THE GENERATION OF Ca2+ SIGNALS IN PLANTS · 24 Apr 2004 19:45 AR AR213-PP55-16.tex...

27 Apr 2004 15:9 AR AR213-PP55-16.tex AR213-PP55-16.sgm LaTeX2e(2002/01/18) P1: GDL10.1146/annurev.arplant.55.031903.141624

Annu. Rev. Plant Biol. 2004. 55:401–27doi: 10.1146/annurev.arplant.55.031903.141624

Copyright c© 2004 by Annual Reviews. All rights reservedFirst published online as a Review in Advance on January 12, 2004

THE GENERATION OF Ca2+ SIGNALS IN PLANTS

Alistair M. Hetherington and Colin BrownleeDepartment of Biological Sciences, Lancaster Environment Center, University ofLancaster, Lancaster LA1 4YQ, United Kingdom; email: [email protected] Biological Association of the United Kingdom, The Laboratory, Citadel Hill,Plymouth PL1 2PB, United Kingdom; email: [email protected]

Key Words signal transduction, stimulus-response coupling, nodulation factor,guard cells, calcium channels

■ Abstract The calcium ion is firmly established as a ubiquitous intracellular sec-ond messenger in plants. At their simplest, Ca2+-based signaling systems are composedof a receptor, a system for generating the increase in [Ca2+]cyt, downstream compo-nents that are capable of reacting to the increase in [Ca2+]cyt, and other cellular systemsresponsible for returning [Ca2+]cyt to its prestimulus level. Here we review the vari-ous mechanisms responsible for generating the stimulus-induced increases in [Ca2+]cyt

known as Ca2+ signals. We focus particularly on the mechanisms responsible for gen-erating [Ca2+]cyt oscillations and transients and use Nod Factor signaling in legumeroot hairs and stimulus-response coupling in guard cells to assess the physiologicalsignificance of these classes of Ca2+ signals.

CONTENTS

INTRODUCTION . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 402DISSECTING THE COMPONENTS OF Ca2+ SIGNALS . . . . . . . . . . . . . . . . . . . . . 403

Entry Points: Ca2+-Permeable Channels . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 403Plasma Membrane Depolarization-Activated Ca2+ Channels (DACCs) . . . . . . . . . . 403Plasma Membrane Hyperpolarization-Activated Ca2+ Channels

(HACCs) . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 404Plasma Membrane NSCCs Indicate Diverse Functions Including Ca2+

Signaling . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 405Endomembrane Ca2+ Channels: Unidentified Players with ImportantRoles . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 406

Shaping the Ca2+ Signal: Pumping and Binding! . . . . . . . . . . . . . . . . . . . . . . . . . . . 407Functional Roles: Evidence from Disruption Studies . . . . . . . . . . . . . . . . . . . . . . . . 408ER Ca2+ Binding and Homeostasis . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 408Mitochondria: Energetic Players in a Dynamic Ca2+ Network . . . . . . . . . . . . . . . . 409Mitochondria as Ca2+ Buffers: “Marks and Sparks” . . . . . . . . . . . . . . . . . . . . . . . . 409

PROPAGATION OF PLANT Ca2+ SIGNALS . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 410[CA2+]cyt OSCILLATIONS AND TRANSIENTS . . . . . . . . . . . . . . . . . . . . . . . . . . . . 411

Calcium Oscillations in Legume Root Hair NF Signaling . . . . . . . . . . . . . . . . . . . . 411

1543-5008/04/0602-0401$14.00 401

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24 Apr 2004 19:45 AR AR213-PP55-16.tex AR213-PP55-16.sgm LaTeX2e(2002/01/18)P1: GDL

402 HETHERINGTON ¥ BROWNLEE

Generation of NF-Induced [Ca2+]cyt Oscillations in Root Hair Cells. . . . . . . . . . . . 412Stimulus-Induced Oscillations and Transients of Guard Cell [Ca2+]cyt . . . . . . . . . . 413Generating Stimulus-Induced Oscillations and Transients in GuardCell [Ca2+]cyt . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 413

Significance of [Ca2+]cyt Oscillations and Transients in Plant CellSignaling . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 416

CONCLUSIONS AND FUTURE PERSPECTIVES. . . . . . . . . . . . . . . . . . . . . . . . . . 418

INTRODUCTION

The calcium ion (Ca2+) is firmly established as a ubiquitous intracellular secondmessenger in plants (114, 116). An increase in the cytosolic concentration of Ca2+

([Ca2+]cyt) couples a diverse array of signals and responses. At their simplest, Ca2+-based signaling systems are composed of a receptor, a system for generating theincrease in [Ca2+]cyt, downstream components that are capable of reacting to theincrease in [Ca2+]cyt, and other cellular systems responsible for returning [Ca2+]cyt

to its prestimulus level. This apparently simple scheme hides considerable com-plexity. Although it has been argued recently that plant Ca2+ elevations may moreoften act as switches in the signaling process (118) rather than encoding specificinformation, it is clear that the cell has multiple mechanisms for generating in-creases in [Ca2+]cyt (116, 136), which suggests the capacity to produce highly com-plex spatio-temporal patterns of [Ca2+]cyt elevation. Downstream of the stimulus-induced [Ca2+]cyt increase (or Ca2+ signal), the cell possesses an array of proteinsthat can respond to changes in [Ca2+]cyt such as calmodulin (CaM) (85, 120),Ca2+-dependent protein kinases (19, 54), and CaM-binding proteins (113, 143).

Further complexity arises with the involvement of other intracellular messen-gers and signaling proteins that can modulate Ca2+ signaling. In recognition ofthis increasing complexity, it is perhaps more appropriate to represent intracellularsignaling as a network. Within a network-based organization a stimulus-inducedincrease in [Ca2+]cyt can be regarded as one of a small number of nodes or hubsresponsible for orchestrating the events that comprise the full cellular response.The possibility that guard cell signaling might be organized as and exhibit prop-erties reminiscent of a specific type of network known as a scale-free network hasbeen discussed (57, 58).

In this review we only consider the processes involved in generating Ca2+ sig-nals. Currently, our understanding of how complex spatio-temporal patterns of[Ca2+]cyt elevation contribute to controlling responses in stimulated cells is frag-mentary. Nevertheless, recent work has provided strong evidence that informationis encoded in [Ca2+]cyt oscillations and transients, at least in guard cells (2, 3).To assess the physiological importance of [Ca2+]cyt oscillations and transients wereview how they are generated. Although we restrict our discussion to the roles ofmitochondria and plasma membrane and endomembrane Ca2+-permeable chan-nels in this process, it should not be forgotten that nuclei (106, 107) and chloroplasts

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CALCIUM SIGNALING 403

(65, 115) also contribute to Ca2+ signal generation. The physiological context ofour discussions is provided by focusing on the roles of [Ca2+]cyt transients andoscillations in Nod Factor (NF) signaling and stimulus-response coupling in guardcells.

DISSECTING THE COMPONENTS OF Ca2+ SIGNALS

Entry Points: Ca2+-Permeable Channels

Ca2+-permeable channels are the key entry points for Ca2+ into the cytosol (116).Elevation of [Ca2+]cyt can arise via increased influx and/or decreased efflux (Fig-ure 1), although generally most Ca2+ signals initiate from increased Ca2+ channelactivity. In animal cells, specific [Ca2+]cyt elevations have been ascribed to Ca2+

influx through specific Ca2+-selective channels, at both the whole-cell (50) andsingle-channel level (145). The pharmacological profiles and selectivities of plantplasma membrane Ca2+-permeable channels often differ significantly from theiranimal counterparts (136). In plants Ca2+ permeable, rather than Ca2+-selective,channels predominant and there is a general lack of specificity of pharmaco-logical agents. These differences may reflect signaling strategies of plants thatrely largely on adaptive rather than behavioral responses as well as the differ-ent modes of energization and more hyperpolarized nature of most plant plasmamembranes.

The known properties of the major groups of plant Ca2+-permeable channelshave been surveyed in several recent comprehensive reviews (26, 130, 136). At themolecular level these have been broadly classed as nonselective cation channels(NSCCs). They are encoded by 41 genes of thecyclic nucleotide gated channel(CNGC) or glutamate receptor channel(GLR) families (see below) and thetwopore calcium channel(TPC) gene. However, in only a few cases have physiolog-ically characterized channels been mapped to their molecular counterparts. Herewe focus on those channels that have been implicated in bringing about specificCa2+ elevations.

Plasma Membrane Depolarization-ActivatedCa2+ Channels (DACCs)

Plasma membrane depolarization is a frequently observed response to various bi-otic (32, 81) and abiotic (104) stimuli that also give rise to [Ca2+]cyt elevations,which suggests the involvement of DACCs (135). Nonselective, cation-permeableDACCs have been described in a number of cell types such asArabidopsisrootcells (128). Their activity appears to be downregulated by the microtubule cy-toskeleton. However, their pharmacology is relatively unexplored and there are noreports directly relating DACC activity in situ with elevations in [Ca2+]cyt. Theseare difficult channels to characterize due to small currents and activity rundown,although channel recruitment in whole-cell recordings is possible with strong

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depolarization or microtubule disrupters (128). DACCs from cereal root plasmamembrane have also been characterized in lipid bilayers (136). The molecularidentities of DACCs have not been verified.AtTPC1, which has homology withthe animalα1 subunit of voltage-dependent Ca2+ channels, and theScCCH/MID1gene in yeast (41) has been proposed as a candidate gene (137). However, evidencethatAtTPC1encodes a depolarization-activated channel is indirect so far. Expres-sion ofAtTPC1in ScCCH1-deficient yeast enhances Ca2+ uptake and growth rate,and in Arabidospis,AtTPC1is required for the [Ca2+]cyt elevation in response tosucrose supply (41). The sucrose-induced Ca2+ rise depends on the expression ofsucrose symporter genes [AtSUC1andAtSUC2], and it is plausible that depolar-ization associated with sucrose/H+ cotransport underlies channel activation.

Plasma Membrane Hyperpolarization-ActivatedCa2+ Channels (HACCs)

Ca2+ channels that underlie hyperpolarization-activated Ca2+ currents across theplasma membrane have been relatively well characterized. They were first de-scribed in tomato cells (43) and were activated in response to fungal elicitors(44). Significantly, aequorin-transformed parsley cells show a biphasic elevationof [Ca2+]cyt in response to aPhytophoraoligopeptide elicitor (13).

HACCs have also been described in root hair apices, and root epidermal, endo-dermal, and cortical cells from the elongation zone and in guard cells (116). Theiractivation at hyperpolarized membrane potentials (more negative than−100 mV)leads to elevation of Ca2+ in guard cells (49, 109), and they play a critical rolein the response of stomata to abscisic acid (ABA). In root hairs their preferentialactivity at the growing apex (129) suggests a role in the sustained tip-localized el-evation of [Ca2+]cyt, and their presence in elongating root cells (68) suggests a rolein sustained Ca2+ influx. Spontaneous or evoked hyperpolarization leads to inwardCa2+ currents and corresponding elevation of [Ca2+]cyt (49) in Vicia guard cells.The observed [Ca2+]cyt elevation is relatively slow, occurring over tens of seconds.Grabov & Blatt (49) suggested that ABA-induced [Ca2+]cyt elevation could resultfrom hyperpolarization-induced HACC activity, although it is also likely that Ca2+

release from intracellular stores plays a significant role in the ABA response.In addition to regulation by membrane voltage, Ca2+

cyt (129) and reactive oxygenspecies (ROS), specifically OH∗, increase HACC activity inArabidopsisroot hairs(40). Molecular evidence for upstream activation of HACC activity by ROS, and anessential role in cell growth comes from work with theArabidopsis rhd2[AtrbohC]nicotinamide adenine dinucleotide phosphate (NADPH) oxidase mutant. Thesemutants fail to elongate root hairs and are defective in ROS production and thegeneration of the root hair apical [Ca2+]cyt gradient (40, 140). Both mutant andwild-type cells have similar OH∗-activated HACC activity. This study suggests thatROS is an essential upstream signal regulating cell growth via HACC regulation. Inguard cells, ROS (specifically H2O2) (76, 108) and phosphorylation (73) increaseHACC activity (see below), but in contrast to root hairs, elevated Ca2+

cyt decreases it

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(53). This apparent differential regulation by Ca2+ and ROS raises the possibilitythat different isoforms occur in these two cell types.

