Biochimica et Biophysica Acta 1477 (2000) 112^121
www.elsevier.com/locate/bba
Review
The systemin signaling pathway: di¡erential activation of plant defensive
genes
Clarence A. Ryan
Institute of Biological Chemistry, Washington State University, Pullman, WA 99164-6340, USA
Abstract
Systemin, an 18-amino-acid polypeptide released from wound sites on tomato leaves caused by insects or other mechanical
damage, systemically regulates the activation of over 20 defensive genes in tomato plants in response to herbivore and
pathogen attacks. Systemin is processed from a larger prohormone protein, called prosystemin, by proteolytic cleavages.
However, prosystemin lacks a signal sequence and glycosylation sites and is apparently not synthesized through the secretory
pathway, but in the cytoplasm. The polypeptide activates a lipid-based signal transduction pathway in which the 18:3 fatty
acid, linolenic acid, is released from plant membranes and converted to the oxylipin signaling molecule jasmonic acid. A
wound-inducible systemin cell surface receptor with an Mr of 160 000 has recently been identified. The receptor regulates an
intracellular cascade including, depolarization of the plasma membrane, the opening of ion channels, an increase in
intracellular Ca2‡ , activation of a MAP kinase activity and a phospholipase A2 activity. These rapid changes appear to play
important roles leading to the intracellular release of linolenic acid from membranes and its subsequent conversion to
jasmonic acid, a potent activator of defense gene transcription. Although the mechanisms for systemin processing, release,
and transport are still unclear, studies of the timing of the synthesis and of the intracellular localization of wound- and
systemin-inducible mRNAs and proteins indicates that differential syntheses of signal pathway genes and defensive genes are
occurring in different cell types. This signaling cascade in plants exhibits extraordinary analogies with the signaling cascade
for the inflammatory response in animals. ß 2000 Elsevier Science B.V. All rights reserved.
Keywords: Tomato; Prosystemin; Octadecanoid pathway; Systemic induction; Plant^herbivore interaction; Plant^pathogen interaction ;
(Lycopersicon esculentum)
1. Introduction
Plants have a variety of chemical defenses to protect themselves against herbivores [1,2]. These in-
Abbreviations: ABA, abscisic acid; ACC, 1-aminocyclopropane-carboxylic acid; AOS, allene oxide synthase; CaM, calmodulin; CPI, metallocarboxypeptidase inhibitor; CaMV, cauli£ower
mosaic virus; CDI, cathepsin D inhibitor; GUS, L-glucuronidase; JA, jasmonic acid; LOX, lipoxygenase; MeJA, methyl
jasmonate; OGA, oligogalacturonic acid; OPDA, 12-oxy-phytodienoic acid; PG, polygalacturonase; PLA2 , phospholipase A2 ;
PPO, polyphenol oxidase
clude the production of chemicals, from small organics to large proteins and enzymes, that have various
deterrent e¡ects on attacking herbivores that consume them. Other responses commonly found among
many plant genera produce volatile signals at sites of
larval attacks that attract insect predators such as
wasps and mites [1].
Many plants respond to herbivore attacks by activating defense genes in leaves whose products inhibit
digestive proteases of herbivores and reduce the nutritional quality of the ingested proteins, making the
attackers ill [3]. In tomato plants, wounding causes a
systemic reprogramming of leaf cells that results in
0167-4838 / 00 / $ ^ see front matter ß 2000 Elsevier Science B.V. All rights reserved.
PII: S 0 1 6 7 - 4 8 3 8 ( 9 9 ) 0 0 2 6 9 - 1
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Fig. 1. Systemic wound responsive genes. Asterisks indicate gene products identi¢ed in cell suspension cultures. Other genes or gene
products have been identi¢ed in tomato leaves.
the synthesis of over 20 defense-related proteins [4^
9]. This is analogous to the in£ammatory and acute
phase responses of animals in response to trauma
[4,10]. Most of the newly synthesized proteins fall
into functional groups that include (i) antinutritional
proteins, including both proteinase inhibitors and
polyphenol oxidase (PPO), (ii) signal pathway components, and (iii) proteinases (Fig. 1). Several other
genes are also activated, but they do not fall into the
above groupings and their roles in defense are obscure.
