesis and sig-

naling during fruit ripening. This knowledge has

increased our ability to modify ethylene synthesis

and responses in fruit crops to lessen the effects of

postharvest deterioration. However many funda-

mental biological questions remain to be addressed:

How does the transition from system 1 to system

2 modes of ethylene synthesis occur at the onset of

ripening, what is the involvement of the rin,nor,

and Cnr loci in mediating this transition, and how is

the system 2 ethylene synthesis perpetuated?

What are the functions of the individual ethylene

signaling components and how is this network of

proteins assembled in vivo? Is the expansion of gene

families in tomato, and perhaps other ﬂeshy fruit-

bearing species, linked to special ethylene signaling

requirements in these species? What are the

downstream targets of the EILs and ERFs in ripening

fruits and how do these transcription factors activate

or repress speciﬁc ripening-related pathways? How

are tissue-speciﬁc ethylene signaling effects, such as

those mediated by the GR protein and the Al3 and

Al4 loci, achieved?

What role does ethylene play in the ripening of

nonclimacteric fruits? What is the signiﬁcance, if

any, of small transient increases in ethylene pro-

duction in species such as grape and strawberry? In

nonclimacteric fruits, do changes in ethylene sen-

sitivity rather than ethylene synthesis mediate

physiologic changes during ripening? Could non-

climacteric fruits carry mutations within compo-

nents of the ethylene synthesis or signaling

pathways? Within closely related species such as

capsicums and melons, what is the molecular

identity of loci that control climacteric ethylene

production, how widespread is this natural varia-

tion in fruit crop species, and can it be harnessed for

generating new varieties with broad consumer ap-

peal but with enhanced shipping and storage capa-

bility?

Clearly there is still much to discover within the

ﬁeld of ethylene biology as it relates to both the

biological and practical aspects of fruit ripening. The

questions outlined above are complex and will re-

quire multidisciplinary experimental approaches to

resolve but will ultimately provide important

information regarding the mechanism by which this

hormone functions to regulate fruit ripening.

ACKNOWLEDGMENTS

Work highlighted in this review, which was per-

formed by C.B. and others in the Giovannoni

laboratory, was supported by the USDA-NCGRI

(2002-35304-12530), National Science Foundation

(DBI-0501778, DBI- 0605659), and the United

States Department of Agriculture – Agricultural

Research Service.

REFERENCES

Adams-Phillips L, Barry C, Giovannoni J. 2004a. Signal trans-

duction systems regulating fruit ripening. Trends Plant Sci

9:331–338.

Adams-Phillips L, Barry C, Kannan P, Leclercq J, Bouzayen M,

Giovannoni J. 2004b. Evidence that CTR1-mediated ethylene

signal transduction in tomato is encoded by a multigene family

whose members display distinct regulatory features. Plant Mol

Biol 54:387–404.

Aharoni A, Keizer LC, van-den-Broeck HC, Blanco-Portales R,

Munoz-Blanco J, Bois G, Smit P, De-Vos RC, OÕConnell AP.

2002. Novel insight into vascular, stress, and auxin-dependent

and -independent gene expression programs in strawberry, a

non-climacteric fruit. Plant Physiol 129:1019–1031.

Alba R, Payton P, Fei Z, McQuinn R, Debbie P, Martin GB,

Tanksley SD, Giovannoni JJ. 2005. Transcriptome and selected

metabolite analyses reveal multiple points of ethylene control

during tomato fruit development. Plant Cell 17:2954–2965.

Alonso JM, Chamarro J, Granell A. 1995. Evidence for the

involvement of ethylene in the expression of speciﬁc RNAs

during maturation of the orange, a nonclimacteric fruit. Plant

Mol Biol 29:385–390.

Ayub R, Guis M, BenAmor M, Gillot L, Roustan JP, Latche A,

Bouzayen M, Pech JC. 1996. Expression of ACC oxidase anti-

sense gene inhibits ripening of cantaloupe melon fruits. Nat

Biotechnol 14:862–866.

Barry CS, Giovannoni JJ. 2006. Ripening in the tomato Green-

ripe mutant is inhibited by ectopic expression of a protein that

disrupts ethylene signaling. Proc Natl Acad Sci U S A 103:7923–

7928.

Barry CS, Blume B, Bouzayen M, Cooper W, Hamilton AJ,

Grierson D. 1996. Differential expression of the 1-aminocy-

clopropane-1-carboxylate oxidase gene family of tomato. Plant

J 9:525–535.

Barry CS, Llop-Tous MI, Grierson D. 2000. The regulation of

1-aminocyclopropane-1-carboxylic acid synthase gene expres-

sion during the transition from system-1 to system-2 ethylene

synthesis in tomato. Plant Physiol 123:979–986.

Ethylene and Fruit Ripening 155

Barry CS, McQuinn RP, Thompson AJ, Seymour GB, Grierson D,

Giovannoni JJ. 2005. Ethylene insensitivity conferred by the

Green-ripe and Never-ripe 2 ripening mutants of tomato. Plant

Physiol 138:267–275.

Bauchot A, Mottram D, Dodson A, John P. 1998. Effect of am-

inocyclopropane-1-carboxylic acid oxidase antisense gene on

the formation of volatile esters in Cantaloupe Charentais

melon (cv. Vedrantais). J Agric Food Chem 46:4787–4792.

Bleecker AB, Kende H. 2000. Ethylene: A gaseous signal mole-

cule in plants. Annu Rev Cell Dev Biol 16:1–18.

Bleecker AB, Estelle MA, Somerville C, Kende H. 1988. Insensi-

tivity to ethylene conferred by a dominant mutation in Ara-

bidopsis thaliana. Science 241:1086–1089.

Chang C, Kwok SF, Bleecker AB, Meyerowitz EM. 1993. Ara-

bidopsis ethylene-response gene ETR1: Similarity of product to

two-component regulators. Science 262:539–544.

Chao Q, Rothenberg M, Solano R, Roman G, Terzaghi W, Ecker

J-R. 1997. Activation of the ethylene gas response pathway in

Arabidopsis by the nuclear protein ETHYLENE-INSENSITIVE3

and related proteins. Cell 89:1133–1144.

Chen G, Alexander L, Grierson D. 2004a. Constitutive expression

of EIL-like transcription factor partially restores ripening in the

ethylene-insensitive Nr tomato mutant. J Exp Bot 55:1491–

1497.

Chen GP, Hackett R, Walker D, Taylor A, Lin ZF, Grierson D.

2004b. Identiﬁcation of a speciﬁc isoform of tomato lipoxy-

genase (TomloxC) involved in the generation of fatty acid-

derived ﬂavor compounds. Plant Physiol 136:2641–2651.

Chervin C, El-Kereamy A, Roustan JP, Latche A, Lamon J,

Bouzayen M. 2004. Ethylene seems required for the berry

development and ripening in grape, a non-climacteric fruit.