The molecular identity of HACC is not known. However, two possibilities havebeen proposed. First, mutation in theAtCNGC2gene underlies the defense nodeath (dnd) mutation inArabidopsis. AtCNGC2has inward-rectifying propertieswhen expressed inXenopusand is nonselective for cations. This led Demidchikand colleagues (25) to suggest that it may be an HACC. However, the apparent in-sensitivity to cyclic nucleotides of theArabidopsisroot hair HACC (V. Demidchik& J.M. Davies, unpublished data) argues against the identity of the HACC as aCNGC. Second, White et al. (137) proposed that an annexin may underlie HACCactivity based on their Ca2+permeability, localization, and known channel-formingproperties of annexins in animal cells (45). Annexins comprise a small family inArabidopsis(21) that have been linked with Ca2+ signaling and fluxes (20, 137)and the Ca2+-dependent regulation of membrane cycling (17). Analysis of annexinmutants should shed more light on their roles as Ca2+-permeable channels.

Plasma Membrane NSCCs Indicate Diverse FunctionsIncluding Ca2+ Signaling

NSCCs have been characterized inArabidopsisroot epidermal cells and mediatesteady-state Ca2+ influx at more depolarized membrane potentials than HACCs(25). These rapidly activating channels show little voltage dependence of activationand are active over a wide range of membrane potentials. They also co-occur withHACCs in the same cell type. Such channels may provide a crucial Ca2+ influxpathway at moderate membrane potentials as found, for example, in root epidermalcells (86), increasing the range of potentials over which plasma membrane channelsmay play a role in Ca2+ signaling.

CNGCs are represented by 20 members inArabidopsis. In animals their activityis increased by cAMP or cGMP. Members of theArabidopsisCNGC family possessoverlapping nucleotide-binding and CaM-binding regions at their C terminal (116,126, 137). Binding CaM and cyclic nucleotides provides the potential for dualregulation by distinct signaling pathways and possible mechanisms for integratingCa2+ and cyclic nucleotide signals (126). However, direct regulation of Ca2+ fluxor [Ca2+]cyt dynamics by cyclic nucleotides for plant CNGCs remains to be shown.Cyclic nucleotide modulation of a Na+-permeable channel has been reported inArabidopsisroot protoplasts, but cGMP or cAMP inhibited rather than enhancedactivity (87). CNGCs probably have a wide range of functions not all related toCa2+ signaling. However, human kidney (HEK) cells transformed withAtCNGC2showed that [Ca2+]cyt increases after treatment with membrane-permeant cAMPand cGMP (80). The demonstration of a role for cAMP in pollen tube growthregulation raises the possibility that CNGC regulation could be involved in theformation of the apical [Ca2+]cyt gradient that directs pollen tube growth (97).

The 20 members of the GLR family are also good candidates for nonselectiveCa2+-permeable channels (24, 26). GLRs have fundamental roles in animal cells

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in mediating fast chemical transmission and long-term synaptic potentiation (29).Plant GLR ligand binding is suggested by the similarities between the animaland plant genes in the ligand-binding domains (24, 26). Glutamate can inducerelatively fast (within a few seconds) increases in [Ca2+]cyt in aequorin-transformedwhole Arabidopsisseedlings (27). Ca2+ influx is implicated in this response bythe inhibitory effects of La3+.

Endomembrane Ca2+ Channels: Unidentified Playerswith Important Roles

Because many Ca2+ responses in plant cells involve Ca2+ release from internalstores it is important to understand the nature and regulation of the endomem-brane Ca2+ release pathways. However, linking the activity of specific endomem-brane Ca2+ channels to [Ca2+]cyt elevations is difficult because they cannot bemonitored electrophysiologically in intact cells. None have yet been character-ized at the molecular level in plants. Obvious homologues of animal inositol (1,4, 5) trisphosphate (InsP3) or ryanodine receptors that mediate InsP3 or cyclicADP ribose (cADPR)-induced Ca2+ release, respectively, have not been identi-fied in theArabidopsisgenome. Nevertheless, there is compelling evidence tosuggest fundamental roles for endomembrane channels based on direct patchclamp recordings, caged release, pharmacology, imaging, and more recently, ge-netic studies. A variety of endomembrane channels that could potentially giverise to cytosolic Ca2+ elevations have been characterized at the physiologicallevel. These include voltage-dependent calcium channels (VDCCs), slow vac-uolar (SV) channels, and InsP3-, and cADPR–sensitive Ca2+ release channels inthe vacuolar membrane (116). Ca2+ release from endoplasmic reticulum (ER)-enriched fractions has also been demonstrated in response to InsP3 (98), cADPR(101), and nicotinic acid adenine dinucleotide phosphate (NAADP) (102), althoughCa2+-permeable ER channels have been monitored directly in only two plant sys-tems (Bryoniatendris andLepidiumroot tips) by incorporation into lipid bilayers(70, 71).

The SV channel has been well studied with respect to its potential role inbringing about Ca2+ release from the vacuole during guard cell signaling. Therehas been considerable discussion about whether this channel can indeed give riseto Ca2+ flux into the cytosol under normal physiological conditions (137); atcytosolic Mg2+ concentrations the activation threshold for this channel is in thephysiological range (−3 to−50 mV, cytosol negative). This channel can potentiallygive rise to Ca2+-induced Ca2+ release (CICR) from the vacuole because its activityalso increases with elevated cytosolic Ca2+. The SV channel is also activated byCaM and the type IIB phosphoprotein phosphatase, calcineurin, which activatesat low concentrations but inhibits at high concentrations. The SV channel is alsoactivated significantly by increased cytosolic pH in the physiological range (7.3 to8.0), potentially providing a link between observed increases in pH in response toABA (11).

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Shaping the Ca2+ Signal: Pumping and Binding!

To act as an effective signal [Ca2+]cyt needs to be maintained at sufficiently lowlevels to allow small influxes through channels to bring about significant changesin [Ca2+]cyt. Adenosine triphosphate (ATP)-fueled pumps (Ca2+-ATPases) andtransporters driven by the electrochemical gradients of other ions, particularlyH+ in higher plants (H+/Ca2+ exchangers), play critical housekeeping roles inmaintaining low resting [Ca2+]cyt in the unstimulated cell. However, they may alsobe important in determining the peak amplitudes and duration of Ca2+ transients(Figure 1). Evidence that they play active roles in shaping Ca2+ signals comes fromthe knowledge that certain classes of pumps and transporters can be regulated byCa2+/CaM, among other factors (see below). However, although the properties oftransporters and pumps have been elucidated in some detail in various plant celltypes, we await concrete evidence to link a specific pump or transporter directlywith a specific Ca2+ signaling event. It is also becoming clear that disrupting theexpression or activity of one type of pump or transporter can lead to compensatorychanges in the activity or expression of other pumps or transporters. However,despite these problems, functions for pumps and transporters in stress signalingand Ca2+ homeostasis are beginning to emerge.

In general Ca2+-ATPases belong to a group of high-affinity pumps whereasH+/Ca2+ exchangers have lower affinity (42, 112, 124). The Ca2+-ATPases can begrouped as type IIA or IIB based on their homologies with animal counterparts.Type IIA ATPases include the ER-type Ca2+ ATPases (ECA) whereas type IIBcontain the autoinhibited Ca2+-ATPases (ACA) (31). Each group contains multiplemembers (4ECAand 10ACAgenes inArabidopsis). Although there is evidence foran ER location for type IIA Ca2+-ATPases in plants (82, 83), plasma membrane,vacuole, and Golgi locations are also evident (38, 42, 105).

Type IIB Ca2+-ATPases in animal cells are targeted to the plasma membranewhereas plant IIB members are also targeted to endomembranes (42, 60). For ex-ample, ACA8 occurs on the plasma membrane (14) but ACA2 has been identifiedwith an ER location (82) and ACA4 occurs on small vacuolar membranes butnot the large central vacuole, which suggests functional diversity within the vac-uolar membrane system in relation to Ca2+ homeostasis or signaling (42). TypeIIA and IIB Ca2+-ATPases show distinctly different modes of regulation. TypeIIB (ACA) members contain an N-terminal autoinhibitory domain (in contrast toanimal counterparts, which contain a C-terminal autoinhibitory domain) and canbe activated directly through binding Ca2+/CaM (56). ACA activity can be inhib-ited by calcium-dependent protein kinase (CDPK) binding and phosphorylation(63). In plants no direct regulation of a type IIA ATPase has been demonstrated,which may suggest a constitutive role in maintaining resting cytosolic Ca2+ levels.However, animal type IIB pumps are highly regulated by an inhibitory subunitphospholamban and possibly also ER lumen Ca2+ content (31).

Physiological and molecular data indicate vacuolar localization of H+/Ca2+

exchangers such as theArabidopsiscation exchanger CAX1, although there is also

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physiological evidence for H+/Ca2+ exchange at other locations such as the plasmamembrane (67), the chloroplast thylakoid membrane (35), and the nuclear envelope(30). CAX transporter activity can be regulated via an N-terminal autoinhibitorydomain. Pittman & Hirschi (111, 112) postulated that this is effected via interactionwith a regulatory protein. However, the N terminal does not interact with CaM(112), unlike the ACA Ca2+-ATPases, and the nature of the regulatory proteinremains unknown.

Functional Roles: Evidence from Disruption Studies

Recent manipulation of expression of both Ca-ATPases and H+/Ca2+ exchangershas begun to shed light on their roles in Ca2+homeostasis. T-DNA insertion mutantsof ECA1(139) showed reduced growth on Ca2+-depleted media. This was the firstknockout of a Ca2+-ATPase gene and provided evidence for a role forECA1 inER Ca2+ homeostasis. Overexpression ofCAX1 in tobacco plants gave rise toincreased vacuolar H+/Ca2+ exchange activity and increased plant Ca2+ levels.CAX1overexpressing plants showed severe Ca2+ deficiency symptoms, probablydue to reduced [Ca2+]cyt levels. These plants also showed increased vacuolar H+

pumping. Knockout ofCAX1(18) led to increased tolerance to both Ca2+deficiencyand abiotic stress and also resulted in decreased vacuolar V-ATPase H+ pumpingactivity. Significantly, these knockouts led to increased tonoplast Ca2+-ATPaseactivity and increased expression of other H+/Ca2+ exchangersCAX3andCAX4.Although evidence for a direct involvement in Ca2+ signaling was not providedin these studies, a possible signaling role was suggested by the impaired hormoneresponses and auxin-regulated gene expression ofcax1mutants.

These results from overexpression and knockout experiments suggest complexinteractions between the activities and expression levels of different pumps andtransporters, and the operation of a highly integrated compensatory network thatmay also include primary H+ pumps. Direct monitoring of Ca2+ transients inplants where specific pump or transporter activity has been manipulated is ea-gerly awaited, and their precise cellular localizations need to be cataloged. In frogoocytes, overexpression of a type IIA Ca2+-ATPase led to increased frequency ofCa2+ spiking (77). Although there is currently no direct evidence for a role forCa2+ pumps or transporters in shaping Ca2+ signals, there is evidence for a role ofa V-type H+-ATPase in determining the patterns of Ca2+ signals.Arabidopsis det3mutants that are defective in a V-type H+ ATPase show disrupted Ca2+ signalingin stomatal guard cells in response to elevated [Ca2+]ext and oxidative stress butnot to ABA or cold (3). The simplest mechanistic interpretation of this result mayrelate to energization of a CAX that is involved in generating stimulus-specific[Ca2+]cyt oscillations.