The wound-inducible proteinase inhibitors [6,7,9^
11] cumulatively represent a wide range of speci¢cities that include all four known mechanistic classes
of proteases. PPO is also wound-inducible in leaves
[5] and, when ingested by herbivores together with
phenolics, can crosslink proteins, rendering them less
digestible. Together, the proteinase inhibitors and
PPO provide formidable barriers to protein digestion.
Wound-induced synthesis of components of the
signal transduction pathway (cf. Section 3) appears
to be a strategy of the plant to amplify its ability to
mount a maximal defense response against the attacking predators. Wound-inducible signal pathway
genes in tomato leaves include prosystemin [12],
CaM [13], LOX [14,15], and AOS (G. Howe, C.A.
Ryan, unpublished). The latter two enzymes are
components of the octadecanoid pathway [16] and
they reside in the chloroplast [14,15,17] where they
participate in the conversion of linolenic acid to signaling molecules, phytodienoic acid and jasmonic
acid, which are oxylipin analogs of prostaglandins
[18]. Whether other members of the signal pathway-associated genes are also synthesized in response
to herbivore attacks has not been determined, but
evidence using cell suspension cultures suggests that
other signaling-associated genes in intact plants may
also be wound-inducible, including the systemin receptor [19], ACC synthase (a key enzyme in ethylene
biosynthesis) [20], and an NADPH oxidase (an enzyme involved in the production of active oxygen
species) [21,22].
The ¢nal category of wound-inducible genes in
Fig. 1 is comprised of several proteinases, including
endopeptidases and exopeptidases. The function of
these proteinases is still unclear, but some may be
involved in protein processing, and others may
have a role in protein turnover and remodeling.
Knowledge of the localization of the synthesis of
these proteinases in leaves may provide some clues
to their functional signi¢cance in the defense response.
2. Systemin and prosystemin
A primary wound signal for the signaling cascade
is an 18-amino-acid polypeptide hormone called systemin [23] that is released at wound sites by chewing
herbivores (wounding) (Fig. 2). Systemin is active at
extraordinarily low levels (i.e., fmol/plant) [23] and
ranks among the most potent gene activators known.
Alanine scanning mutagenesis of the systemin sequence revealed that all residues except Thr17 could
be replaced without completely eliminating activity.
Ala17 is essentially inactive in inducing defense
genes, but it is a potent antagonist of systemin, [24]
suggesting that systemin-A17 can bind a receptor but
cannot activate the signal transduction system. Ra-
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C.A. Ryan / Biochimica et Biophysica Acta 1477 (2000) 112^121
Fig. 2. Amino acid sequence of prosystemin. The underlined sequences indicate amino acids within conserved regions resulting
from gene duplication^elongation events. The systemin sequence
is indicated within the C-terminus.
dioactively labeled systemin, when placed on fresh
wounds on tomato leaves has been shown to move
from wound sites to petiole phloem, supporting its
possible role as a mobile systemic wound signal
[23,25]. Systemin is processed from a 200-aminoacid precursor protein called prosystemin [12] (Fig.
2). Prosystemin is found at low levels in leaves of
unwounded plants, but increases several fold in response to wounding [12], apparently to amplify the
wound signal in plants when under attack. The enzymes involved in the processing of prosystemin have
not been identi¢ed, but possible processing enzymes
may include one or more of the wound-inducible
enzymes shown in Fig. 1. The processing apparently
takes place as a result of wounding, which is thought
to mix prosystemin with proteolytic enzymes from
another cellular compartment(s), resulting in the release of systemin.