Plant Sci 167:1301–1305.

Ciardi JA, Tieman DM, Jones JB, Klee HJ. 2001. Reduced

expression of the tomato ethylene receptor gene LeETR4 en-

hances the hypersensitive response to Xanthomonas campestris

pv. vesicatoria. Mol Plant Microbe Interact 14:487–495.

Clark KL, Larsen PB, Wang X, Chang C. 1998. Association of the

Arabidopsis CTR1 Raf-like kinase with the ETR1 and ERS

ethylene receptors. Proc Natl Acad Sci U S A 95:5401–5406.

Dandekar AM, Teo G, Deﬁlippi BG, Uratsu SL, Passey AJ, Kader

AA, Stow JR, Colgan RJ, James DJ. 2004. Effect of down-reg-

ulation of ethylene biosynthesis on fruit ﬂavor complex in

apple fruit. Transgenic Res 13:373–384.

Deﬁlippi BG, Dandekar AM, Kader AA. 2004. Impact of sup-

pression of ethylene action or biosynthesis on ﬂavor metabo-

lites in apple (Malus domestica Borkh) fruits. J Agric Food Chem

52:5694–5701.

Deﬁlippi BG, Kader AA, Dandekar AM. 2005. Apple aroma:

alcohol acyltransferase, a rate limiting step for ester biosyn-

thesis, is regulated by ethylene. Plant Sci 168:1199–1210.

Dopico B, Lowe AL, Wilson ID, Merodio C, Grierson D. 1993.

Cloning and characterization of avocado fruit mRNAs and their

expression during ripening and low-temperature storage. Plant

Mol Biol 21:437–449.

El-Kereamy A, Chervin C, Roustan JP, Cheynier V, Souquet JM,

Moutounet M, Raynal J, Ford C, Latche A, Pech JC, Bouzayen

M. 2003. Exogenous ethylene stimulates the long-term

expression of genes related to anthocyanin biosynthesis in

grape berries. Physiol Plant 119:175–182.

Fan XT, Mattheis JP, Fellman JK. 1998. A role for jasmonates in

climacteric fruit ripening. Planta 204:444–449.

Fei ZJ, Tang X, Alba RM, White JA, Ronning CM, Martin GB,

Tanksley SD, Giovannoni JJ. 2004. Comprehensive EST anal-

ysis of tomato and comparative genomics of fruit ripening.

Plant J 40:47–59.

Fei ZJ, Tang XM, Alba R, Giovannoni J. 2006. Tomato Expression

Database (TED): a suite of data presentation and analysis tools.

Nucleic Acids Res 34:D766–D770.

Flores F, El Yahyaoui F, Billerbeck G de , Romojaro F, Latche A,

Bouzayen M, Pech JC, Ambid C. 2002. Role of ethylene in the

biosynthetic pathway of aliphatic ester aroma volatiles in

Charentais Cantaloupe melons. J Exp Bot 53:201–206.

Flores FB, Martinez-Madrid M, Ben Amor M, Pech JC, Latche A,

Romojaro F. 2004. Modiﬁed atmosphere packaging confers

additional chilling tolerance on ethylene-inhibited cantaloupe

Charentais melon fruit. European Food Res Technol 219:614–

619.

Fox AJ, Pozo-Insfran D Del , Lee JH, Sargent SA, Talcott ST. 2005.

Ripening-induced chemical and antioxidant changes in bell

peppers as affected by harvest maturity and postharvest eth-

ylene exposure. Hortscience 40:732–736.

Fridman E, Carrari F, Liu YS, Fernie AR, Zamir D. 2004. Zooming

in on a quantitative trait for tomato yield using interspeciﬁc

introgressions. Science 305:1786–1789.

Frye CA, Tang DZ, Innes RW. 2001. Negative regulation of de-

fense responses in plants by a conserved MAPKK kinase. Proc

Natl Acad Sci U S A 98:373–378.

Gagne J-M, Smalle J, Gingerich D-J, Walker J-M, Yoo S-D,

Yanagisawa S, Vierstra R-D. 2004. Arabidopsis EIN3-binding F-

box 1 and 2 form ubiquitin-protein ligases that repress ethyl-

ene action and promote growth by directing EIN3 degradation.

Proc Natl Acad Sci U S A 101:6803–6808.

Gao Z, Chen Y-F, Randlett MD, Zhao X-C, Findell JL, Kieber JJ,

Schaller GE. 2003. Localization of the Raf-like kinase CTR1 to

the endoplasmic reticulum of Arabidopsis through participa-

tion in ethylene receptor signaling complexes. J Biol Chem

278:34725–34732.

Giovannoni JJ. 2004. Genetic regulation of fruit development

and ripening. Plant Cell 16:S170–S180.

Given NK, Venis MA, Grierson D. 1988. Hormonal regulation of

ripening in the strawberry a non-climacteric fruit. Planta

174:402–406.

Goldschmidt EE, Huberman M, Goren R. 1993. Probing the role

of endogenous ethylene in the degreening of citrus-fruit with

ethylene antagonists. J Plant Growth Regul 12:325–329.

Grifﬁths A, Barry C, Alpuche-Solis AG, Grierson D. 1999. Eth-

ylene and developmental signals regulate expression of lipox-

ygenase genes during tomato fruit ripening. J Exp Bot 50:793–

798.

Grumet R, Fobes JF, Herner RC. 1981. Ripening behavior of wild

tomato species. Plant Physiol 68:1428–1432.

Guis M, Botondi R, BenAmor M, Ayub R, Bouzayen M, Pech

JC, Latche A. 1997. Ripening-associated biochemical traits

of Cantaloupe Charentais melons expressing an antisense

ACC oxidase transgene. J Am Soc Horticult Sci 122:748–

751.

Guo H, Ecker JR. 2003. Plant responses to ethylene gas are

mediated by SCFEBF1/EBF2-dependent proteolysis of EIN3

transcription factor. Cell 115:667–677.

Guo H, Ecker JR. 2004. The ethylene signaling pathway: New

insights. Curr Opin Plant Biol 7:40–49.

Gur A, Semel Y, Cahaner A, Zamir D. 2004. Real time QTL of

complex phenotypes in tomato interspeciﬁc introgression lines.

Trends Plant Sci 9:107–109.

Gussman CD, Goffreda JC, Gianfagna TJ. 1993. Ethylene pro-

duction and fruit-softening rates in several apple fruit ripening

variants. Hortscience 28:135–137.

Gutterson N, Reuber TL. 2004. Regulation of disease resistance

pathways by AP2/ERF transcription factors. Curr Opin Plant

Biol 7:465–471.

156 Cornelius S. Barry and others

Guzman P, Ecker JR. 1990. Exploiting the triple response of

Arabidopsis to identify ethylene-Related mutants. Plant Cell

2:513–524.

