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Abiotic stress signalling pathways: specificity and cross-talk
Heather Knight and Marc R. Knight
Plants exhibit a variety of responses to abiotic stresses that enable them to tolerate and survive adverse conditions. As we learn more about the signalling pathways leading to these responses, it is becoming clear that they constitute a network that is interconnected at many levels. In this article, we discuss the ‘cross-talk’ between different signalling pathways and question whether there are any truly specific abiotic stress signalling responses.

Heather Knight* Marc R. Knight Dept Plant Sciences, University of Oxford, Oxford, UK OX1 3RB. *e-mail: heather.knight@plants. ox.ac.uk

Plants encounter a wide range of environmental insults during a typical life cycle and have evolved mechanisms by which to increase their tolerance of these through both physical adaptations and interactive molecular and cellular changes that begin after the onset of stress. The first step in switching on such molecular responses is to perceive the stress as it occurs and to relay information about it through a signal transduction pathway. These pathways eventually lead to physiological changes, such as guard cell closure, or to the expression of genes and resultant modification of molecular and cellular processes. Our knowledge about the signalling pathways leading from stimulus to end response in plants has increased over recent years. It is increasingly apparent that the linearpathways that we have been studying are actually only part of a more complex signalling network and that there is much overlap between its branches, with, for instance, many genes inducible by more than one particular stimulus. In this article, we discuss two aspects of these abiotic stress signalling networks, namely cross-talk and specificity. We define ‘cross-talk’ as any instance of two signalling pathways from different stressors that converge. This might take the form of different pathways achieving the same end or of pathways interacting and affecting each other’s outcome, including the flux through one pathway affecting another. These might act in an additive or negatively regulatory way, or might compete for a target (Fig. 1). We define specificity as any part of a signalling pathway that enables distinction between two or more possible outcomes and that thus might link a particular stimulus to a particular end response and not to any other end responses. Opportunities for both cross-talk and specificity can occur within a particular pathway.
Cross-talk

nature, however, the plant encounters stress combinations concurrently or separated temporally and must present an integrated response to them. In the case of phytochrome signalling, the two pathways leading to red-light-induced CHS and CAB gene expression negatively regulate flux through one another1,2. Seemingly separate abiotic stress signalling pathways are also likely to interact in a similar manner. In addition, several abiotic stress pathways share common elementsthat are potential ‘nodes’ for cross-talk. Cross-talk can also occur between pathways in different organs of the plant when a systemic signal such as hydrogen peroxide moves from a stimulated cell into another tissue to elicit a response3.
Specificity

When stress signalling pathways are examined in the laboratory, they are usually considered in isolation from other stresses to simplify interpretation. In

In spite of considerable overlap between many abiotic stress signalling pathways, there might, in some instances, be a benefit to producing specific, inducible and appropriate responses that result in a specific change suited to the particular stress conditions encountered. One advantage would be to avoid the high energy cost of producing stress-tolerance proteins, exemplified by the dwarf phenotype of plants constitutively overexpressing the frost tolerance protein DREB1A (Ref. 4). In some cases, the signal transduction pathways triggered by different stresses are common to more than one stress type. One possible reason for this is that, under certain conditions, the two stresses cannot be distinguished from one another. Alternatively, each stress might require the same protective action (or at least some common elements). The discovery of separate sensing mechanisms for each stress would invalidate the first suggestion but the second is true in several cases. For example, dehydration protection is required in plants undergoing either freezing or drought and the production of antioxidants and scavenging enzymes (e.g.catalase and peroxidases) that protect against oxidative damage affords protection against a variety of different abiotic (and biological) stresses5. Most abiotic stresses tested have been shown to elicit rises in cytosolic free calcium levels ([Ca2+]cyt) and to involve protein phosphatases and kinases [including mitogen-activated protein kinase (MAPK) cascades]. However, are any of these components truly specific to one stress and which of them are ‘nodes’ at which cross-talk occurs? In the following sections, we consider different classes of signalling component in turn, and examine their potential