ER Ca2+ Binding and Homeostasis

Ca2+-binding proteins also play important roles in Ca2+ homeostasis in plants, al-though a direct signaling role is not yet evident. Overexpression of maize

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calreticulin (CRT) in tobacco leads to increased Ca2+ storage in the ER (110).Arabidopsisplants overexpressing CRT show increased resistance to Ca2+ deple-tion, and expression of CRT antisense leads to increased sensitivity to low Ca2+.This suggests that the ER can provide an exchangeable store of Ca2+ for [Ca2+]cyt

homeostasis. Vacuolar Ca2+-binding proteins have also been identified (141), al-though their role in Ca2+ signaling or homeostasis has not been determined.

Mitochondria: Energetic Players in a Dynamic Ca2+ Network

The role(s) of mitochondria in signaling in plants cells are relatively unexplored.There is evidence that they may function to provide a source of Ca2+ in maizesuspension cells in response to anoxia (122), or respond to elevated [Ca2+]cyt byincreasing their Ca2+ content inFucusrhizoid cells (22). Recently, Logan & Knight(84) targeted aequorin to the mitochondrial matrix and recorded average cellular“resting” [Ca2+]mit around 200 nM, approximately double that of [Ca2+]cyt. Treat-ments that caused elevation of [Ca2+]cyt also gave rise to elevations of [Ca2+]mit.However, although [Ca2+]mit elevations were comparable in kinetics (though lowerin amplitude) to those in the cytosol in response to cold and osmotic treatments,significantly different kinetics were observed in response to touch stimuli, and[Ca2+]cyt recovered to resting levels much more rapidly than [Ca2+]mit. In responseto H2O2, [Ca2+]mit elevated to a similar amplitude as [Ca2+]cyt. These results indi-cate separate mechanisms for regulating [Ca2+]mit and [Ca2+]cyt, and existence ofmitochondria-specific Ca2+ signals. Logan & Knight (84) proposed that the ele-vations in [Ca2+]mit might be involved in stimulating dehydrogenases of the TCAcycle, which in turn would stimulate respiration and increased ATP production.Similar mechanisms have been elucidated in animal cells (10, 51).

Mitochondria as Ca2+ Buffers: “Marks and Sparks”

In animal cells, evidence is emerging for a very tight coordination of ER/SR Ca2+

release and uptake by individual mitochondria (51). It is likely that very highly lo-calized [Ca2+]cyt “sparks,” resulting from Ca2+ flux although endomembrane InsP3

or ryanodine receptors, permit targeted delivery of Ca2+ to neighboring mitochon-dria, resulting in transient [Ca2+]cyt elevations in the matrix of individual mito-chondria (“marks”). This points to a high degree of coordination of cytoplasmicand mitochondrial Ca2+ signaling, and highly localized regulation of mitochon-drial function that is coordinated with localized signaling activity of the cytosol.However, there is evidence that mitochondria play an important role in regulatingthe extent of Ca2+cyt signal propagation by highly localized buffering. For example,Malli et al. (89) have shown, using DsRed and yellow chameleon Ca2+ indicatorstargeted to the mitochondria and ER, that localized [Ca2+]cyt elevations in responseto histamine were only evident in mitochondrial-free zones and did not occur closeto mitochondria. This heterogeneity of Ca2+ elevation could allow simultaneousactivation of plasma membrane Ca2+-activated and Ca2+-inhibited channels in thesame cell. Thus, by rapidly removing Ca2+ from the cytosol, mitochondria can

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play an important role in the spatiotemporal patterning of Ca2+ signals. Recentlyit was shown in HeLa cells that mitochondrial uptake required sustained release ofCa2+ from nearby locations on the ER membrane and balanced uptake of Ca2+ intothe ER via the activity of plasma membrane Ca2+ channels (123). Although wecan only speculate that similar mechanisms exist in plants (Figure 2), the data ofLogan & Knight (84) are broadly consistent with results from mammalian cells. Itwill be interesting to determine whether the mean increases in [Ca2+]mit measuredwith aequorin reflect a heterogeneity of [Ca2+

mit] between individual mitochondriain plant cells.

Polarized growing plant cells offer an obvious choice for investigating the role ofmitochondria in [Ca2+]cyt spatial regulation. Regulating mitochondrial movementsin growing pollen tubes and root hairs may be especially significant. The apical[Ca2+]cyt gradient in growing lily pollen tubes is associated with large measuredfluxes of Ca2+ into the tube apex (59). These Ca2+ fluxes show oscillations withthe same frequency as oscillations in the apical [Ca2+]cyt gradient and growth.However, although it is clear that the [Ca2+]cyt gradient at the pollen tube apex isrequired for growth (59), the peak Ca2+ elevation at the growing tube apex lagsbehind the growth peak by approximately 4 s, and the maximum Ca2+ influx atthe tube tip lags behind growth by approximately 15 s. Studies of mitochondriadistribution in pollen tubes show that they are excluded from the extreme apex(28) but they also move with cytoplasmic streaming in a “reverse fountain” pattern(39) that continually delivers and removes mitochondria to the subapical region.Whether these mitochondria play a role in the periodic removal or buffering of[Ca2+]cyt in the pollen tube apex remains to be determined.

PROPAGATION OF PLANT Ca2+ SIGNALS

Experimental demonstration of the components required for propagated intracel-lular release provides compelling evidence that plant cells, like animal cells, cangenerate Ca2+ signals that propagate through the cytosol via intracellular Ca2+

release. Most recorded Ca2+ waves in animal cells fall into the “fast Ca2+ wave”category that propagate within a narrow range of velocities that may reflect thehighly conserved nature of the intracellular propagation mechanisms (64). How-ever, slow Ca2+ waves also occur in animal cells, particularly those that propagatebetween cells and in developing systems (134), and it has been proposed thatboth fast and slow waves propagate via reaction-diffusion mechanisms (64, 134).In plants, very few intracellular propagating waves have been visualized directly,although Campbell et al. (15) showed the existence of intercellular waves that prop-agate slowly through tissue in wholeArabidopsisseedlings. Although [Ca2+]cyt

elevations that appear to propagate as waves have been observed in guard cells,their velocity is approximately ten times slower than typical fast Ca2+ waves (49).So far the only recorded fast Ca2+ wave described in detail in a plant or algalcell is that of the osmotically induced wave in theFucusrhizoid (48). Althoughthis will likely be triggered by activating the mechanosensitive plasma membrane

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channels (127), it propagated by elemental “sparks” through ER-rich regions (48).Fast [Ca2+]cyt responses of higher plants, such as those in response to osmoticstimuli, that are likely to involve intracellular Ca2+ release (125) are potentiallygood candidates for studying plant Ca2+ waves. The question remains of how weexplain the preponderance of relatively slow increases in cytosolic Ca2+ in termsof known mechanisms for Ca2+ elevation in plants. The rate of rise of [Ca2+]cyt

and velocity of propagation in any cell depends on several factors, including thekinetics of receptor activation, rate of production and diffusion of second messen-gers such as InsP3, the rate of permeation of Ca2+ through channels (which alsodepends on their selectivity), the length of time the channels stay open (presum-ably a sustained elevation of Ca2+ requires a sustained increased channel-openprobability), and the regulation of efflux systems. The highly convoluted natureof the ER endomembrane system may allow Ca2+ release sites to be positioned inclose proximity to one another, whereas the reaction-diffusion kinetics along themore linear vacuolar membrane may be significantly different.

Examples of significant deviation from the typical animal reaction-diffusionmodel for Ca2+ waves do exist in plants. For example, a slowly propagating Ca2+

signal occurs in the maize egg during in vitro fertilization (8). In this system, ele-vation of Ca2+ is brought about by a slow wave of Ca2+ influx across the plasmamembrane, which propagates around the egg. Another example is provided by cy-toplasmic droplets from the giant algaNitella that display Ca2+ spikes in responseto hypo-osmotic treatment. The most likely explanation for this phenomenon is theactivation of mechanosensitive Ca2+-permeable endomembrane channels (69).

[CA2+]cyt OSCILLATIONS AND TRANSIENTS

In animal cells, stimulus-induced [Ca2+]cyt oscillations are well documented andtheir mechanisms of generation and the potential for the information to be encodedin the frequency of [Ca2+]cyt oscillation was recently reviewed (10). [Ca2+]cyt oscil-lations and transients also occur in plant cells with, to the best of our knowledge,the first report being IAA-stimulated [Ca2+]cyt oscillations in maize coleoptiles(37). The frequency, period, and amplitude of [Ca2+]cyt oscillations vary amongdifferent cell types and in response to different stimuli (36), and the possibility that[Ca2+]cyt oscillations might encode information has been discussed (36, 93, 114,116, 117), as have mechanisms for generating [Ca2+]cyt oscillations (9, 55). Todiscuss how [Ca2+]cyt oscillations and transients are generated, and their possibleroles in plant cell signaling, we focus on NF signaling in legume root hair cellsand stimulus-response coupling in guard cells.

Calcium Oscillations in Legume Root Hair NF Signaling

NFs are bacterial lipochitooligosaccharide signals that play an important role in theearly stages of nodule development. They stimulate root hair cells of susceptibleplants to undergo morphological changes that lead to the entrapment and invasion

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of the bacteria. The role of NFs in this process has been comprehensively reviewed(16, 46, 81). Among the NFs’ induced changes in susceptible root hair are increasesin [Ca2+]cyt, which include oscillations. There are two phases to NF-induced roothair [Ca2+]cyt elevations. The first phase is rapid. InMedicago truncatulait consistsof a rapid spike followed by a sustained [Ca2+]cyt increase or plateau that lasts for3–4 min. Approximately 10 mins after NF application, the second phase occurs,which consists of “calcium spiking” in the nuclear region (7, 119). A typical spike inM. sativaor M. truncatulaconsists of a rapid increase of [Ca2+]cyt (approximately500 nM) followed by a more gradual return to resting levels. This produces anasymmetric peak with a sharp rising phase and a slower recovery. The durationof the whole spike from onset to return to resting [Ca2+]cyt is between 30–120 s,and the frequency of occurrence is typically approximately 1 min (7, 33). BecauseCa2+ spiking is an example of an oscillatory phenomenon we refer to it as [Ca2+]cyt

oscillation.

Generation of NF-Induced [Ca2+]cyt Oscillationsin Root Hair Cells

In M. truncatula, (34) NF-induced [Ca2+]cyt oscillations were inhibited separatelyby 50µM 2-APB and 50 mM caffeine, which are putative antagonists of intracel-lular store-mediated Ca2+ release. 5µM cyclopiazonic acid (CPA), an inhibitorof type IIA calcium ATPases in plants, 10µM 2,5-di-(t-butyl)-1,4-hydroquinone(BHQ), an inhibitor of mammalian sarco(endo)plasmic reticulum calcium AT-Pases (SERCA), and the phosphatidyl inositol specific phospholipase C (PI-PLC)inhibitor U73122 (20µM) also inhibited NF-induced root hair [Ca2+]cyt oscil-lations. Because both U73122 (121) and CPA (83) inhibit plant enzymes thedata obtained by Engstrom and colleagues support a role for PI-PLC and TypeIIA Calcium ATPases in NF generation of [Ca2+]cyt oscillations. No effects on[Ca2+]cyt oscillations were observed when the root hairs were treated with the pu-tative plasma membrane calcium channel blockers verapamil (100µM) and LaCl3(1 mM) (34). However, it might be unwise to rule out the involvement of plasma-membrane Ca2+ channels because it is possible that plants possess Ca2+ channelsin the plasma membrane that are La3+- and verapamil-insensitive.

Genetic approaches have also been used to identify components involved in theNF signaling pathway. Walker and colleagues (132) investigated the occurrence ofNF-induced [Ca2+]cyt oscillations in the pea nodulation defective mutantssym8,sym10, andsym19. In all three of these mutants NF-induced [Ca2+]cyt oscillationswere blocked, suggesting that the products of all three genes are upstream ofand required for the generation of NF-induced [Ca2+]cyt oscillations. Long andcolleagues (131) reached similar results and conclusions using thedmi1anddmi2nodulation mutants ofM. truncatula.