Hackett RM, Ho C-W, Lin Z, Foote HCC, Fray RG, Grierson D. 2000.

Antisense inhibition of the Nr gene restores normal ripening to

the tomato Never-ripe mutant, consistent with the ethylene

receptor-inhibition model. Plant Physiol 124:1079–1085.

Haji T, Yaegaki H, Yamaguchi M. 2001. Changes in ethylene

production and ﬂesh ﬁrmness of melting, nonmelting and

stony hard peaches after harvest. J Jpn Soc Horticult Sci

70:458–459.

Haji T, Yaegaki H, Yamaguchi M. 2003. Softening of stony hard

peach by ethylene and the induction of endogenous ethylene

by 1-aminocyclopropane-1-carboxylic acid (ACC). J Jpn Soc

Horticult Sci 72:212–217.

Hall AE, Chen QG, Findell JL, Schaller GE, Bleecker AB. 1999.

The relationship between ethylene binding and dominant

insensitivity conferred by mutant forms of the ETR1 ethylene

receptor. Plant Physiol 121:291–299.

Harada T, Sunako T, Wakasa Y, Soejima J, Satoh T, Niizeki M.

2000. An allele of the 1-aminocyclopropane-1-carboxylate

synthase gene (Md-ACS1) accounts for the low level of eth-

ylene production in climacteric fruits of some apple cultivars.

Theoret Appl Genet 101:742–746.

Hirschberg J. 2001. Carotenoid biosynthesis in ﬂowering plants.

Curr Opin Plant Biol 4:210–218.

Hobson GE, Nichols R, Davies JN, Atkey PT. 1984. The inhibition

of tomato lycopersicon-esculentum fruit ripening by silver. J

Plant Physiol 116:21–30.

Hua J, Meyerowitz EM. 1998. Ethylene responses are negatively

regulated by a receptor gene family in Arabidopsis thaliana. Cell

94:261–271.

Iannetta PPM, Laarhoven LJ, Medina-Escobar N, James EK,

McManus MT, Davies HV, Harren FJM. 2006. Ethylene and

carbon dioxide production by developing strawberries show a

correlative pattern that is indicative of ripening climacteric

fruit. Physiol Plant 127:247–259.

Jacob-Wilk D, Holland D, Goldschmidt EE, Riov J, Eyal Y. 1999.

Chlorophyll breakdown by chlorophyllase: isolation and

functional expression of the Chlase1 gene from ethylene-

treated citrus fruit and its regulation during development. Plant

J 20:653–661.

Jarret RL, Tigchelaar EC, Handa AK. 1984. Ripening behavior of

the green-ripe tomato lycopersicon-esculentum mutant. J Am

Soc Horticult Sci 109:712–717.

Katz E, Lagunes PM, Riov J, Weiss D, Goldschmidt EE. 2004.

Molecular and physiological evidence suggests the existence of

a system II-like pathway of ethylene production in non-cli-

macteric Citrus fruit. Planta 219:243–252.

Kerr EA. 1958. Mutations of chlorophyll retention in ripe fruit.

Rep Tomato Genet Coop 8:22 .

Kerr EA. 1982. Never ripe-2 (Nr-2) a slow ripening mutant

resembling Nr and Gr. Rep Tomato Genet Coop 32:33 .

Kieber JJ, Rothenberg M, Roman G, Feldmann KA, Ecker JR.

1993. CTR1, a negative regulator of the ethylene response

pathway in Arabidopsis, encodes a member of the Raf family of

protein kinases. Cell 72:427–441.

Klee HJ. 2004. Ethylene signal transduction. Moving beyond

Arabidopsis. Plant Physiol 135:660–667.

Klee HJ, Hayford MB, Kretzmer KA, Barry GF, Kishore GM.

1991. Control of ethylene synthesis by expression of a bacterial

enzyme in transgenic tomato plants. Plant Cell 3:1187–1194.

Lanahan MB, Yen H-C, Giovannoni JJ, Klee HJ. 1994. The never

ripe mutation blocks ethylene perception in tomato. Plant Cell

6:521–530.

Lashbrook CC, Tieman DM, Klee HJ. 1998. Differential regulation

of the tomato ETR gene family throughout plant development.

Plant J 15:243–252.

Leclercq J, Adams-Phillips LC, Zegzouti H, Jones B, Latche A,

Giovannoni JJ, Pech J-C, Bouzayen M. 2002. LeCTR1, a to-

mato CTR1-like gene, demonstrates ethylene signaling ability

in Arabidopsis and novel expression patterns in tomato. Plant

Physiol 130:1132–1142.

Lelievre JM, Latche A, Jones B, Bouzayen M, Pech JC. 1997.

Ethylene and fruit ripening. Physiol Plant 101:727–739.

Lin Z, Hackett RM, Payton S, Grierson D. 1998. A tomato se-

quence, TCTR2 (accession no. AJ005077), encoding an Ara-

bidopsis CTR1 homologue. Plant Physiol 117:1126 .

Lincoln JE, Fischer RL. 1988. Regulation of gene-expression by

ethylene in wild-type and rin tomato (lycopersicon-esculen-

tum) fruit. Plant Physiol 88:370–374.

Lincoln JE, Cordes S, Read E, Fischer RL. 1987. Regulation of

gene expression by ethylene during lycopersicon-esculentum

tomato fruit development. Proc Natl Acad Sci U S A 84:2793–

2797.

Litt A, Irish VF. 2003. Duplication and diversiﬁcation in the A-

PETALA1/FRUITFULL ﬂoral homeotic gene lineage: Implica-

tions for the evolution of ﬂoral development. Genetics

165:821–833.

Liu Y, Schiff M, Dinesh-Kumar S-P. 2002. Virus-induced gene

silencing in tomato. Plant J 31:777–786.

Liu YS, Gur A, Ronen G, Causse M, Damidaux R, Buret M, Hir-

schberg J, Zamir D. 2003. There is more to tomato fruit colour

than candidate carotenoid genes. Plant Biotechnol J 1:195–

207.

Malcomber ST, Kellogg EA. 2005. SEPALLATA gene diversiﬁca-

tion: brave new whorls. Trends Plant Sci 10:427–435.

Manning K, Tor M, Poole M, Hong Y, Thompson AJ, King GJ,

Giovannoni JJ, Seymour GB. 2006. A naturally occurring

epigenetic mutation in a gene encoding an SBP-box tran-

scription factor inhibits tomato fruit ripening. Nat Genet

38:948–952.

Marty I, Douat C, Tichit L, Jungsup K, Leustek T, Albagnac G.

2000. The cystathionine-gamma-synthase gene involved in

methionine biosynthesis is highly expressed and auxin-re-

pressed during wild strawberry (Fragaria vesca L.) fruit ripening.

Theoret Appl Genet 100:1129–1136.