A critical role for prosystemin and systemin in
signaling defense genes has been established by transforming tomato plants with a fused gene comprised
of a prosystemin cDNA in its antisense orientation,
driven by the 35S CaMV promoter [12]. Plants ex-
pressing the gene are severely impaired in their systemic induction of both inhibitor I and II proteins in
response to wounding. The plants were also compromised in their ability to defend themselves against
attacking Manduca sexta larvae [26]. As expected,
supplying antisense plants with systemin resulted in
the expression of the defensive genes, con¢rming that
the plants were capable of responding to the wound
signal, but could not produce it. Additionally, placing systemin or recombinant prosystemin on fresh
wounds of antisense plants reestablished the wound
response to wild-type levels in distal leaves [27]. The
characteristics of systemin, including its potency, its
mobility in the plant, the wound-inducibility of the
prosystemin gene, the e¡ects of the antisense gene in
blocking systemic wound signaling, and the reestablishment of the systemic wound response in antisense
plants, have led to the conclusion that systemin is a
systemic wound hormone that plays a central role in
regulating the expression of defense genes in response
to pest attacks.
Tomato plants transformed with the prosystemin
cDNA in its correct or `sense' orientation regulated
by a constitutive promoter expressed high levels of
prosystemin mRNA in leaves [28]. Surprisingly, the
plants constitutively synthesized wound-inducible defensive proteins throughout the plants in the absence
of wounding, apparently due to an abnormal release
of systemin from the overexpression of prosystemin.
Inhibitors I and II were found to accumulate to extraordinary high levels of several hundred Wg inhibitor protein per g leaf tissue. Analyses of the proteins
that accumulated in the transgenic plants (Fig. 3)
have led to the identi¢cation of several systemin-inducible genes in tomato leaves that are shown in Fig.
1. When wild-type tomato plants were grafted onto
the transgenic prosystemin rootstocks that overexpressed prosystemin, they synthesized high levels of
proteinase inhibitors, indicating that systemin, or a
defense signal produced by systemin, was transported
through the graft to the wild-type plants where it
also activated defense genes in the absence of wounding.
Recombinant prosystemin, synthesized in Escherichia coli [29], was as active as systemin in inducing
the synthesis of systemic wound response proteins
when supplied to young tomato plants through their
cut stems [27]. It is not known if full-length prosys-
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115
tivity of systemin, while the prosystemin lacking the
systemin sequence was totally inactive [27]. This indicated that the systemin sequence is likely the only
sequence within prosystemin that can activate the
signal transduction pathway.
Prosystemin has been isolated from potato, black
nightshade, and bell pepper [30]. All are homologous
to tomato systemin, and sequence identities among
them range from 73% to 88%. Systemins were synthesized from their deduced sequences and tested for
their abilities to induce proteinase inhibitor synthesis
in tomato. Potato and pepper were as active as tomato systemin, whereas nightshade systemin was 10fold less active. Tomato systemin is inactive in inducing proteinase inhibitor synthesis in leaves of tobacco, although a semi-puri¢ed polypeptide fraction
from tobacco leaves is highly active in inducing proteinase inhibitor synthesis in tobacco leaves (G.
Pearce, C.A. Ryan, unpublished). This suggests that
either the systemin gene has changed signi¢cantly
during evolution, or that other polypeptides may be
present in tobacco that serve a similar signaling role
as systemin in response to wounding. This suggests
that if polypeptides are defense-signaling molecules
throughout the plant kingdom, they may have minimally conserved structures or diverse structures that
may require individual isolation and characterization.
Fig. 3. SDS^PAGE of the soluble proteins extracted from
leaves of wild-type (WT) and transgenic tomato plants constitutively expressing the prosystemin gene. Arrows indicate proteins
that constitutively accumulate in the transgenic plants. These
same proteins accumulate systemically in wild-type plants in response to wounding.
temin is active, or if a processed form interacts with
the systemin receptor. Proteinases have been identi¢ed in apoplast £uids that can degrade prosystemin
to small polypeptides that retain immunoreactivity
against systemin antibodies [27]. Thus, it is likely
that systemin is processed from prosystemin when
supplied to plants via the transpiration stream. Additionally, recombinant prosystemin analogs were synthesized in E. coli in which either the prosystemin
sequence was mutated at residue 195, corresponding
Thr17 to Ala17 of systemin, or else the entire systemin sequence was deleted. The Ala17 analog exhibited
less than 1% of the proteinase inhibitor inducing ac-
3. The systemin signaling pathway
An updated original model [31] for the activation
of defense genes by systemin in tomato plants is
shown in Fig. 4. Using 125 I-labeled systemin, a systemin-binding protein has recently been identi¢ed in
the plasma membranes of leaf cells [32] and, more
recently, on the surface of Lycopersicon peruvianum
suspension cultured cells [19]. The properties of the
binding protein indicate that it is the receptor that is
involved in systemin-mediated signal transduction.