Shaw & Long (119) recently investigated whether the first rapid NF-induced[Ca2+]cyt increase is required for generating NF-induced [Ca2+]cyt oscillations.When they challengedM. truncatulawith 10 nM NF they observed the familiar

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two-phase Ca2+ response. However, when they applied 1 nM NF they did not ob-serve any rapid NF-induced [Ca2+]cyt increase, although they saw the NF-induced[Ca2+]cyt oscillations. These results suggest that generating [Ca2+]cyt oscillationsdoes not require the first NF-induced [Ca2+]cyt elevation.

Stimulus-Induced Oscillations and Transientsof Guard Cell [Ca2+]cyt

Stimulus-induced increases in guard cell [Ca2+]cyt are part of signal transductionnetwork(s) responsible for coupling ABA and other external signals to alterationsin stomatal aperture (11, 12, 57, 93, 117). Stimulus-induced oscillations in guardcell [Ca2+]cyt were observed whenCommelina communis, Fura-2-loaded guardcells were bathed in 0.1 mM [Ca2+]ext (symmetrical oscillations with an averageamplitude of approximately 430 nM and a mean period of 8.3 min) or 1.0 mM[Ca2+]ext (asymmetrical oscillations with an amplitude of 625–850 nM and a meanperiod of 13.6 min) (94). Oscillations in guard cell [Ca2+]cyt were more extensivelyinvestigated inArabidopsisusing cameleon [Ca2+]cyt technology (5). Using thisapproach (3), 1 mM [Ca2+]ext induced oscillations with an average amplitude ofapproximately 160 nM and an average period of 2.6 min, whereas 10 mM [Ca2+]ext

induced oscillations with an average amplitude of 1020 nM and a period of 6.6min. In a separate study, 10 mM [Ca2+]ext induced [Ca2+]cyt oscillations with aperiod of approximately 8.2 min (2). The periods of the [Ca2+]ext-induced oscilla-tions reported using the cameleon technique correlate well with data obtained inthe same species using the fluorescent ratiometric [Ca2+]cyt indicator Fura-2 (4).ABA can also induce [Ca2+]cyt oscillations inC. communis(121) andArabidopsis.ABA-induced [Ca2+]cyt oscillations have been reported in differentArabidopsisecotypes (Col, Ler, WS), and when they occur they are either in the form of regu-lar oscillations with a magnitude of about 500 nM and a period of approximately7.8 min or have no obvious periodicity. Schroeder and colleagues refer to theseirregular increases as “transients” and we adopt this terminology here (2, 6, 61, 66,72, 75, 76). Similar to earlier work (47, 90, 91), not all guard cells responded toABA with detectable increases in [Ca2+]cyt. Transients were also observed whenguard cells were challenged with a yeast elicitor (72), 100µM H2O2, and cold (3).

Generating Stimulus-Induced Oscillationsand Transients in Guard Cell [Ca2+]cyt

Ca2+ influx across the plasma membrane and release from internal stores bothcontribute to the generation of [Ca2+]cyt oscillations. InC. communisthe results ofmanganese quench experiments and application of verapamil suggests that plasmamembrane calcium-permeable channels participate in the generation of [Ca2+]ext-induced [Ca2+]cyt oscillations (94). A similar conclusion was reached from experi-ments in whichArabidopsisguard cells bathed in Ca2+-free media failed to exhibit[Ca2+]cyt transients when stimulated by a yeast elicitor or ABA (72).

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Although it is likely that more than one guard cell plasma membrane calciumchannel is involved in the generation of [Ca2+]cyt oscillations (53, 72), most at-tention has focused on an HACC inVicia faba(53) andArabidopsis(where it isknown as ICa) (108). InVicia the open probability of the HACC is increased byABA, the concentration of extracellular divalent cations (52, 53), and inhibitors ofType 1 and 2A phosphoprotein phosphatases (73). This latter result, coupled withthe observation that ATP hydrolysis was required for channel activity, led Kohler& Blatt (73) to propose that phosphorylation of sites on or close to the channelfacilitates opening. Increasing [Ca2+]cyt from 200 nM to 2µM reduces the openprobability of the channel and provides a mechanism for feedback inhibition ofcalcium influx (53). InArabidopsisthe HACCs are permeable to several cations,including Mg2+, require cytosolic NAD(P)H, and are involved in the generation of[Ca2+]cyt increases by H2O2, ABA (108), and pathogenic elicitors (72). Schroederand colleagues proposed a model for ABA action onArabidopsisguard cells thatinvolves ABA stimulating H2O2 production and the resulting H2O2 increasingthe open probability of HACC (108). The suggestion that ABA action involvesH2O2 is corroborated by investigations carried out inVicia guard cells (144), andSchroeder and colleagues further suggest that ABA and a fungal elicitor signalingpathways converge on HACC (72). They also place the Type 2C phosphopro-tein phosphatases encoded by theABI1-1 andABI2-1 genes upstream of ICa inABA signaling (4, 99). The most recent work shows that although ABA-induced[Ca2+]cyt transients were not abolished, they were significantly reduced inAra-bidopsisNADPH oxidase catalytic subunit mutants (atrbohD/F double mutant),supporting a role for HACCs and ROS inArabidopsisguard cell ABA signaling(76). However, the Blatt laboratory’s (74) recent work inVicia questions whetherH2O2 really is a critical second messenger in guard cell ABA signaling. Kohlerand colleagues (74) found that inVicia although H2O2 regulated two key cellularcomponents involved in guard cell turgor control (HACC and inward rectifying K+

channels) in a manner reminiscent of ABA, and H2O2stimulates [Ca2+]cyt increasesin C. communis(92) andArabidopsis(108) it also depresses guard cell outward-rectifying K+ channels (74). This latter action is the direct opposite of what hap-pens in guard cell ABA signaling. Ignoring possible species-specific differencesas an explanation for these apparently conflicting sets of data, it seems likely that,as guard cell ABA signaling is partially disrupted in theArabidopsisAtrbohD/Fdouble mutant (76), H2O2 and possibly other ROS are involved in some but notall aspects of guard cell ABA signaling. Finally, there is evidence that Type 2Aphosphoprotein phosphatase (PP2A) activity is involved in the regulation of ABA-induced [Ca2+]cyt transients. In the loss of functionrcn1 mutant, which encodesthe regulatory subunit of a guard cell–expressed PP2A, stomatal responsivenessto ABA was greatly reduced, as was the probability of ABA-induced increases in[Ca2+]cyt (75).

There are also data indicating that release of Ca2+ from internal stores con-tributes to the generation of stimulus induced [Ca2+]cyt transients and oscillations.There is evidence pointing to a role for the PI-PLC–InsP3 system in generating

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[Ca2+]cyt oscillations and transients in guard cell ABA signaling. In bothC. com-munisandArabidopsisguard cells the pharmacological PI-PLC inhibitor U73122interferes with ABA–induced alterations in stomatal aperture and ABA-induced[Ca2+]cyt oscillations (121) and transients (72). U73122 also interferes with ABA-induced K+ efflux from guard cells (88).

Is it possible to place PI-PLC in the sequence of events leading to the gener-ation of ABA-induced [Ca2+]cyt oscillations and transients? Hunt and colleagues(62) present a model in which a primary ABA-generated increase in [Ca2+]cyt acti-vates guard cell PI-PLC, which then generates InsP3, leading to Ca2+ release frominternal stores. In this model PI-PLC–InsP3 amplifies the primary ABA-inducedincrease in [Ca2+]cyt and contributes to the rising phase of the oscillation. For thissystem to generate a series of oscillations, a separate mechanism for repriming[Ca2+]cyt to a level whereby it could activate PI-PLC would be required after thecompletion of each oscillation and the return of [Ca2+]cyt to resting levels. ABA-induced [Ca2+]cyt influx through the plasma membrane HACC (discussed above)might be a prime candidate to fulfill such a role. Although PI-PLC plays a role inthe generation of guard cell ABA-induced [Ca2+]cyt oscillations it is not apparentlyinvolved in [Ca2+]ext-induced [Ca2+]cyt oscillations inC. communis(121).

Just as MacRobbie (88) showed that the mechanism used to generate ABA-induced increases in [Ca2+]cyt depends on the concentration of applied ABA, it ispossible that other Ca2+-mobilizing messengers might be involved in ABA sig-naling. In guard (78) and other plant cells (1) cADPR induces Ca2+ release fromvacuoles. There is evidence that cADPR might be involved in the generation ofABA-induced [Ca2+]cyt oscillations and transients. Microinjecting guard cells withcADPR induces increases in guard cell [Ca2+]cyt that include oscillations (78) andnicotinamide, which blocks cADPR synthesis and interferes with ABA-induced[Ca2+]cyt calcium transients (72). However, in contrast to the situation with thePI-PLC–InsP3 system, it is unlikely that cADPR is involved in Ca2+ signal ampli-fication or the phenomenon of CICR because cADPR-gated currents downregulateover the range of guard cell [Ca2+]cyt elevated by cADPR microinjection (78). Thislatter finding suggests that cADPR might be involved in the initial priming phaserequired in the generation of [Ca2+]cyt oscillations and transients. Understandingthe contribution of cADPR to stimulus-induced [Ca2+]cytoscillations and transientswill be greatly facilitated with the identification of the enzyme systems responsiblefor its production and its intracellular receptors.

Another molecule capable of generating increases in [Ca2+]cyt in animal andplant cells is sphingosine-1-phosphate (S1P) (138). When applied toC. communisguard cells 6µM S1P induced asymmetrical [Ca2+]cyt oscillations with a periodof 3.8 min and peaks of up to 50 nM above resting levels of [Ca2+]cyt. 50 nMSIP induced symmetrical [Ca2+]cyt oscillations with a shorter period (2.8 min)but greater peak height (100 nM above resting levels of [Ca2+]cyt) (103). Recentwork from the Assmann lab (23) resulted in the construction of a model for theinvolvement of S1P in guard cell ABA signaling, in which ABA activates sphin-gosine kinase inArabidopsisguard cells resulting in the production of elevated

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SIP. The SIP then interacts with an unidentified receptor followed by interactionswith the product of theGPA1gene and generation of [Ca2+]cyt increases (23). Cur-rently, we do not know whether other Ca2+-mobilizing messengers such as inositolhexakisphosphate (InsP6) (79) and NAADP (101) are involved in generating[Ca2+]cyt oscillations or transients.

As with NF signaling in root hair cells, genetic approaches have also resultedin significant insights into the generation of [Ca2+]cyt transients and oscillations.Schroeder and coworkers investigated whether the products of genes involved inABA signaling are involved in the generation of [Ca2+]cytoscillations and transientsby examining ABA-induced [Ca2+]cyt elevations in the correspondingArabidopsismutants. TenµM ABA induced [Ca2+]cyt transients in 86% of wild-type guardcells but failed to induce [Ca2+]cyt increases in anyabi2-1 guard cells (6). Theresult suggests that the product of theABI2-1 gene (a type 2C phosphoproteinphosphatases) is required for the generating ABA-induced [Ca2+]cyt transients.Stomata of theArabidopsis rcn1mutant are less sensitive to ABA. 14% ofrcn1guard cells showed two or more [Ca2+]cyt transients when treated with 5 uM ABA,in contrast to the 44% of wild-type stomata that exhibited two or more [Ca2+]cyt

transients. The conclusion from this work is that the type 2A phosphoproteinphosphatase encoded by theRCN1gene is also involved in the generation of ABA-induced [Ca2+]cyt transients (75). Similar experimental approaches have shown thattheERA1gene product (encoding a farnesyltransferase) (6), the catalytic subunitsof NADPH oxidase encoded byAtRBOHDandAtRBOHFgenes (76), the productof theGCA2gene (2), and the mRNA cap binding protein encoded byABH1(61)can influence ABA-induced [Ca2+]cyt transients.