Matarasso N, Schuster S, Avni A. 2005. A novel plant cysteine

protease has a dual function as a regulator of 1-aminocyclo-

propane-1-carboxylic acid synthase gene expression. Plant Cell

17:1205–1216.

McMurchie EJ, McGlasson WB, Eaks IL. 1972. Treatment of fruit

with propylene gives information about biogenesis of ethylene.

Nature 237:235–236.

Moeder W, Barry CS, Tauriainen AA, Betz C, Tuomainen J,

Utriainen M, Grierson D, Sandermann H, Langebartels C,

Kangasjarvi J. 2002. Ethylene synthesis regulated by biphasic

induction of 1-aminocyclopropane-1-carboxylic acid synthase

and 1-aminocyclopropane-1-carboxylic acid oxidase genes is

required for hydrogen peroxide accumulation and cell death in

ozone-exposed tomato. Plant Physiol 130:1918–1926.

Mueller LA, Tanksley SD, Giovannoni JJ, Eck J van , Stack S,

Choi D, Kim BD, Chen MS, Cheng ZK, Li CY, Ling HQ, Xue YB,

Seymour G, Bishop G, Bryan G, Sharma R, Khurana J, Tyagi A,

Chattopadhyay D, Singh NK, Stiekema W, Lindhout P, Jesse T,

Lankhorst RK, Bouzayen M, Shibata D, Tabata S, Granell A,

Botella MA, Giullano G, Frusciante L, Causse M, Zamir D.

2005. The Tomato Sequencing Project, the ﬁrst cornerstone of

the International Solanaceae Project (SOL). Comp Funct Ge-

nomics 6:153–158.

Ethylene and Fruit Ripening 157

Nakano R, Ogura E, Kubo Y, Inaba A. 2003. Ethylene biosyn-

thesis in detached young persimmon fruit is initiated in calyx

and modulated by water loss from the fruit. Plant Physiol

131:276–286.

Nakatsuka A, Murachi S, Okunishi H, Shiomi S, Nakano R,

Kubo Y, Inaba A. 1998. Differential expression and internal

feedback regulation of 1-aminocyclopropane-1-carboxylate

synthase, 1-aminocyclopropane-1-carboxylate oxidase, and

ethylene receptor genes in tomato fruit during development

and ripening. Plant Physiol 118:1295–1305.

OÕMalley RC, Rodriguez FI, Esch JJ, Binder BM, OÕDonnell P,

Klee HJ, Bleecker AB. 2005. Ethylene-binding activity, gene

expression levels, and receptor system output for ethylene

receptor family members from Arabidopsis and tomato. Plant J

41:651–659.

Oeller PW, Min Wong L, Taylor LP, Pike DA, Theologis A. 1991.

Reversible inhibition of tomato fruit senescence by antisense

RNA. Science 254:437–439.

Oetiker JH, Olson DC, Shiu O-Y, Yang SF. 1997. Differential

induction of seven 1-aminocyclopropane-1-carboxylate syn-

thase genes by elicitor in suspension cultures of tomato

(Lycopersicon esculentum). Plant Mol Biol 34:275–286.

Olmedo G, Guo HW, Gregory BD, Nourizadeh SD, Aguilar-He-

nonin L, Li HJ, An FY, Guzman P, Ecker JR. 2006. ETHYLENE-

INSENSITIVE5 encodes a 5¢ﬁ3¢exoribonuclease required for

regulation of the EIN3-targeting F-box proteins EBF1/2. Proc

Natl Acad Sci U S A 103:13286–13293.

Oraguzie NC, Iwanami H, Soejima J, Harada T, Hall A. 2004.

Inheritance of the Md-ACS1 gene and its relationship to fruit

softening in apple (Malus x domestica Borkh.). Theoret Appl

Genet 108:1526–1533.

Payton S, Fray RG, Brown S, Grierson D. 1996. Ethylene receptor

expression is regulated during fruit ripening, ﬂower senescence

and abscission. Plant Mol Biol 31:1227–1231.

Pedraza-Lopez A, Blanco-Portales R, Lopez-Raez J, Medina-

Escobar N, Munoz-Blanco J, Rodriguez A. 2006. Character-

ization of a strawberry late-expressed and fruit-speciﬁc peptide

methionine sulphoxide reductase. Physiol Plant 126:129–139.

Perin C, Gomez-Jimenez M, Hagen L, Dogimont C, Pech J-C,

Latche A, Pitrat M, Lelievre J-M. 2002. Molecular and genetic

characterization of a non-climacteric phenotype in melon re-

veals two loci conferring altered ethylene response in fruit.

Plant Physiol 129:300–309.

Picton S, Barton SL, Bouzayen M, Hamilton AJ, Grierson D.

1993a. Altered fruit ripening and leaf senescence in tomatoes

expressing an antisense ethylene-forming enzyme transgene.

Plant J 3:469–481.

Picton S, Gray J, Barton S, Abubakar U, Lowe A, Grierson D.

1993b. CDNA cloning and characterisation of novel ripening-

related mRNAs with altered patterns of accumulation in the

ripening inhibitor (rin) tomato ripening mutant. Plant Mol Biol

23:193–207.

Potuschak T, Lechner E, Parmentier Y, Yanagisawa S, Grava S,

Koncz C, Genschik P. 2003. EIN3-dependent regulation of

plant ethylene hormone signaling by two Arabidopsis F box

proteins: EBF1 and EBF2. Cell 115:679–689.

Purvis AC, Barmore CR. 1981. Involvement of ethylene in

chlorophyll degradation in peel of citrus-fruits. Plant Physiol

68:854–856.

Resnick JS, Wen C-K, Shockey JA, Chang C. 2006. REVERSION-

TO-ETHYLENE SENSITIVITY1, a conserved gene that regulates

ethylene receptor function in Arabidopsis. Proc Natl Acad Sci

USA 103:7917–7922.

Rick CM. 1956. New mutants. Rep Tomato Genet Coop 6:22–23.

Rose JKC, Bennett AB. 1999. Cooperative disassembly of the

cellulose-xyloglucan network of plant cell walls: parallels be-

tween cell expansion and fruit ripening. Trends Plant Sci

4:176–183.

Rottmann WH, Peter GF, Oeller PW, Keller JA, Shen NF, Nagy

BP, Taylor LP, Campbell AD, Theologis A. 1991. 1 Aminocy-

clopropane-1-carboxylate synthase in tomato is encoded by a

multigene family whose transcription is induced during fruit

and ﬂoral senescence. J Mol Biol 222:937–962.

Sato T, Kudo T, Akada T, Wakasa Y, Niizeki M, Harada T. 2004.

Allelotype of a ripening-speciﬁc 1-aminocyclopropane-1-car-

boxylate synthase gene deﬁnes the rate of fruit drop in apple. J

Am Soc Horticult Sci 129:32–36.