The dissociation constant of the systemin^receptor
interaction with the suspension cultured cell receptor
was 0.17 nM and is in the range of Kd s found for
polypeptide^receptor interactions in animal systems.
The receptor exhibited a speci¢c and reversible interaction with systemin, and the binding was shown to
increase several fold in response to MJ, suggesting
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Fig. 4. A current model for the systemic signaling pathway for
defensive genes in tomato plants that are activated by herbivore
attacks (wounding). The interaction of systemin with its membrane receptor initiates intracellular events that activate a
PLA2 . The phospholipase releases LA from membranes, the
production of JA, and the activation of defensive genes.
that the receptor is a systemic wound response protein. An Mr of 160 000 was determined for the receptor by photoa¤nity labeling and electrophoretic
analysis [19]. The isolation of the receptor protein
and its cDNA should provide a new avenue of research to seek interactive signaling components to
further elucidate the early events of signaling.
The interaction of systemin with its receptor regulates a complex cascade of intracellular event s that
are all orchestrated to activate a PLA2 to release
linolenic acid from membranes. These events include
a depolarization of the plasma membrane [33], the
opening of ion channels [32^34], an increase in the
concentration of intracellular Ca2‡ [34], the inactivation of a plasma membrane proton ATPase [35], the
activation of a MAP kinase [36,37], the synthesis of
calmodulin, and the activation of a PLA2 [38,39].
Release of linolenic acid from membranes leads to
its conversion to the oxylipins OPDA and JA that
regulate defense genes. JA, together with ethylene
has been shown [40] to activate transcription of the
defense-related genes by a mechanism that is still
obscure. Investigations of the individual components
of this pathway are important not only for the
understanding of the intracellular events leading to
plant defense gene activation, but of events leading
to stress and developmental responses that are regulated in plants by JA [18,41^44].
The identi¢cation of a systemin receptor on the
cell surface raises important questions regarding the
transport of systemin throughout the plants and its
delivery to the surface of the distal cells. Although
systemin was found to be mobile in the phloem, its
long-distance transport following wounding is not
well understood. For systemin to be regulating systemic activation of defense genes, it must be either
transported throughout the plants or somehow involved in a cascade that operates over long distances.
In any event, systemin appears to exert its e¡ects
through an external cell surface receptor. If systemin
is transported in the phloem then it is likely that it is
transported to companion cells and phloem parenchyma where it could be delivered to the apoplast,
presumably by a speci¢c transporter. On the other
hand, when systemin is initially released by wounding it may travel through the apoplast to nearby
vascular bundle cells where it can interact with receptors to activate the synthesis in the cells of both
prosystemin and its processing enzymes. This in turn
would produce more systemin, which could move
through the phloem and apoplast to distal cells to
continue a cascade through the plant. Another possibility is that the initially released systemin might
activate synthesis of prosystemin and a prosystemin
transporter that would deliver nascent prosystemin
to the apoplast where processing and receptor binding would both occur. In this scenario, the transport
of systemin through the plant would occur by apoplastic transport and phloem loading to e¡ect transport to more distal cells, cascading through the plant.
This latter scenario would require systemin to be
transported from the phloem to the apoplast to interact with the receptor. Both scenarios ¢t several
known facts, including the timing of movement of
the wound signal (systemin) through the petioles
(from 15 to 90 min), the tissue speci¢city of prosystemin synthesis in the vascular bundles (see Section
4), the movement of the wound signal from prosystemin transgenic rootstalks through grafts to wildtype scions in the absence of wounding, and the ob-
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servations that the addition of systemin or prosystemin to wound sites on leaves of prosystemin antisense tomato plants caused distal leaves to activate
defense genes [27].