Significance of [Ca2+]cyt Oscillations and Transientsin Plant Cell Signaling

In animal cells experimental evidence supports the hypothesis that informationis encoded in [Ca2+]cyt oscillations (10). In plant cells it has been suggested that[Ca2+]cyt oscillations and transients might encode information that helps to dictatethe nature of the downstream response (36, 55, 93, 114, 116, 117). We focus onthree experimental systems, pollen tube growth, NF signaling in root hair cells, andstimulus-response coupling in guard cells, to assess the physiological significanceof [Ca2+]cyt oscillations and transients in plant cell signaling.

The role of [Ca2+]cyt in the control of pollen tube growth was the subject of arecent comprehensive review (59). However, in brief, the growing pollen tube isan endogenous oscillator and accompanying growth oscillations are oscillationsin [Ca2+]cyt, pHcyt, membrane potential, exo and endocytosis, cell wall thickness,and the fluxes of extracellular Ca2+, H+, K+, and Cl−. Because it has long beenrecognized that Ca2+ is essential for pollen tube growth, an attractive propositionwas that [Ca2+]cyt oscillations might be the coordinator or pacemaker controllinggrowth. Current evidence suggests that this is unlikely for two reasons. First,[Ca2+]cyt oscillations follow rather than anticipate changes in pollen tube growthrate (95), and second, pollen tube [Ca2+]cyt oscillations have been observed in

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the absence of growth (96). Accordingly, the current situation in this system isthat there are well-characterized [Ca2+]cyt oscillations but their physiological roleremains to be established.

Currently, a similar situation holds for NF-induced [Ca2+]cyt oscillations inlegume root hairs, although this may change in the near future as a result of workby Shaw & Long (119). They showed that by judicial choice of NF concentrationit is possible to induce [Ca2+]cyt oscillations in the absence of the first NF-induced[Ca2+]cyt elevation. This seems to provide the ideal experimental system to addressthe question of the physiological role of [Ca2+]cyt oscillations. Given the perinu-clear location of the [Ca2+]cyt oscillations, an obvious experiment to conduct wouldbe a comparative analysis of 1 nM NF- (induces only [Ca2+]cyt oscillations) and10 nM NF- (induces first-phase [Ca2+]cyt elevations and second-phase oscillations)induced gene expression.

Currently, the most compelling evidence that [Ca2+]cyt oscillations and tran-sients might be responsible for encoding specific signaling information comes fromguard cells. McAinsh and coworkers showed that that it is possible to correlate thepattern of [Ca2+]cyt oscillation with both the strength of the external stimulus andthe extent of the final response (stomatal closure) in [Ca2+]ext signaling (94). How-ever, these data revealed a correlation rather than provided evidence of a causalrelationship between [Ca2+]cyt oscillations and the control of stomatal aperture.Allen and colleagues (3) established a much stronger link between [Ca2+]cyt oscil-lations and the control of stomatal aperture using theArabidopsis det3mutant thatexhibits a 60% reduction in expression of the C-subunit of the V-type H+-ATPase.The authors hypothesized that a reduction in V-type H+-ATPase activity might re-sult in reduced Ca2+ sequestration into internal stores, and this in turn might affectstimulus-induced [Ca2+]cyt oscillations. Whatever the precise effects of the reduc-tion in expression of theDET3gene the authors observed, in contrast to wild-typeguard cells, thatdet3guard cells failed to exhibit [Ca2+]cyt oscillations and reduc-tions in guard cell turgor in response to 1 and 10 mM [Ca2+]ext. When total guardcell [Ca2+]ext-stimulated [Ca2+]cyt was integrated over a 30-minute period it wassignificantly higher in thedet3mutant than in wild type. This result clearly showsthat the reason thedet3mutant failed to close its stomata in response to [Ca2+]ext

was not due to an inability to increase [Ca2+]cyt, but rather due to the failure togenerate [Ca2+]cyt oscillations that seemed to underlie the phenotype. Allen andcolleagues also showed that experimentally induced [Ca2+]cyt oscillations broughtabout (rescued) closure in thedet3mutant. These data provide strong evidencethat [Ca2+]cyt oscillations are required in the signaling pathway leading to stomatalclosure.

With evidence for the importance of [Ca2+]cyt oscillations in guard cell sig-naling the next question to address is what their specific role is in the controlof stomatal aperture. One possibility is that [Ca2+]cyt oscillations are responsi-ble for holding the guard cell in a steady low-turgor state while protecting itfrom the presumed deleterious effects of long-term exposure to elevated [Ca2+]cyt

(94). However, although this might be beneficial to the guard cell it is not a re-quirement in guard cell signaling because there are examples of closure-inducing,

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[Ca2+]cyt-elevating stimuli that do not bring about [Ca2+]cyt oscillations such asABA (47, 91, 99), CO2 (133), and H2O2 (92). Instead, Schroeder and colleaguespropose that [Ca2+]cyt controls stomatal aperture by two mechanisms, based onexperiments carried out on theArabidopsis gca2mutant. They distinguish be-tween short-term stomatal closure that occurs in response to elevated [Ca2+]cyt andlong-term (steady-state) closure, the degree of which is programmed or controlledby [Ca2+]cyt oscillations within a defined range of frequency, transient number,duration, and amplitude (2).

However, some additional factors need to be considered before any generaliza-tions can be made about the role of [Ca2+]cyt oscillations in guard cells. Perhapsthe most obvious issue to consider is that ABA-, CO2-, and H2O2-induced stom-atal closure can occur in the absence of [Ca2+]cyt oscillations (47, 90–92, 133).It is possible that the experiments in these studies were not long enough for thestomata to reach steady state and thus failed to exhibit [Ca2+]cyt oscillations. Afurther complication concerns the occurrence of spontaneous [Ca2+]cyt transientsin guard cells (72). The lack of closure in the stomata exhibiting spontaneous[Ca2+]cyt transients is presumably due to the fact that the period of the sponta-neous [Ca2+]cyt transients was shorter than ABA-induced [Ca2+]cyt oscillations andtransients associated with closure. However, there are some difficulties with thisexplanation because 10 mM [Ca2+]ext induces [Ca2+]cyt oscillations with a similarperiod to the spontaneous [Ca2+]cytoscillations and 10 mM [Ca2+]extcloses stomata(3, 4).

In summary, [Ca2+]cyt oscillations and transients are found in different plantcells, and their mechanism of generation and possible physiological significancehave been the subjects of considerable scrutiny in three well-studied plant signalingsystems. Although we are learning much about the origin of [Ca2+]cyt oscillationsand transients, pinning down their physiological role has proved more difficult. Todate, the clearest evidence that stimulus-induced [Ca2+]cyt oscillations (as opposedto stimulus-induced nonoscillatory [Ca2+]cyt elevations) are required during sig-naling comes from the work performed on thedet3mutant (3). However, with theidentification of new signaling mutants and ways of manipulating Ca2+ homeosta-sis it is likely that we are poised to learn much more about the possible functionsof [Ca2+]cyt oscillations and transients in plant cell signaling.

CONCLUSIONS AND FUTURE PERSPECTIVES

It was recently proposed, on the basis of a well-argued case, that many (but notall) Ca2+ signals act as simple on-off binary switches and thus the Ca2+ signalis an unlikely mechanism for ensuring fidelity in plant signaling systems (118).However, as Scrase-Field & Knight (118) acknowledge and we discuss here, theguard cell provides examples of where Ca2+ signals do appear to encode theinformation for defining the outcome of responses (2, 3). The question that wenow need to address is whether the guard cell is unique in using the Ca2+ signal

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to help provide fidelity in stimulus-response coupling, or whether it exists in othercells but we have failed to detect it so far. Unless the symplastic isolation of matureguard cells is a factor, there is no obvious reason why stomata should have evolveda unique method of signaling.

Guard cells have provided important insights into signaling because it is possibleto investigate the effects of different stimuli on Ca2+ dynamics at the single-celllevel while monitoring a robust readout (changes in stomatal aperture). When theSchroeder lab combined the experimental advantages of the guard cell with the useof mutants the definitive evidence that Ca2+ signals can encode information wasobtained. Before we conclude that information-encoding Ca2+ signals are uniqueto guard cells we need to thoroughly investigate whether they occur in other celltypes. In this respect, NF signaling holds great promise, especially if a robustreadout, such as an alteration in gene expression, can be identified similar to theway Shaw & Long (119) were able to experimentally isolate NF-induced [Ca2+]cyt

oscillations from other NF-induced [Ca2+]cyt elevations.The technology for addressing whether the binary switch or the Ca2+ signal

model predominates in plant cell signaling is advancing on several fronts. In thisreview we identify a range of targets for potential disruption and modification. Theavailability of mutants displaying lesions in these targets is increasing and shouldenable investigation of the effect of manipulating Ca2+ signals on stimulus-inducedresponses. Significant advances are also being made in the use of FRET-basedCa2+ indicators such as cameleons and pericams (100, 142) that can be expressedin a targeted manner. These advances in reporter technology are being matchedby fast improving camera– and scanning-based imaging technology, and severaloptions are now available for monitoring Ca2+ from total internal reflection flu-orescence (TIRF) for near-membrane fast events in wall-free cells to multipho-ton microscopy for deeper penetration and improved signal-to-noise ratio (39) inwhole tissues and plants. Combining these techniques with robust cellular or tissueresponses will allow the global significance of Ca2+ signals to be assessed. Over-all, the field is poised for exciting developments and insights over the next fewyears.

ACKNOWLEDGMENTS

The authors acknowledge the support of the BBSRC and NERC for funding sig-naling research in their laboratories.

NOTE ADDED IN PROOF

Recently, a specific receptor for apoplastic Ca2+ that is expressed in guard cellsand is required for oscillatory elevations in guard cell Ca2+ and stomatal closurein response to elevated [Ca2+

ext] was cloned fromArabidopsis(53a). This suggestsa specific external messenger role for [Ca2+

ext]. It will be interesting to determinethe downstream components of this signal transduction pathway.

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The Annual Review of Plant Biologyis online at http://plant.annualreviews.org

LITERATURE CITED

1. Allen CG, Muir SR, Sanders D. 1995.Release of Ca2+ from individual plantvacuoles by both InsP3 and cyclic ADP-ribose.Science268:735–37

2. Allen GJ, Chu SP, Harrington CL, Schu-macher K, Hoffman T, et al. 2001. Adefined range of guard cell calcium oscil-lation parameters encodes stomatal move-ments.Nature411:1053–57

3. Allen GJ, Chu SP, Schumacher K, Shi-mazaki CT, Vafeados D, et al. 2000. Al-teration of stimulus-specific guard cellcalcium oscillations and stomatal clos-ing in Arabidopsis det3mutant.Science289:2338–42

4. Allen GJ, Kuchitsu K, Chu SP, MurataY, Schroeder JI. 1999. Arabidopsisabi1-1 and abi2-1 phosphatase mutations re-duce abscisic acid-induced cytoplasmiccalcium rises in guard cells.Plant Cell11:1785–89

5. Allen GJ, Kwak JM, Chu SP, Llopis J,Tsien RY, et al. 1999. Cameleon calciumindicator reports cytoplasmic calcium dy-namics in Arabidopsis guard cells.PlantJ. 19:735–47

6. Allen GJ, Murata Y, Chu SP, Nafisi M,Schroeder JI. 2002. Hypersensity of ab-scisic acid-induced cytosolic calcium in-creases in the Arabidopsis farnesyltrans-ferase mutantera1-2.Plant Cell14:1649–62

7. Deleted in proof8. Antoine AF, Faure J-E, Cordiero S, Du-

mas C, Rougier M, Feijo JA. 2000. Acalcium influx is triggered and propa-gates in the zygote as a wavefront duringin vitro fertilization of flowering plants.Proc. Natl. Acad. Sci. USA97:10643–48

9. Bauer CS, Plieth C, Bethmann B, PopescuO, Hansen U-P, et al. 1998. Strontium-induced repetitive calcium spikes in aunicellular green alga.Plant Physiol.117:545–47

9a. Ben Amor B, Shaw SL, Oldroyd GED,Maillet F, Penmetsa RV, et al. 2003. TheNFP locus ofMedicago truncatulacon-trols an early step of Nod factor signaltransduction upstream of a rapid calciumflux and root hair deformation.Plant J.34:495–506