Schauer N, Semel Y, Roessner U, Gur A, Balbo I, Carrari F, Pleban

T, Perez-Melis A, Bruedigam C, Kopka J, Willmitzer L, Zamir D,

Fernie AR. 2006. Comprehensive metabolic proﬁling and

phenotyping of interspeciﬁc introgression lines for tomato

improvement. Nat Biotechnol 24:447–454.

Sisler EC. 2006. The discovery and development of compounds

counteracting ethylene at the receptor level. Biotechnol Adv

24:357–367.

Slater A, Maunders MJ, Edwards K, Schuch W, Grierson D. 1985.

Isolation and characterization of complementary DNA clones

for tomato Lycopersicon-esculentum cultivar Ailsa-Craig po-

lygalacturonase and other ripening-related proteins. Plant Mol

Biol 5:137–148.

Solano R, Stepanova A, Chao Q, Ecker JR. 1998. Nuclear events

in ethylene signaling: A transcriptional cascade mediated by

ethylene-insensitive3 and ethylene-response-factor1. Genes

Dev 2:3703–3714.

Stewart I, Wheaton TA. 1972. Carotenoids in citrus—their

accumulation induced by ethylene. J Agric Food Chem 20:448–

449.

Sunako R, Sakuraba W, Senda M, Akada S, Ishikawa R, Niizeki

M, Harada T. 1999. An allele of the ripening-speciﬁc 1-am-

inocyclopropane-1-carboxylic acid synthase gene (ACS1) in

apple fruit with a long storage life. Plant Physiol 119:1297–

1303.

Symons GM, Davies C, Shavrukov Y, Dry IB, Reid JB, Thomas

MR. 2006. Grapes on steroids. Brassinosteroids are involved in

grape berry ripening. Plant Physiol 140:150–158.

Tang DZ, Christiansen KM, Innes RW. 2005. Regulation of plant

disease resistance, stress responses, cell death, and ethylene

signaling in Arabidopsis by the EDR1 protein kinase. Plant

Physiol 138:1018–1026.

Tatsuki M, Haji T, Yamaguchi M. 2006. The involvement of 1-

aminocyclopropane-1-carboxylic acid synthase isogene, Pp-

ACS1, in peach fruit softening. J Exp Bot 57:1281–1289.

Tesniere C, Pradal M, El-Kereamy A, Torregrosa L, Chatelet P,

Roustan JP, Chervin C. 2004. Involvement of ethylene sig-

nalling in a non-climacteric fruit: new elements regarding the

regulation of ADH expression in grapevine. J Exp Bot 55:2235–

2240.

Thompson AJ, Tor M, Barry CS, Vrebalov J, Orﬁla C, Jarvis MC,

Giovannoni JJ, Grierson D, Seymour GB. 1999. Molecular and

genetic characterization of a novel pleiotropic tomato-ripening

mutant. Plant Physiol 120:383–389.

Tieman DM, Klee HJ. 1999. Differential expression of two novel

members of the tomato ethylene-receptor family. Plant Physiol

120:165–172.

Tieman DM, Taylor MG, Ciardi JA, Klee HJ. 2000. The tomato

ethylene receptors NR and LeETR4 are negative regulators of

ethylene response and exhibit functional compensation within

a multigene family. Proc Natl Acad Sci U S A 97:5663–5668.

158 Cornelius S. Barry and others

Tieman DM, Ciardi JA, Taylor MG, Klee HJ. 2001. Members of

the tomato LeEIL (EIN3-like) gene family are functionally

redundant and regulate ethylene responses throughout plant

development. Plant J 26:47–58.

Tieman D, Taylor M, Schauer N, Fernie AR, Hanson AD, Klee HJ.

2006. Tomato aromatic amino acid decarboxylases participate

in synthesis of the ﬂavor volatiles 2-phenylethanol and 2-

phenylacetaldehyde. Proc Natl Acad Sci U S A 103:8287–8292.

Tigchelaar EC, McGlasson WB, Buescher RW. 1978. Genetic

regulation of tomato fruit ripening. HortScience 13:508–513.

Tournier B, Sanchez-Ballesta M-T, Jones B, Pesquet E, Regad F,

Latche A, Pech J-C, Bouzayen M. 2003. New members of the

tomato ERF family show speciﬁc expression pattern and di-

verse DNA-binding capacity to the GCC box element. FEBS

Lett 550:149–154.

Trainotti L, Pavanello A, Casadoro G. 2005. Different ethylene

receptors show an increased expression during the ripening of

strawberries: does such an increment imply a role for ethylene

in the ripening of these non-climacteric fruits? J Exp Bot

56:2037–2046.

Van-der-Hoeven R, Ronning C, Giovannoni J, Martin G, Tanks-

ley S. 2002. Deductions about the number, organization, and

evolution of genes in the tomato genome based on analysis of a

large expressed sequence tag collection and selective genomic

sequencing. Plant Cell 14:1441–1456.

Vardhini BV, Rao SSR. 2002. Acceleration of ripening of tomato

pericarp discs by brassinosteroids. Phytochemistry 61:843–847.

Villavicencio L, Blankenship SM, Sanders DC, Swallow WH.

1999. Ethylene and carbon dioxide production in detached

fruit of selected pepper cultivars. J Am Soc Horticult Sci

124:402–406.

Vrebalov J, Ruezinsky D, Padmanabhan V, White R, Medrano D,

Drake R, Schuch W, Giovannoni J. 2002. A MADS-box gene

necessary for fruit ripening at the tomato ripening-inhibitor

(rin) locus. Science 296:343–346.

Watkins CB. 2006. The use of 1-methylcyclopropene (1-MCP) on

fruits and vegetables. Biotechnol Adv 24:389–409.

Whitelaw CA, Lyssenko NN, Chen L, Zhou D, Mattoo AK, Tucker

ML. 2002. Delayed abscission and shorter internodes correlate

with a reduction in the ethylene receptor LeETR1 transcript in

transgenic tomato. Plant Physiol 128:978–987.

Wilkinson JQ, Lanahan MB, Clark DG, Bleecker AB, Chang C,

Meyerowitz EM, Klee HJ. 1997. A dominant mutant receptor

from Arabidopsis confers ethylene insensitivity in heterologous

plants. Nat Biotechnol 15:444–447.

Wilkinson JQ, Lanahan MB, Yen H-C, Giovannoni JJ, Klee HJ.

1995. An ethylene-inducible component of signal transduction

encoded by Never-ripe. Science 270:1807–1809.

Yanagisawa S, Yoo S-D, Sheen J. 2003. Differential regulation of

EIN3 stability by glucose and ethylene signalling in plants.

Nature 425:521–525.

Yang SF, Hoffman NE. 1984. Ethylene biosynthesis and its reg-

ulation in higher plants. Annu Rev Plant Physiol 35:155–190.