Other primary signals have been proposed for activating defense genes in response to herbivore attacks, including electrical pulses, changes in hydraulic pressure, and the plant hormones ABA and
ethylene [45^48]. Electrical phenomena accompany
wounding but no evidence has been forthcoming to
de¢ne how weak electrical signals can play a role in
systemic signal transduction. The propagation of
both electrical and hydraulic pulses are very fast
compared to the known velocity of the movement
of the wound signal in plants [49]. Additionally, girdling petioles of young tomato plants with a stream
of hot air that kills all cells except the xylem elements
prevented systemic signaling of proteinase inhibitor I
in upper, ungirdled leaves in response to wounding
[49]. Under these conditions, hydraulic signals would
have been able to move through the xylem to unwounded leaves. Malone and Alorcan [50] reported
that wounding girdled tomato leaves induced inhibitory activity against elastase in upper, unwounded
leaves. It is possible that an entirely di¡erent signaling pathway is activating elastase inhibitors, but
these latter experiments did not give consistent results, and speci¢c inhibitor quanti¢cation was not
recorded, so that basal levels of inhibitor at the onset
of experiments could not be assessed. There is no
doubt that electrical and hydraulic waves do propagate throughout the plants in response to wounding,
but their e¡ects on plant defense activation still remains obscure. In this regard, in the grafted plants
mentioned earlier in which `sense' prosystemin transgenic rootstalks caused the induction of proteinase
inhibitors in the wild-type scions in the absence of
wounding, no generation of electrical or hydraulic
signals could have occurred.
In wild-type plants, breaching the vascular system
has been proposed to perturb membranes throughout the plants [37], so that a priming of the signal
transduction system through a hydraulic event might
e¡ect an early transient activation of the MAP kinases and the PLA2 that are involved in the early
signaling events, similar to a `touch response'. Transient signaling in response to hydraulic signals may
be part of an early alarm system to activate signaling
117
pathway components, but weak, transient, electrical,
or hydraulic signals alone cannot sustain the activation of defense genes over a period of several hours.
ABA appears to play an overall role in determining whether the plant can mount a defense response
[51,52]. Tomato plants with a mutation in ABA biosynthesis were found to be de¢cient in the wound
response, and the response could be restored by exogenously ABA applied to leaves [51]. ABA does not
behave as if it is a primary component of the signaling pathway since it does not act as an inducer of
proteinase inhibitor synthesis in tomato plants, as do
wounding, systemin, or other elicitors [52], but it is
required for the plants to respond maximally [51].
Other plant hormones are also important to the defense response, including auxin, which inhibits the
response [53], and ethylene [40] which is a component
of the signaling pathway.
4. Di¡erential signaling by systemin ^ a modi¢ed
model
In comparing the timing of the wound-inducible
expression of proteinase inhibitor genes with those
of the signal transduction pathway, it became clear
that two classes of wound- and systemin-inducible
genes were di¡erentially regulated. As shown in
Fig. 5, mRNAs coding for proteinase proteins
mRNAs appeared much later than mRNAs coding
for signal pathway proteins. The mRNAs encoding
proteinase inhibitors I and II [11], CPI ([54]; M.
Diaz, C.A. Ryan, in preparation), and an aspartic
proteinase inhibitor, CDI [4,7], were ¢rst detectable
in northern analyses at about 2 h after the plants
were wounded, and the levels continued to increase
through the next 8 h. On the other hand, mRNAs
encoding wound-inducible LOX [15], CaM [13], AOS
(G. Howe, C.A. Ryan, unpublished), and prosystemin [12] were ¢rst detectable within about 30 min,
and were maximally induced 2^3 h after wounding.
This was 5^6 h before the proteinase inhibitor
mRNAs reached maximal levels. AOS mRNA has
been reported to be wound inducible in Arabidopsis
leaves [55] with induction kinetics similar to LOX
mRNAs in tomato leaves, beginning at 0.5 h after
wounding and maximizing at about 2^4 h, then declining thereafter.