10. Berridge MJ, Bootman MD, RoderickHL. 2003. Calcium signalling: dynamics,homeostasis and remodelling.Nat. Rev.Mol. Cell Biol.4:517–29

11. Blatt MR. 2000. Cellular signaling andvolume control in stomatal movements inplants.Annu. Rev. Cell Dev. Biol.16:221–41

12. Blatt MR. 2000. Ca2+ signalling andcontrol of guard-cell volume in stom-atal movements.Curr. Opin. Plant Biol.3:196–204

13. Blume B, Nurnberger T, Nass N, ScheelD. 2000. Receptor-mediated increase incytoplasmic free calcium required for ac-tivation of pathogen defense in parsley.Plant Cell12:1425–40

14. Bonza MC, Morandini P, Luoni L, GeislerM, Palmgren MG, De Michelis MI. 2000.At-ACA8 encodes a plasma membrane-localized calcium-ATPase ofArabidop-sis with a calmodulin-binding domain atthe N terminus.Plant Physiol.123:1495–96

15. Campbell AK, Trewavas AJ, Knight MR.1996. Calcium imaging shows differentialsensitivity to cooling and communicationin luminous transgenic plants.Cell Cal-cium19:211–18

16. Cardenas L, Holdaway-Clarke TL,Sanchez F, Quinto C, Feijo JA, et al.2000. Ion changes in legume root hairsresponding to Nod factors.Plant Physiol.123:443–51

17. Carroll AD, Moyen C, Van Kesteren P,Tooke F, Battey NH, Brownlee C. 1998.Calcium, GTP and annexins modulate

Ann

u. R

ev. P

lant

Bio

l. 20

04.5

5:40

1-42

7. D

ownl

oade

d fr

om a

rjou

rnal

s.an

nual

revi

ews.

org

by C

NR

S-m

ulti-

site

on

12/2

8/07

. For

per

sona

l use

onl

y.



exocytosis from maize root protoplasts.Plant Cell10:1267–76

18. Cheng NH, Pittman JK, Barkla BJ, Shi-gaki T, Hirschi KD. 2003. The Arabidop-sis cax1 mutant exhibits impaired ionhomeostasis, development and hormonalresponses, and reveals interplay amongvacuolar transporters.Plant Cell15:347–64

19. Cheng SH, Willmann MR, Chen HC,Sheen J. 2002. Calcium signaling throughprotein kinases. The Arabidopsis calcium-dependent protein kinase gene family.Plant Physiol.129:469–85

20. Clark GB, Dauwalder M, Roux S. 1994.Immunolocalization of an annexin-likeprotein in corn.Adv. Space Res.14:341–46

21. Clark GB, Sessions A, Eastburn DJ, RouxSJ. 2001. Differential expression of mem-bers of the annexin multigene family inArabidopsis.Plant Physiol.126:1072–84

22. Coleho SM, Taylor AR, Ryan KP, Sousa-Pinto I, Brown MT, Brownlee C. 2002.Spatiotemporal patterning of reactiveoxygen production and Ca2+ wave prop-agation inFucusrhizoid cells.Plant Cell14:2369–81

23. Coursol S, Fan L-M, Le Stunff H, SpiegelS, Gilroy S, Assmann SM. 2003. Sphin-golipid signalling in Arabidopsis guardcells involves heterotrimeric G proteins.Nature423:651–54

24. Davenport R. 2001. Glutamate receptorsin plants.Ann. Bot.90:549–57

25. Demidchik V, Bowen HC, Maathuis FJM,Shabala SN, Tester MA, et al. 2002.Arabidopsis thalianaroot non-selectivecation channels mediate calcium uptakeand are involved in growth.Plant J.32:799–808

26. Demidchik V, Davenport RJ, Tester M.2002. Nonselective cation channels inplants.Annu. Rev. Plant Biol.53:67–107

27. Dennison KL, Spalding EP. 2000.Glutamate-gated calcium fluxes inArabidopsis. Plant Physiol.124:1511–14

28. Derksen J, Rutten T, Lichtscheidl IK,

Dewin A, Pierson ES, Rongen G. 1995.Quantitative analysis of the distributionof organelles in tobacco pollen tubes-implications for exocytosis and endocy-tosis.Protoplasma188:267–76

29. Dingledine R, Borges K, Bowie D,Traynelis SF. 1999. The glutamate recep-tor ion channels.Pharm. Rev.51:7–61

30. Downie L, Priddle J, Hawes C, Evans DE.1998. A calcium pump at the higher plantnuclear envelope?FEBS Lett.429:44–48

31. East JM. 2000. Sarco(endo)plasmicreticulum calcium pumps: recent ad-vances in our understanding of structure/function and biology.Mol. Membr. Biol.17:189–200

32. Ehrhardt DW, Atkinson EM, Long SR.1992. Depolarization ofAlfalfa root hairplasma membrane potential byRhizobiumnodulation signals.Science 256:998–1000

33. Ehrhardt DW, Wais R, Long SR. 1996.Calcium spiking in plant root hairs re-sponding to rhizobium nodulation signals.Cell 85:673–81

34. Engstrom EM, Ehrhard DW, Mitra RM,Long SR. 2002. Pharmacological analysisof Nod factor-induced calcium spiking inMedicago truncatula. Evidence for the re-quirement of Type IIA calcium pumps andphosphoinositide signalling.Plant Phys-iol. 128:1390–401

35. Ettinger WF, Clear AM, Fanning KJ, PeckML. 1999. Identification of a Ca2+/H+ an-tiport in the plant chloroplast thylakoidmembrane.Plant Physiol.119:1379–85

36. Evans NH, McAinsh MR, Hethering-ton AM. 2001. Calcium oscillations inplants. Curr. Opin. Plant Biol. 4:415–20

37. Felle HH. 1988. Auxin causes oscilla-tions of cytoplasmic free calcium andpH in Zea mayscoleoptiles.Planta 174:495–99

38. Ferrol N, Bennett AB. 1996. A singlegene may encode differentially localizedCa2+-ATPases in tomato.Plant Cell8:1159–69

Ann

u. R

ev. P

lant

Bio

l. 20

04.5

5:40

1-42

7. D

ownl

oade

d fr

om a

rjou

rnal

s.an

nual

revi

ews.

org

by C

NR

S-m

ulti-

site

on

12/2

8/07

. For

per

sona

l use

onl

y.



39. Fiejo J, Moreno N. 2003. Imaging plantcells by two-photon excitation.Proto-plasma.In press

40. Foreman J, Demidchik V, Bothwell JH,Mylona A, Miedema H, et al. 2003. Reac-tive oxygen species produced by NADPHoxidase regulate plant cell growth.Nature422:442–46

41. Furuichi T, Cunningham KW, MutoS. 2001. A putative two pore channelAtTPC1 mediates Ca2+ flux in Arabidop-sisleaf cells.Plant Cell Physiol.42:900–5

42. Geisler M, Axelse KB, Harpe JF, Palm-gren MG. 2000. Molecular aspectsof higher plant P-type Ca2+-ATPases.Biochim. Biophys. Acta1465:52–78

43. Gelli A, Blumwald E. 1997. Hyper-polarization-activated Ca2+-permeablechannels in the plasma membrane oftomato cells.J. Membr. Biol.155:35–45

44. Gelli A, Higgins VJ, Blumwald E. 1997.Activation of plant plasma membraneCa2+-permeable channels by race-specificfungal elicitors.Plant Physiol.113:269–79

45. Gerke V, Moss SE. 2002. Annexins:from structure to function.Physiol. Rev.82:331–71

46. Geurts R, Bisseling T. 2002. Rhizo-bium nod factor perception and signalling.Plant Cell14:S239–49

47. Gilroy S, Fricker MD, Read ND, Tre-wavas AJ. 1991. Role of calcium in signaltransduction of Commelina guard cells.Plant Cell3:333–43

48. Goddard H, Manison NFH, Tomos D,Brownlee C. 2000. Elemental propagationof calcium signals in response-specificpatterns determined by environmentalstimulus strength.Proc. Natl. Acad. Sci.USA97:1932–37

49. Grabov A, Blatt MR. 1998. Membranevoltage initiates Ca2+ waves and poten-tiates Ca2+ increases with abscisic acidin stomatal guard cells.Proc. Natl. Acad.Sci. USA95:4778–83

50. Grimaldi M, Maratos M, Verma A. 2003.Transient receptor potential channel acti-

vation causes a novel form of [Ca2+] os-cillations and is not involved in capacitita-tive Ca2+ entry in glial cells.J. Neurosci.23:4737–45

51. Hajnoczky G, Csordas G, Yi M. 2002.Old players in a new role: mitochondria-associated membranes, VDAC, and ryan-odine receptors as contributors to calciumsignal propagation from the endoplasmicreticulum to the mitochondria.Cell Cal-cium32:363–77

52. Hamilton DWA, Hills A, Blatt MR. 2001.Extracellular Ba2+ and voltage interact togate Ca2+ channels at the plasma mem-brane of stomatal guard cells.FEBS Lett.491:99–103

53. Hamilton DWA, Hills A, Kohler B, BlattMR. 2000. Ca2+ channels at the plasmamembrane of stomatal guard cells are ac-tivated by hyperpolarization and abscisicacid.Proc. Natl. Acad. Sci. USA97:4967–72

53a. Han S, Tang R, Anderson LK, Wo-erner TE, Pei Z-M. 2003. A cell sur-face receptor mediates extracellular Ca2+

sensing in guard cells.Nature425:196–200

54. Harmon AC, Gribskov M, Gubrium E,Harper JF. 2001. The CDPK superfamilyof protein kinases.New Phytol.151:175–83

55. Harper JF. 2001. Dissecting calcium oscil-lators in plants.Trends Plant Sci.6:395–97

56. Harper JF, Hong B, Hwang I, Guo HQ,Stoddard R, et al. 1998. A novel calmod-ulin-regulated Ca2+-ATPase (ACA2)from Arabidopsiswith an N-terminal au-toinhibitory domain.J. Biol. Chem.273:1099–106

57. Hetherington AM. 2001. Guard cell sig-naling.Cell 107:711–14

58. Hetherington AM, Woodward FI. 2003.The role of stomata in sensing and drivingenvironmental change.Nature 424:901–1008

59. Holdaway-Clark TL, Hepler PK. 2003.Control of pollen tube growth: the role

Ann

u. R

ev. P

lant

Bio

l. 20

04.5

5:40

1-42

7. D

ownl

oade

d fr

om a

rjou

rnal

s.an

nual

revi

ews.

org

by C

NR

S-m

ulti-

site

on

12/2

8/07

. For

per

sona

l use

onl

y.



of ion gradients and fluxes.New Phytol.159:539–63

60. Hong B, Ichida A, Wang Y, Gens JS,Pickard BG, Harper JF. 1999. Identifi-cation of a calmodulin-regulated Ca2+-ATPase in the endoplasmic reticulum.Plant Physiol.119:1165–76

61. Hugouvieux V, Kwak JM, Schroeder JI.2001. An mRNA cap binding protein,ABH1, modulates early abscisic acidsignal transduction in Arabidopsis.Cell106:477–87

62. Hunt L, Mills JN, Pical C, Leckie CP,Aitken FL, et al. 2003. Phospholipase C isrequired for the control of stomatal aper-ture by ABA.Plant J.34:47–55

63. Hwang I, Sze H, Harper JF. 2000. Acalcium-dependent protein kinase can in-hibit a calmodulin-stimulated Ca2+ pump(ACA2) located in the endoplasmic retic-ulum of Arabidopsis. Proc. Natl. Acad.Sci. USA97:6224–29

64. Jaffe LF. 2002. On the conservation offast calcium wavespeeds.Cell Calcium32:217–29

65. Johnson CH, Knight MR, Kondo T, Mas-son P, Sedbrook J, et al. 1995. Circadianoscillations of cytoplasmic and chloro-plastic free calcium in plants.Science269:1863–65