Yen H-C, Lee S, Tanksley SD, Lanahan MB, Klee HJ, Giovannoni

JJ. 1995. The tomato Never-ripe locus regulates ethylene-

inducible gene expression and is linked to a homolog of the

Arabidopsis ETR1 gene. Plant Physiol 107:1343–1353.

Yokotani N, Tamura S, Nakano R, Inaba A, Kubo Y. 2003.

Characterization of a novel tomato EIN3-like gene (LeEIL4). J

Exp Bot 54:2775–2776.

Yokotani N, Tamura S, Nakano R, Inaba A, McGlasson WB, Kubo

Y. 2004. Comparison of ethylene- and wound-induced re-

sponses in fruit of wild-type, rin and nor tomatoes. Postharvest

Biol Technol 32:247–252.

Zarembinski TI, Theologis A. 1994. Ethylene biosynthesis and

action: A case of conservation. Plant Mol Biol 26:1579–1597.

Zegzouti H, Jones B, Frasse P, Marty C, Maitre B, Latche A, Pech

J-C, Bouzayen M. 1999. Ethylene-regulated gene expression in

tomato fruit: Characterization of novel ethylene-responsive

and ripening-related genes isolated by differential display. Plant

J 18:589–600.

Zheng XY, Wolff DW. 2000. Ethylene production, shelf-life and

evidence of RFLP polymorphisms linked to ethylene genes in

melon (Cucumis melo L.). Theoret Appl Genet 101:613–624.

Zhou D, Kalaitzis P, Mattoo AK, Tucker ML. 1996. The mRNA for

an ETR1 homologue in tomato is constitutively expressed in

vegetative and reproductive tissues. Plant Mol Biol 30:1331–

1338.

Ethylene and Fruit Ripening 159

The Physical and Structural Effects of 1-MCP on Four Different Apple Cultivars during Storage

Article

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The impact of the ethylene inhibitor, 1-methylcyclopropene (1-MCP), on four apple cultivars (Braeburn, Fuji, Jazz and Golden Delicious) over 150 days of storage at 2 °C was assessed. Proton transfer reaction quadrupole mass spectrometry (PTR-QUAD-MS) was used to monitor changes in VOC composition, while texture analysis and X-ray microcomputer tomography (µ-CT) scanning were used to study microstructural changes. The application of 1-MCP on apples reduced VOC emissions, concurrently maintaining a firmer texture compared to the untreated apples at each time point. The µ-CT scanning revealed how changes in specific morphological characteristics such as anisotropy, connectivity and porosity, size and shape, as well as the interconnectivity of intracellular spaces (IS) influenced texture even when porosity was similar. Additionally, this study showed that the porosity and connectivity of IS were associated with VOC emission and increased simultaneously. This study highlights how the morphological parameters of an apple can help explain their ripening process during long-term storage and how their microstructure can influence the release of VOCs.

Genome-wide identification and expression pattern analysis of the ACS gene family during fruit development in peach

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Jan 2023

Solving the puzzle of climacteric fruit ripening: EMB1444-like and its regulatory function

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J EXP BOT

Francesca Bellinazzo

This article comments on: Zhao W, Wang S, Li W, Shan X, Naeem M, Zhang L, Zhao L. 2023. The transcription factor EMB1444-like affects tomato fruit ripening by regulating YELLOW-FRUITED TOMATO 1, a core component of ethylene signaling transduction. Journal of Experimental Botany 74, 6563–6574.

Impact of Climate Change on Altered Fruit Quality with Organoleptic, Health Benefit, and Nutritional Attributes

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J AGR FOOD CHEM

Tattoo-like Flexible Ethylene Sensor for Plant Stress Monitoring in Real-time

Article

Full-text available

Oct 2023
IEEE SENS J

This paper presents a flexible tattoo sensor for real-time monitoring of gaseous ethylene released from plant leaves. To the best of our knowledge, this is the first reported plant tattoo sensor that can measure and monitor a plant hormone in situ. The tattoo sensor can be easily installed on leaves and removed when required using a stick-and-peel mechanism. Furthermore, temperature and humidity sensors were incorporated to correct the ethylene measurements based on the specific microclimate of the leaf. All sensors were chemiresistive in nature, causing variations in resistance as a result of changes in temperature, humidity, or ethylene. The ethylene sensor demonstrated a sensitivity of 0.6295 kΩ/ppm and a limit of detection of 0.13 ppm. The measurements were processed by an onboard data logger, transmitted wirelessly to the cloud, and accessed on a smartphone. Plants were subjected to multiple stressors, including water, temperature, and dark stressors, and the real-time ethylene measurements were recorded with the tattoo sensor. A statistically significant difference, characterized by p values less than 0.05, was observed between different types of stresses. Roll-to-roll fabrication of these tattoos will enable their large-scale implementation in greenhouses and agricultural fields. Index Terms-abiotic stress, electrochemical sensor, ethylene, plant hormone, plant wearables, tattoo sensor.

The First Signal to Initiate Fruit Ripening is Generated in the Cuticle: An Hypothesis

Chapter

Full-text available

Oct 2023

The paradigm that has prevailed for a long time sustains that ethylene is the first signal that initiates fruit ripening. However, in this manuscript, we present the hypothesis that a signal generated from the cuticle induces the synthesis of ethylene, and therefore, it is the initial signal that triggers the fruit-ripening phenomena. Among the experimental evidence supporting the hypothesis, we can mention that cuticle components released during the plant pathogenic attack can induce the synthesis of ethylene in plants. Also, it has been found that in fungi, a cuticle component can activate a transcription factor by phosphorylation, which induces the transcription of a gene encoding a cutinase. Besides, studies with plant tissues experiencing a high rate of cell expansion have shown that there is a careful synchronization between the demand of cuticle components and biosynthesis, which suggests that the plant cell can sense the moment in which the fruit would stop growing by cell expansion, and initiate the ripening phenomena. In this chapter, experimental evidences supporting the physiological role of the fruit cuticle in the fruit ripening phenomena will be presented and reviewed with the goal to show a possible role of the fruit cuticle in the onset of fruit ripening.

Fruit Development: Growth, Maturation and Ripening

Chapter

Nov 2023

Genome-Wide Analysis of the Polygalacturonase Gene Family Sheds Light on the Characteristics, Evolutionary History, and Putative Function of Akebia trifoliata

Article

Full-text available

Nov 2023
INT J MOL SCI

Polygalacturonase (PG) is one of the largest families of hydrolytic enzymes in plants. It is involved in the breakdown of pectin in the plant cell wall and even contributes to peel cracks. Here, we characterize PGs and outline their expression profiles using the available reference genome and transcriptome of Akebia trifoliata. The average length and exon number of the 47 identified AktPGs, unevenly assigned on 14 chromosomes and two unassembled contigs, were 5399 bp and 7, respectively. The phylogenetic tree of 191 PGs, including 47, 57, 51, and 36 from A. trifoliata, Durio zibethinus, Actinidia chinensis, and Vitis vinifera, respectively, showed that AktPGs were distributed in all groups except group G and that 10 AktPGs in group E were older, while the remaining 37 AktPGs were younger. Evolutionarily, all AktPGs generally experienced whole-genome duplication (WGD)/segmental repeats and purifying selection. Additionally, the origin of conserved domain III was possibly associated with a histidine residue (H) substitute in motif 8. The results of both the phylogenetic tree and expression profiling indicated that five AktPGs, especially AktPG25, could be associated with the cracking process. Detailed information and data on the PG family are beneficial for further study of the postharvest biology of A. trifoliata.