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C.A. Ryan / Biochimica et Biophysica Acta 1477 (2000) 112^121
Fig. 5. The di¡erential induction of mRNAs of systemic wound
response proteins. Young tomato plants were wounded on the
lower leaves at time zero and assayed by Northern blotting at
the times indicated. Inh I, proteinase inhibitor I; Inh II, proteinase inhibitor II; CDI, cathepsin D inhibitor (and aspartic
proteinase inhibitor); CPI, metallocarboxypeptidase inhibitor;
CaM, calmodulin; LOX, lipoxygenase; ProSys, prosystemin.
Ultrastructural studies of the intracellular localization of proteinase inhibitor I, synthesized in response
to wounding, had revealed previously that the inhibitor is sequestered in the central vacuoles of mesophyll cells of tomato leaves [56]). Inhibitor II has also
been found in leaf vacuoles [57,58], and it is likely
that other proteinase inhibitors with long half lives
are also sequestered in vacuoles of mesophyll and/or
palisade cells. In contrast, when the promoter region
of the wound-inducible prosystemin gene was fused
with the GUS reporter gene, and tomato plants were
transformed with the fused gene, the plants responded to wounding and MeJA by synthesizing
the GUS protein [59]. The localization of GUS activity was visualized by supplying a colorimetric substrate to freehand sections of leaf tissues. The
wound-inducible GUS activity was only found in
the vascular bundles of the major and minor veins
of the leaves. This was con¢rmed by tissue printing
using speci¢c antibodies prepared against prosystemin and visualized before and after wounding using
a colorimetric substrate. The AOS gene in Arabidopsis [55] that is a component of the defense signaling
pathway is also speci¢cally expressed in the vascular
bundles in response to wounding (E. Weiler, personal
communication). We hypothesize that the signal
transduction genes as a group are `early genes' that
are expressed in the vascular bundle cells, while the
defensive proteinase inhibitor genes are `late genes'
that are expressed in palisade and spongy mesophyll
cells (Fig. 6).
The di¡erences in timing and localization of these
two functionally di¡erent groups of proteins suggests
a scenario in which the signal transduction pathway
is initially activated in the vascular bundles to amplify the production of second messengers, which could
include OPDA and/or JA, derivatives of these molecules [60], or other signaling molecules that are transported to other cell types. The second messengers are
then transported to the palisade and mesophyll cells
where the defense proteins are synthesized and compartmented.
A comparison of the relative time courses of the
various components of the signal transduction pathway from the initial release of systemin to the production of proteinase inhibitors is shown in Fig. 7.
This scenario di¡ers signi¢cantly from a recently proposed signaling model [48] in which systemin was
suggested to be a signal released in response to jasmonic acid. In this latter model, jasmonic acid and
ethylene were not considered downstream systemic
signals because increases in ethylene and jasmonic
acid cannot be detected levels in unwounded leaves
of wounded plants [48]. The author did not consider
that JA is likely transient and only present at very
Fig. 6. A modi¢ed model to indicate the di¡erential signaling
of signal pathway components and the proteinase inhibitors in
di¡erent cell types.
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Fig. 7. A time-course representation of the progression of the
production of signal pathway components in tomato leaves in
response to wounding at time 0. H, a transient hydraulic pulse
due to the breaching of the vascular system by wounding,
which initiates the signaling events.
low concentrations in speci¢c cell types. There is no
evidence available to bear on the minimum quantities
of these compounds that can activate genes, or
whether OPDA or JA is the signaling component.
Several examples are known of JA-inducible genes
being activated without measurable increases in JA.
As stated by Weiler et al. [61], `clearly, results based
solely on analysis of JA can no longer be regarded as
complete'. Bowles also concluded [48], based on unpublished data, that Ala17-systemin, the potent antagonist of systemin mentioned earlier, inhibits the
action of OGA elicitors. In contrast, through extensive experimentation (G. Pearce, C.A. Ryan, unpublished), we have shown conclusively that Ala17-systemin has no e¡ect on the proteinase inhibitor
inducing activity of either OGA or chitosan, and
therefore systemin must be perceived independently
from these elicitors, likely through di¡erent receptors. A recent report has demonstrated that chitosan,
OGA, and systemin all activate PLA2 activity in tomato plants [39], and all three elicitors act via the
octadecanoid pathway [62] although it is not yet
known if the same PLA2 is involved with signaling
by all three elicitors.