66. Jung J-J, Kim Y-W, Kwak JM, Hwang J-U, Young J, et al. 2002. Phosphatidyl 3-and 4-phosphate are required for normalstomatal movements.Plant Cell14:2399–412

67. Kasai M, Muto S. 1990. Ca2+ pump andCa2+/H+ antiporter in plasma membranevesicles isolated by aqueous two-phasepartitioning from corn leaves.J. Membr.Biol. 114:133–42

68. Kiegle E, Gilliham M, Haseloff J, TesterM. 2000. Hyperpolarisation-activated cal-cium currents found only in cellsfrom the elongation zone ofArabidop-sis thaliana roots. Plant J. 21:225–29

69. Kikuyama M, Tazawa M. 2001. Mecha-nosensitive Ca2+ release from intracellu-

lar stores inNitella flexilis. Plant CellPhysiol.42:358–65

70. Klusener B, Boheim G, Liss H,Engelberth J, Weiler EW. 1995.Gadolinium-sensitive, voltage-dependentcalcium-release channels in the endo-plasmic-reticulum of a higher-plantmechanoreceptor organ.EMBO J. 14:2708–14

71. Klusener B, Weiler EW. 1999. A calcium-selective channel from root-tip endomem-branes of garden cress.Plant Physiol.119:1399–405

72. Klusener B, Young JJ, Murata Y, Allen GJ,Mori IC, et al. 2002. Convergence of cal-cium signalling pathways of pathogenicelicitors and abscisic acid in Arabidopsisguard cells.Plant Physiol.130:2152–63

73. Kohler B, Blatt MR. 2002. Protein phos-phorylation activates the guard cell Ca2+

channel and is a prerequisite for gating byabscisic acid.Plant J.32:185–94

74. Kohler B, Hills A, Blatt MR. 2003. Con-trol of guard cell ion channels by hydrogenperoxide and abscisic acid indicates theiraction through alternate signalling path-ways.Plant Physiol.131:358–88

75. Kwak JM, Moon J-H, Murata Y, KuchitsuK, Leonhardt N, et al. 2002. Disruptionof a guard cell-expressed protein phos-phatase 2A regulatory subunit, RCN1,confers abscisic acid insensitivity in Ara-bidopsis.Plant Cell14:2849–61

76. Kwak JM, Mori IC, Pei ZM, LeonhardtN, Torres MA, et al. 2003. NADPH ox-idase AtrbohD and AtrbohF genes func-tion in ROS-dependent ABA signaling inArabidopsis. EMBO J.22:2623–33

77. Lechleiter JD, John LM, Camacho P.1998. Ca2+ wave dispersion and spi-ral wave entrainment inXenopus lae-visoocytes overexpressing Ca2+ATPases.Biophys. Chem.72:123–29

78. Leckie CP, McAinsh MR, Allen GJ,Sanders D, Hetherington AM. 1998. Ab-scisic acid-induced stomatal closure me-diated by cyclic ADP-ribose.Proc. Natl.Acad. Sci. USA95:15837–42

Ann

u. R

ev. P

lant

Bio

l. 20

04.5

5:40

1-42

7. D

ownl

oade

d fr

om a

rjou

rnal

s.an

nual

revi

ews.

org

by C

NR

S-m

ulti-

site

on

12/2

8/07

. For

per

sona

l use

onl

y.



79. Lemtiri-Chlieh F, MacRobbie EAC, WebbAAR, Manison NF, Brownlee C, et al.2003. Inositol hexakisphosphate mobi-lizes an endomembrane store of calciumin guard cells.Proc. Natl. Acad. Sci. USA100:10091–95

80. Leng Q, Mercier RW, Hua BG, Fromm H,Berkowitz GA. 2002. Electrophysiologi-cal analysis of cloned cyclic nucleotide-gated ion channels.Plant Physiol.128:400–10

81. Lhuissier FGP, de Ruijter NCA, SiebererBJ, Esseling JJ, Emons AMC. 2001. Timecourse of cell biological events evoked inlegume root hairs by rhizobium Nod fac-tors: state of the art.Ann. Bot.87:289–302

82. Liang F, Cunningham KW, Harper JF,Sze H. 1997. ECA1 complements yeastmutants defective in Ca2+ pumps andencodes an endoplasmic reticulum-typeCa2+-ATPase in Arabidopsis thaliana.Proc. Natl. Acad. Sci. USA94:8579–84

83. Liang F, Sze H. 1998. A high-affinity Ca2+

pump, ECA1, from the endoplasmic retic-ulum is inhibited by cyclopiazonic acidbut not by thapsigargin.Plant Physiol.118:817–25

84. Logan DC, Knight MR. 2003. Mitochon-drial cytosolic calcium dynamics are dif-ferentially regulated in plants.Plant Phys-iol. 133:21–24

85. Luan S, Kudla J, Rodriguez-ConcepcionM, Yalovsky S, Gruissem W. 2002.Calmodulins and calcineurin B-like pro-teins: calcium sensors for specific signalresponse coupling in plants.Plant Cell14:S389–400

86. Maathuis FJM, Sanders D. 1993. Ener-gization of potassium uptake inArabidop-sis thaliana. Planta191:302–7

87. Maathuis FJM, Sanders D. 2001. Sodiumuptake in Arabidopsis roots is regu-lated by cyclic nucleotides.Plant Physiol.127:1617–25

88. MacRobbie EAC. 2000. ABA activatesmultiple Ca2+ fluxes in stomatal guardcells, triggering vacuolar K+ (Rb+) re-

lease.Proc. Natl. Acad. Sci. USA97:12361–68

89. Malli R, Frieden M, Osibow K, GraierWF. 2003. Mitochondria efficiently buffersubplasmalemmal Ca2+ elevation dur-ing agonist stimulation.J. Biol. Chem.278:10807–15

90. McAinsh MR, Brownlee C, Hethering-ton AM. 1990. ABA induced elevationof guard cell cytosolic calcium precedesstomatal closure inCommelina commu-nis. Nature343:186–88

91. McAinsh MR, Brownlee C, HetheringtonAM. 1992. Visualising changes in cytoso-lic free calcium during the response ofstomatal guard cells to abscisic acid.PlantCell 4:1113–22

92. McAinsh MR, Clayton H, MansfieldTA, Hetherington AM. 1996. Changes instomatal behaviour and guard cell cy-tosolic free calcium in response to ox-idative stress.Plant Physiol.111:1031–42

93. McAinsh MR, Hetherington AM. 1998.Encoding specificity in calcium signallingsystems.Trends Plant Sci.3:32–36

94. McAinsh MR, Webb AAR, Taylor JE,Hetherington AM. 1995. Stimulus-indu-ced oscillations in guard cell cytoplas-mic free calcium.Plant Cell7:1207–19

95. Messerli MA, Danuser G, Robinson KR.2000. Periodic increases in elongation rateprecede increases in cytosolic Ca2+ dur-ing pollen tube growth.Dev. Biol.222:84–98

96. Messerli MA, Robinson KR. 2003. Ionicand osmotic disruptions of the lily pollentube oscillator: testing proposed models.Planta217:147–57

97. Moutinho A, Hussey PJ, Trewavas AJ,Malho R. 2001. cAMP acts as a secondmessenger in pollen tube growth and re-orientation.Proc. Natl. Acad. Sci. USA98:10481–86

98. Muir SR, Sanders D. 1997. Inositol1,4,5 trisphosphate-sensitive Ca2+ releaseacross nonvascular membranes in cauli-flower.Plant Physiol.114:1511–21

Ann

u. R

ev. P

lant

Bio

l. 20

04.5

5:40

1-42

7. D

ownl

oade

d fr

om a

rjou

rnal

s.an

nual

revi

ews.

org

by C

NR

S-m

ulti-

site

on

12/2

8/07

. For

per

sona

l use

onl

y.



99. Murata Y, Pei Z-M, Mori IC, SchroederJI. 2001. Abscisic acid activation ofplasma membrane Ca2+ channels inguard cells requires cytosolic NAD(P)Hand is differentially disrupted upstreamand downstream of reactive oxygenspecies production in abi1-1 andabi2-1 protein phosphatase 2C mutants.Plant Cell13:2513–23

100. Nagai T, Sawano A, Park ES, MiyawakiA. 2001. Circularly permuted green flu-orescent proteins engineered to senseCa2+. Proc. Natl. Acad. Sci. USA98:3197–202

101. Navazio L, Bewell MA, Siddiqua A, Dick-inson GD, Galione A, Sanders D. 2000.Calcium release from the endoplasmicreticulum of higher plants elicited by theNADP metabolite nicotinic acid adeninedinucleotide phosphate.Proc. Natl. Acad.Sci. USA97:8693–98

102. Navazio L, Mariani P, Sanders D. 2001.Mobilization of Ca2+ by cyclic ADP-ribose from the endoplasmic reticulumof cauliflower florets. Plant Physiol.125:2129–38

103. Ng CK-Y, Carr K, McAinsh MR, Pow-ell B, Hetherington AM. 2001. Drought-induced guard cell signal transduction in-volves sphingosine-1-phosphate.Nature410:596–99

104. Okasaki Y, Ishigami M, Iwasaki N. 2002.Temporal relationship between cytosolicfree Ca2+ and membrane potential duringhypotonic turgor regulation in a brack-ish water charophyteLamprothamniumsuccinctum. Plant Cell Physiol.43:1027–35

105. Ordenes VR, Reyes F, Wolff D, OrellanaA. 2002. A thapsigarin-sensitive Ca2+

pump is present in the pea Golgi apparatusmembrane.Plant Physiol.19:1820–28

106. Pauly N, Knight MR, Thuleau P, GrazianaA, Muto S, et al. 2001. The nucleustogether with the cytosol generates pat-terns of specific cellular calcium signa-tures in tobacco culture cells.Cell Cal-cium30:413–21

107. Pauly N, Knight MR, Thuleau P, van derLuit AH, Moreau M, et al. 2000. Cellsignalling—Control of free calcium inplant nuclei.Nature405:754–55

108. Pei Z-M, Murata Y, Benning G, ThomineS, Klusener B, et al. 2000. Calcium chan-nels activated by hydrogen peroxide medi-ate abscisic acid signalling in guard cells.Nature406:731–34

109. Pei Z-M, Ward JM, Schroeder JI. 1999.Magnesium sensitizes slow vacuolarchannels to physiological cytosolic cal-cium and inhibits fast vacuolar channelsin fava bean guard cell vacuoles.PlantPhysiol.121:977–86

110. Persson S, Wyatt SE, Love J, ThompsonWF, Robertson D, Boss WF. 2001. TheCa2+ status of the endoplasmic reticulumis altered by induction of calreticulin ex-pression in transgenic plants.Plant Phys-iol. 126:1092–104

111. Pittman JK, Hirschi KD. 2001. Regula-tion of CAX1, an ArabidopsisCa2+/H+

antiporter. Identification of an N-terminalautoinhibitory domain.Plant Physiol.127:1020–29

112. Pittman JK, Hirschi KD. 2003. Don’tshoot the (second) messenger: endomem-brane transporters and binding proteinsmodulate cytosolic Ca2+ levels. Curr.Opin. Plant Biol.6:257–62

113. Reddy VS, Ali GS, Reddy ASN. 2002.Genes encoding calmodulin-binding pro-teins in the Arabidopsis genome.J. Biol.Chem.277:9840–52

114. Rudd JJ, Franklin-Tong VE. 2001. Un-ravelling response-specificity in Ca2+ sig-nalling pathways in plant cells.New Phy-tol. 151:7–33

115. Sai JQ, Johnson CH. 2002. Dark-stimulated calcium ion fluxes in the chlo-roplast stroma and cytosol.Plant Cell14:1279–91

116. Sanders D, Pelloux J, Brownlee C, HarperJF. 2002. Calcium at the crossroads of sig-nalling.Plant Cell14:S401–17

117. Schroeder JI, Allen GJ, Hugouvieux V,Kwak JM, Waner D. 2001. Guard cell

Ann

u. R

ev. P

lant

Bio

l. 20

04.5

5:40

1-42

7. D

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12/2

8/07

. For

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signal transduction.Annu. Rev. PlantPhysiol. Plant Mol. Biol.52:627–58