Alleviation of Plant Stress by Molecular Hydrogen

Chapter

Nov 2023

Gasotransmitters and gaseous-signaling molecules are hugely important for controlling cell function and especially so during stress challenges. Past research has concentrated on molecules such as nitric oxide (NO) and hydrogen sulfide (H2S), although others such as ethylene and carbon monoxide (CO) are also important. Here, molecular hydrogen (H2) is added to the mix. H2 has been shown to ameliorate responses to a range of stressors in plants, including exposure to heavy metals, salinity, extreme temperatures, and UV radiation. Clearly, H2 is an important gas, which may be useful for enhancing plant growth and food security in the future. Exogenous treatments with H2 are easy in the form of hydrogen-rich water (HRW), but there are still issues with its wide-spread use. Furthermore, the molecular basis of the action of H2 in cells is still not clear. Here, aspects of the use and the action of H2 in plants are discussed, along with what might be learnt from other species.

Deciphering the mechanisms, hormonal signaling, and potential applications of endophytic microbes to mediate stress tolerance in medicinal plants

Article

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Nov 2023

The global healthcare market in the post-pandemic era emphasizes a constant pursuit of therapeutic, adaptogenic, and immune booster drugs. Medicinal plants are the only natural resource to meet this by supplying an array of bioactive secondary metabolites in an economic, greener and sustainable manner. Driven by the thrust in demand for natural immunity imparting nutraceutical and life-saving plant-derived drugs, the acreage for commercial cultivation of medicinal plants has dramatically increased in recent years. Limited resources of land and water, low productivity, poor soil fertility coupled with climate change, and biotic (bacteria, fungi, insects, viruses, nematodes) and abiotic (temperature, drought, salinity, waterlogging, and metal toxicity) stress necessitate medicinal plant productivity enhancement through sustainable strategies. Plants evolved intricate physiological (membrane integrity, organelle structural changes, osmotic adjustments, cell and tissue survival, reclamation, increased root-shoot ratio, antibiosis, hypersensitivity, etc.), biochemical (phytohormones synthesis, proline, protein levels, antioxidant enzymes accumulation, ion exclusion, generation of heat-shock proteins, synthesis of allelochemicals. etc.), and cellular (sensing of stress signals, signaling pathways, modulating expression of stress-responsive genes and proteins, etc.) mechanisms to combat stresses. Endophytes, colonizing in different plant tissues, synthesize novel bioactive compounds that medicinal plants can harness to mitigate environmental cues, thus making the agroecosystems self-sufficient toward green and sustainable approaches. Medicinal plants with a host set of metabolites and endophytes with another set of secondary metabolites interact in a highly complex manner involving adaptive mechanisms, including appropriate cellular responses triggered by stimuli received from the sensors situated on the cytoplasm and transmitting signals to the transcriptional machinery in the nucleus to withstand a stressful environment effectively. Signaling pathways serve as a crucial nexus for sensing stress and establishing Frontiers in Plant Science CITATION Pandey P, Tripathi A, Dwivedi S, Lal K and Jhang T (2023) Deciphering the mechanisms, hormonal signaling, and potential applications of endophytic microbes to mediate stress tolerance in medicinal plants.

Ripening Behavior of the Green Ripe Tomato Mutant

Article

Full-text available

Sep 1984

The fruit ripening behavior of the green ripe mutant ( Gr ) of tomato ( Lycopersicon esculentum Mill.) was examined. Green ripe fruit are climacteric and evolve increasing amounts of ethylene after harvest; however, the time course for these events is dramatically altered in comparison with ‘Rutgers’. Maximal rates of C 2 H 4 evolution from Gr fruit were achieved 20 days after the initial increase, and 7 to 10 days prior to maximal respiratory rates. Fruit age at harvest did not affect either the rate or the magnitude of these processes. Wavelength scans of pigment extracts from 60 day postharvest Gr fruit indicated low levels of carotenoids. Mutant fruit also remain firm a long time after harvest. Polygalacturonase activity in Gr fruit increases with fruit age, but reaches only 3% to 5% of the total activity in ‘Rutgers’. PG activity was only slightly reduced when extracts were heated to 65°C for 5 min, suggesting that a heat stable isoenzyme of PG is predominant in mature mutant fruit in contrast to ‘Rutgers’ in which 90% to 95% of PG activity in ripe fruit is heat labile under these conditions. When heterozygous, the Gr mutation is dominant in its effects on total PG activity and on the time course from the initiation of C 2 H 4 and respiratory increases to their maximal rates of evolution. The magnitude of other changes in heterozygous mutant fruit was intermediate between normal and homozygous Gr .

Ripening-associated Biochemical Traits of Cantaloupe Charentais Melons Expressing an Antisense ACC Oxidase Transgene

Article

Full-text available

Nov 1997

Transgenic Cantaloupe Charentais melons (Cucumis melo var. cantalupensis Naud. 'Vedrantais') exhibiting strong inhibition of ethylene production were used as a model to discriminate between ethylene-regulated and ethylene-independent ripening pathways. Compared to wild-type fruit, transgenic fruit did not undergo significant yellowing of the rind and softening of the flesh. However, these effects were completely reversed by treating transgenic fruit with 50 μL·L-1 exogenous ethylene. Pigmentation of the flesh occurred early before the onset of the climacteric and was thus unaffected by ethylene inhibition in transgenic fruit. Total soluble solids accumulated at the same rate in both types of fruit until 38 days after pollination when wild-type fruit abscissed. However, as ethylene-inhibited fruit failed to develop a peduncular abscission zone, they remained attached to the plant and accumulated higher amounts of sugars, mainly sucrose. Harvesting transgenic fruit resulted in a small but significant increase of internal ethylene associated with softening of the flesh.

Genetic Regulation of Tomato Fruit Ripening1

Article

Full-text available

Oct 1978

Ripening is a dramatic event in the development of many fleshy fruits. Tomato ripening involves a number of chemical and physical changes which convert the fruit from a relatively inedible state to one of optimal quality (2, 19). These changes appear to be highly synchronized, as evidenced by the fact that respiratory patterns, rate of ethylene production, carotene development, and flavor and textural changes normally associated with the ripening process, occur in close succession during the relatively short period in which the fruit ripens (2, 22). The association of these changes with seed maturation supports the popular view that ripening is of adaptive significance in seed dispersal by rendering fruit attractive to animals responsible for dispersal.