While the downstream message that activates pro-
119
teinase inhibitor synthesis in response to wounding
and systemin may be JA or a derivative in conjunction with ethylene, new evidence from our research
suggests that other candidates for second messengers
are produced in response to wounding and systemin
that could be part of the signaling pathway for proteinase inhibitor synthesis. The candidates are (i) the
OGA derived from the cell walls of vascular bundle
cells by the action of a newly discovered wound-,
systemin- and JA-inducible PG [6], and (ii) reactive
oxygen species [22], known to be produced in tomato
leaves by OGAs [63], through NADPH oxidase [63^
65]. A speci¢c inhibitor of NADPH oxidase, diphenylene iodonium chloride, blocks wound-inducible
H2 O2 generation in tomato leaves [22], and we
have recently shown that this inhibitor powerfully
blocks the synthesis of inhibitor I and II protein
accumulation in response to wounding, systemin,
and MeJA (M. Orozco-Cardenas, C.A. Ryan, in
preparation), although whether the inhibitor is speci¢c for NADPH oxidase in vivo is not known. If
OGA and H2 O2 are indeed second messengers for
the activation of proteinase inhibitor genes in mesophyll cells, they are likely downstream in the signaling pathway since they are inducible by JA and PG
in the vascular bundles. The production of OGA and
H2 O2 as second messengers is kinetically compatible
with the synthesis of proteinase inhibitors since both
are produced later than the signal transduction
mRNAs ([22]; see Fig. 5), but concurrent with the
synthesis of proteinase inhibitor mRNAs (i.e., between 2 and 8 h after wounding).
Reactive oxygen plays a central role in the resistance response of plants against pathogens leading to
incompatibility and the development of systemic acquired resistances [66]. The increase in active oxygen
during herbivore attacks has been suggested to act as
a deterrent to herbivores [66], and the wound-inducible reactive oxygen might potentiate the defense response against pathogens as well. While there is no
evidence to date to link wound-inducible H2 O2 in
tomato leaves to pathogen resistances, the octadecanoid pathway is important to phytoalexin production
in several species of plants [67,68] and the pathway
has been shown through mutant analyses to be a
factor in pathogen resistance in Arabidopsis [69^72].
In a broader context, the defense signaling pathways in plants contain common types of components
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C.A. Ryan / Biochimica et Biophysica Acta 1477 (2000) 112^121
that have contributed to the evolution of information
processing systems for the activation of defenses
against both herbivores and pathogens in particular
ecological niches. For example, receptors have
evolved in plants that can recognize signals unique
to speci¢c pests or pathogens [73^76]. These signals
regulate the tissue-speci¢c and/or cell-speci¢c expression of a variety of defensive genes in di¡erent plants
to protect against speci¢c herbivores and/or pathogens. Herbivore saliva can be a source of signals to
activate defense genes [77], or to signal the production of chemicals to attract predators of insects
chewing on the plants [78,79]. While the various signals and genes produced by herbivore and pathogen
attacks are likely to di¡er among di¡erent plant species, it is also likely that some other signal pathway
components, e.g., ion channels, MAP kinases, and
transcription factors, will have evolved with some
conservation of structures and functions. Perhaps
with time these fundamental components may track
back to ancestral genes common to plant and animal
defense systems. Knowledge of the biochemical details of the evolution of defense signaling pathways
of plants could have an enormous potential in helping to understand the biological characteristics of
species that are found in various ecosystems, as
well as in applying this knowledge to the agricultural
community to increase plant productivity using natural defense strategies
Acknowledgements
Supported in part by Washington State University
College of Agriculture and Home Economics, Project
1791 and Grants DCB 9104542 and DCB 9117795
from the National Science Foundation, and Grant
#9801502 from the U.S. Department of Agriculture.
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