118. Scrase-Field SAMG, Knight MR. 2003.Calcium: just a chemical switch?Curr.Opin. Plant Biol.6:500–6

119. Shaw SL, Long SR. 2003. Nod factorelicits two separable calcium responsesin Medicago truncatularoot hairs.PlantPhysiol.131:976–84

120. Snedden WA, Fromm H. 2001. Calmod-ulin as a versatile calcium signal trans-ducer in plants.New Phytol.151:35–66

121. Staxen I, Pical C, Montgomery LT, GrayJE, Hetherington AM, McAinsh MR.1999. Abscisic acid induces oscillationsin guard-cell cytosolic free calcium thatinvolve phosphoinositide-specific phos-pholipase C.Proc. Natl. Acad. Sci. USA96:1779–84

122. Subbaiah CC, Bush DS, Sachs MM. 1994.Elevation of cytosolic calcium precedesanoxic gene expression in maize suspen-sion cultured cells.Plant Cell6:1747–62

123. Szabadki G, Simoni AM, Rizzuto R.2003. Mitochondrial Ca2+uptake requiressustained Ca2+ release from the endoplas-mic reticulum.J. Biol. Chem.278:15153–61

124. Sze H, Liang F, Hwang I, Curran AC,Harper JF. 2000. Diversity and regulationof Ca2+ pumps: insights from expressionin yeast.Annu. Rev. Plant Physiol. PlantMol. Biol. 51:433–62

125. Takahashi K, Isobe M, Knight MR, Tre-wavas AJ, Muto S. 1997. Hypo-osmoticshock induces increases in cytosolic Ca2+

in tobacco suspension culture cells.PlantPhysiol.113:587–94

126. Talke IN, Blaudez D, Maathuis FJM,Sanders D. 2003. CNGCs: prime tar-gets of plant cyclic nucleotide signalling?Trends Plant Sci.8:286–92

127. Taylor AR, Manison NFH, Fernandez C,Wood J, Brownlee C. 1996. Spatial orga-nization of calcium signalling involved incell volume control in theFucusrhizoid.Plant Cell8:2015–31

128. Thion L, Mazars C, Nacry P, Bouchez

D, Moreau M, et al. 1998. Plasma mem-brane depolarisation-activated calciumchannels, stimulated by microtubule-depolymerizing drugs in wild typeAra-bidopsis thaliana protoplasts, displayconstitutively large activities and a longerhalf life in ton2 mutant cells affected inthe organization of cortical microtubules.Plant J.13:603–10

129. Very AA, Davies JM. 2000. Hyper-polarization-activated calcium channelsat the tip of Arabidopsis root hairs.Proc.Natl. Acad. Sci. USA97:9801–906

130. Very AA, Sentenac H. 2002. Cation chan-nels in theArabidopsisplasma membrane.Trends Plant Sci.7:168–75

131. Wais RJ, Galera C, Oldroyd G, Catoira R,Penmetsa RV, et al. 2000. Genetic analy-sis of calcium spiking response in nodu-lation mutants ofMedicago truncatula.Proc. Natl. Acad. Sci. USA97:13407–12

132. Walker SA, Viprey V, Downie JA. 2000.Dissection of nodulation signaling usingpea mutants defective for calcium spik-ing induced by Nod factors and chitinoligomers.Proc. Natl. Acad. Sci. USA97:13413–18

133. Webb AAR, McAinsh MR, Mansfield TA,Hetherington AM. 1996. Carbon dioxideinduces increases in guard cell cytosolicfree calcium.Plant J.9:297–304

134. Webb SE, Miller AL. 2003. Calcium sig-naling during embryonic development.Nat. Rev. Cell Mol. Biol.4:539–51

135. White PJ. 1998. Calcium channels in theplasma membrane of root cells.Ann. Bot.81:173–83

136. White PJ. 2000. Calcium channels inhigher plants.Biochim. Biophys. Acta1465:171–89

137. White PJ, Bowen HC, Demidchik V,Nichols C, Davies JM. 2002. Genesfor calcium-permeable channels in theplasma membrane of plant root cells.Biochim. Biophys. Acta1564:299–309

138. Worrall D, Ng CK, Hetherington AM.2003. Sphingolipids, new players in plantsignalling.Trend. Plant Sci.8:317–20

Ann

u. R

ev. P

lant

Bio

l. 20

04.5

5:40

1-42

7. D

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org

by C

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12/2

8/07

. For

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139. Wu Z, Liang F, Hong B, Young JC, Suss-man MR. 2002. An ER-bound Ca2+/Mn2+

pump ECA1 supports plant growth andconfers tolerance to Mn2+ stress.PlantPhysiol.130:128–37

140. Wymer CL, Bibikova TN, Gilroy S. 1997.Cytoplasmic free calcium distributionsduring the development of root hairs ofArabidopsis thaliana. Plant J.12:427–39

141. Yuasa K, Maeshima M. 2000. Purifica-tion, properties and molecular cloning of anovel Ca2+ binding protein in radish vac-uoles.Plant Physiol.124:1069–78

142. Zhang J, Campbell RE, Ting AY, TsienRY. 2002. Creating new fluorescent

probes for cell biology.Nat. Rev. Cell Mol.Biol. 3:906–18

143. Zhang L, Lu YT. 2003. Calmodulin-binding protein kinases in plants.Trends.Plant Sci.8:123–27

144. Zhang X, Zhang L, Dong FC, Gao JF,Galbraith DW, Song C-P. 2001. Hydro-gen peroxide is involved in abscisic acid-induced stomatal closure inVicia faba.Plant Physiol.126:1438–48

145. Zou H, Lifshitz LM, Tuft RA, Fogarty KE,Singer JJ. 2002. Visualization of Ca2+ en-try through single stretch-activated cationchannels.Proc. Natl. Acad. Sci. USA99:6404–9

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CALCIUM SIGNALING C-1

Figure 1 Schematic representation of complex interactions between channels, pumps,transporters, and Ca2+-binding proteins in bringing about [Ca2+]cyt homeostasis andstimulus-specific elevations (as represented by the solid black trace) or oscillations of[Ca2+]cyt. Light arrows indicate hypothetical actions in raising or lowering [Ca2+]cyt atparticular points in a Ca2+ transient. The assignment of Ca2+ pumps and transporters todifferent membranes is indicative rather than exhaustive. The combined activity ofchannels and pumps contributes to resting [Ca2+]cyt. NSCCs and HACCs may be par-ticularly involved in maintaining steady Ca2+ influx. The chloroplast and nucleus arenot shown. ACA,ECA, Ca2+-ATPAses; CAX, H+/Ca2+ exchangers; NSCC, nonselec-tive cation channels; DACC, depolarization-activated Ca2+ channels; HACC, hyperpo-larization-activated Ca2+ channels; GLR, glutamate receptor channels; CNGC, cyclicnucleotide gated channels; PM, plasma membrane; mit, mitochondria; ER, endoplas-mic reticulum; CRT, calreticulin; VCaB, vacuolar Ca2+-binding protein.

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C-2 HETHERINGTON ■ BROWNLEE

Figure 2 Hypothetical scheme for Ca2+ cycling between the cytosol, the ER, and mito-chondria. [Ca2+]cyt elevations arising via influx through plasma membrane (PM) orendoplasmic reticulum (ER) channels (e.g., in response to touch, osmotic, or cold) areoffset by mitochondrial and ER uptake or reuptake. Ca2+ efflux from mitochondria isremoved from the cytosol by adjacent ER. [Ca2+]mit elevations stimulate ATP produc-tion, which may be used to fuel Ca2+ efflux via Ca2+- ATPase activity. Green arrowsrepresent the proposed Ca2+ cycling pathway.

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P1: FRK

March 24, 2004 1:1 Annual Reviews AR213-FM

Annual Review of Plant BiologyVolume 55, 2004

CONTENTS

AN UNFORESEEN VOYAGE TO THE WORLD OF PHYTOCHROMES,Masaki Furuya 1

ALTERNATIVE NAD(P)H DEHYDROGENASES OF PLANTMITOCHONDRIA, Allan G. Rasmusson, Kathleen L. Soole,and Thomas E. Elthon 23

DNA METHYLATION AND EPIGENETICS, Judith Bender 41

PHOSPHOENOLPYRUVATE CARBOXYLASE: A NEW ERA OFSTRUCTURAL BIOLOGY, Katsura Izui, Hiroyoshi Matsumura,Tsuyoshi Furumoto, and Yasushi Kai 69

METABOLIC CHANNELING IN PLANTS, Brenda S.J. Winkel 85

RHAMNOGALACTURONAN II: STRUCTURE AND FUNCTION OF ABORATE CROSS-LINKED CELL WALL PECTIC POLYSACCHARIDE,Malcolm A. O’Neill, Tadashi Ishii, Peter Albersheim, and Alan G. Darvill 109

NATURALLY OCCURRING GENETIC VARIATION IN ARABIDOPSISTHALIANA, Maarten Koornneef, Carlos Alonso-Blanco, andDick Vreugdenhil 141

SINGLE-CELL C4 PHOTOSYNTHESIS VERSUS THE DUAL-CELL (KRANZ)PARADIGM, Gerald E. Edwards, Vincent R. Franceschi,and Elena V. Voznesenskaya 173

MOLECULAR MECHANISM OF GIBBERELLIN SIGNALING IN PLANTS,Tai-ping Sun and Frank Gubler 197

PHYTOESTROGENS, Richard A. Dixon 225

DECODING Ca2+ SIGNALS THROUGH PLANT PROTEIN KINASES,Jeffrey F. Harper, Ghislain Breton, and Alice Harmon 263

PLASTID TRANSFORMATION IN HIGHER PLANTS, Pal Maliga 289

SYMBIOSES OF GRASSES WITH SEEDBORNE FUNGAL ENDOPHYTES,Christopher L. Schardl, Adrian Leuchtmann, Martin J. Spiering 315

TRANSPORT MECHANISMS FOR ORGANIC FORMS OF CARBON ANDNITROGEN BETWEEN SOURCE AND SINK, Sylvie Lalonde,Daniel Wipf, and Wolf B. Frommer 341

vii

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March 24, 2004 1:1 Annual Reviews AR213-FM

viii CONTENTS

REACTIVE OXYGEN SPECIES: METABOLISM, OXIDATIVE STRESS,AND SIGNAL TRANSDUCTION, Klaus Apel and Heribert Hirt 373

THE GENERATION OF Ca2+ SIGNALS IN PLANTS,Alistair M. Hetherington and Colin Brownlee 401

BIOSYNTHESIS AND ACCUMULATION OF STEROLS, Pierre Benveniste 429

HOW DO CROP PLANTS TOLERATE ACID SOILS? MECHANISMS OFALUMINUM TOLERANCE AND PHOSPHOROUS EFFICIENCY,Leon V. Kochian, Owen A. Hoekenga, and Miguel A. Pineros 459

VIGS VECTORS FOR GENE SLIENCING: MANY TARGETS,MANY TOOLS, Dominique Robertson 495

GENETIC REGULATION OF TIME TO FLOWER IN ARABIDOPSIS THALIANA,Yoshibumi Komeda 521

VISUALIZING CHROMOSOME STRUCTURE/ORGANIZATION,Eric Lam, Naohiro Kato, and Koichi Watanabe 537

THE UBIQUITIN 26S PROTEASOME PROTEOLYTIC PATHWAY,Jan Smalle and Richard D. Vierstra 555

RISING ATMOSPHERIC CARBON DIOXIDE: PLANTS FACE THE FUTURE,Stephen P. Long, Elizabeth A. Ainsworth, Alistair Rogers,and Donald R. Ort 591

INDEXESSubject Index 629Cumulative Index of Contributing Authors, Volumes 45–55 661Cumulative Index of Chapter Titles, Volumes 45–55 666

ERRATAAn online log of corrections to Annual Review of Plant Biologychapters may be found at http://plant.annualreviews.org/

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