Ripening-induced Chemical and Antioxidant Changes in Bell Peppers as Affected by Harvest Maturity and Postharvest Ethylene Exposure

Article

Full-text available

Jun 2005

Greenhouse-grown bell pepper (Capsicum annuum L. 'Robusta') were harvested at five stages of maturation (10% red to full red) in early winter 2002 (Expt. 1) and at two stages (10% red and full red) in early Spring 2002 (Expt. 2). The fruit were subsequently stored at 20°C in a continuous-flow chamber consisting of either 100 μL·L-1 ethylene (balance air) or air-only (control) at 90% relative humidity (RH). Individual fruit were removed from the chambers upon reaching full red color, and stored at -30°C until physicochemical analyses were conducted. Harvest maturity, and ethylene exposure had no appreciable effect on pulp soluble solids content, total titratable acidity or pH. Exposure to ethylene hastened ripening time compared to the air control but was independent of fruit maturity at harvest. Fruit exposed to ethylene reached full-red color 6.4 days (Expt. 1) and 4 days (Expt. 2) earlier than air-only fruit, respectively. There were no significant phytochemical and antioxidant differences noted for total carotenoids, total ascorbic acid, and soluble phenolics at various maturity stages due to ethylene exposure. Appreciable differences were observed between the two experiments for phytochemicals and antioxidants, as bell peppers from the latter experiment contained at least twice the concentrations of phytochemicals and antioxidant capacity as those from the first experiment. Differences in these parameters between experiments were attributed to environmental factors such as average temperature, day length, and light intensity. Ethylene was demonstrated to be an effective postharvest treatment for accelerating color change in this bell pepper cultivar, permitting earlier harvest without altering phytochemical synthesis rates.

Insensitivity to Ethylene Conferred by a Dominant Mutation in Arabidopsis thaliana

Article

Aug 1988

Ethylene influences a number of developmental processes and responses to stress in higher plants. The molecular basis for the action of ethylene was investigated in mutants of Arabidopsis thaliana that have altered responses to ethylene. One mutant line, which has a dominant mutation at a locus designated etr, lacks a number of responses to ethylene that are present in the wild-type plant. These include inhibition of cell elongation, promotion of seed germination, enhancement of peroxidase activity, acceleration of leaf senescence, and feedback suppression of ethylene synthesis by ethylene. These diverse responses, which occur in different tissues of Arabidopsis, appear to share some common element in their transduction pathways—for example, a single receptor for ethylene. Results of ethylene binding experiments in vivo indicate that this receptor may be affected by the etr mutation.

The ethylene signalling pathway: New insights

Article

Jan 2003
CURR OPIN PLANT BIOL

Allelotype of a Ripening-specific 1-Aminocyclopropane-1-carboxylate Synthase Gene Defines the Rate of Fruit Drop in Apple

Article

Jan 2004

The onset of apple [Malus sylvestris (L.) Mill. Var. domestica (Borkh.)Mansf.] fruit maturity is preceded by the production of ethylene, the ripening hormone, which induces fruit ripening. The amount of ethylene produced by the fruit correlates with the level of transcription of the ripening-specific 1-aminocyclopropane-1-carboxylate (ACC) synthase genes. We have found that an allele (MdACS1-2), which contains an inserted retroposon-like sequence at the 5′-flanking region, is transcribed at a lower level than the wild-type (MdACS1-1). MdACS1-2/2 homozygous fruit produce a lower level of ethylene at the climacteric stage than do the wild type fruit. We have also found that the preharvest drop rates of apple cultivars and strains of MdACS1-2/2 trees have less fruit drop than the MdACS1-1/1 or MdACS1-1/2 trees. Treatment of the MdACS1-1/2 trees with 1-MCP, an ethylene receptor blocker, further decreased fruit drop. Analysis of commercial apple cultivars for the presence of the MdACS1-2/2 allele may help in the early detection of apple cultivars with a low fruit drop rate.

Ethylene Production and Fruit-softening Rates in Several Apple Fruit Ripening Variants

Article

Feb 1993

Ethylene production and fruit softening during postharvest storage of several apple ( Malus domestica Borkh.) ripening variants were compared with two standard cultivars. PA14-238 and D101-110 produced only low levels of ethylene (<10 μl·kg –1 ·hour –1 ) at harvest and throughout most of 86 days of storage at 4C, whereas `Red Chief Delicious' and `Golden Delicious' fruit produced >100 μl ethylene/kg per hour during the same time period. PA14-238 and D101-110 flesh disks converted aminocyclopropane-1-carboxylic acid (ACC) but not methionine (MET) to ethylene. `Red Chief Delicious' readily converted both MET and ACC to ethylene at the end of cold storage. PA14-238 fruit were the firmest and did not soften during postharvest storage; however, D101-110 softened appreciably. NJ55 did not produce ethylene at harvest, but produced a significant amount of ethylene (90 μl·kg –1 ·hour –1 ) during storage. Despite its high capacity to produce ethylene, NJ55 remained nearly as firm as PA14-238 at the end of cold storage.

Ethylene and Carbon Dioxide Production in Detached Fruit of Selected Pepper Cultivars

Article

Jul 1999

Bell peppers (Capsicum annuum L.) are classified as nonclimacteric fruits while some hot peppers have been reported as climacteric. Responses of peppers to exogenously applied ethylene-releasing compounds suggest ethylene involvement in the ripening process. Ethylene production and respiration rates in 13 cultivars of pepper: 'Camelot', 'Cherry Bomb', 'Chiltepin', 'Cubanelle', 'Banana Supreme', 'Habanero', 'Hungarian Wax', 'Mesilla', 'Mitla', 'Savory', 'Sure Fire', 'Tabasco', and 'King Arthur' were studied under greenhouse and field conditions. Fruit from each cultivar were harvested at different maturity stages determined by color, ranging from mature-green to full red-ripe. Carbon dioxide and ethylene production were measured by gas chromatography. Both variables were significantly different among maturity stages for all cultivars. Respiration rates were between 16.5 and 440.3 mg·kg-1·h-1 CO2. Ethylene production ranged from 0.002 to 1.1 μL·kg-1h-1. Two patterns of CO2 production were identified: higher CO2 production for mature-green fruit with successive decreases for the rest of the maturity stages or lower respiration rates for mature-green fruit with an increase in CO2 production either when fruit were changing color or once fruit were almost totally red. A rise in CO2 production was present for most cultivars. Ethylene evolution increased significantly at maturity or before maturity in all cultivars except 'Cubanelle' and 'Hungarian Wax'. Respiration rates and ethylene production were significantly different among cultivars at the mature-green and red stages.

Ethylene Biosynthesis and its Regulation in Higher Plants

Article