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Stimulus 1 A B C D

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these molecules themselves have the potential to encode specificity of response. An early event in the response to many different environmental stresses is an elevation in [Ca2+]cyt (Refs 7,8), which is thought to be the primary stimulus-sensing event for several stresses (e.g. cold)9–11. If this is the case then mechanisms could exist for encoding the information that relates to the particular stress through the calcium signature (see below). Alternatively, the stress might be sensed through other components either in parallel to or upstream of Ca2+ in the pathway. It has been postulated that cold is sensed via changes in membrane fluidity12 and cytoskeletal reorganization13affecting calcium channels.
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Fig. 1. Cross-talk in signalling pathways. (a) Two different stimuli (1 and 2) evoke the same end response via different signalling pathways, using different signalling intermediates (A–D and W–Z, respectively). (b) Positive and negative reciprocal control. Two different stimuli (A and B) activate two signalling pathways (broken arrows), leading to different end responses. (i) Pathways operating totally independently of each other. (ii) Flux through the stimulus-A-mediated pathway negatively regulates the stimulus-B-mediated pathway and inhibits its flux. An example of this is in phytochrome-mediated expression of a chlorophyll a/b binding protein gene (CAB) and a chalcone synthase gene (CHS) by independent pathways, each negatively regulating the other. (iii) Flux through the stimulus-A-mediated pathway positively regulates the stimulus-B-mediated pathway and promotes its flux. (c) (i) Signalling pathway leading from a stimulus (stimulus 1) using a specific signalling component (green) to effect response X. (ii) Stimulus 2 uses this same signalling component to mediate its end response (response Y) and, by out-competing the stimulus-1 pathway, inhibits it. Calcium is an example of this: one stimulus might exhaust a specific pool of calcium andmake it unavailable for use by another stimulus.

contribution to specificity and cross-talk between abiotic stress signalling pathways.
Sensing systems

Specificity might occur at the point of initial stress perception itself. In the case of osmotic stress, the putative osmosensor AtHK1 (Ref. 6), a transmembrane histidine kinase, is thought to be the first component to relay changes in osmotic potential outside the cell to the transduction pathway(s) inside the cell that regulates drought-inducible gene expression. If specific stresses are actually sensed by dedicated receptor molecules,
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The precise kinetics, magnitude and cellular source of stimulus-induced [Ca2+]cyt elevations (the ‘calcium signature’) have been proposed to encode information about the particular stimulus, and to determine the specific end response elicited14. Biphasic elevations, responses lasting from two seconds to tens of minutes and repeated oscillations are among the responses observed after abiotic stress. Studies using animal cells showed that the Ca2+-induced activation of particular transcription factors could be specified by the magnitude and kinetics of an artificially induced [Ca2+]cyt elevation15. This suggests that cells can decode specific information in the [Ca2+]cyt elevation that refers to particular stimuli, and relate this to an appropriate change in response. However, such data do not prove that this system of encoding specificity is actually used by cells. Also, these measurements were all made onpopulations of cells, obscuring the complexity of individual cell responses. In plants, the principle of specificity through calcium signatures has been difficult to show because of problems in generating artificial [Ca2+]cyt elevations to meet specific designs. In some cases, [Ca2+]cyt has been successfully elevated in the absence of an abiotic stress by using a Ca2+ ionophore or Ca2+ channel agonists16,17. The correlation of such [Ca2+]cyt elevations with changes in the end responses usually associated with exposure to the stress shows a requirement for Ca2+ in the transduction of these stimuli. However, it also supports the argument that specificity is not encoded through the calcium signature because it is most unlikely that the artificial [Ca2+]cyt elevations could have the same subcellular source and calcium signature as those normally provoked by that particular stress. It is more likely that the [Ca2+]cyt elevation must achieve a minimum (and perhaps maximum) threshold peak value or total elevation (magnitude × time). Ozone exposure elicits a brief [Ca2+]cyt peak followed by a more prolonged elevation18. Only the second of these is necessary for the induction of GST gene expression and so, even though these data might imply the significance of the calcium signature, they might also imply that the ‘magnitude × time’hypothesis is true.


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In stomatal guard cells, variation in the timing of stimulus-induced Ca2+ oscillations has been correlated with the intensity of boththe stimulus and the resultant end response, with alterations in the signature associated with loss of aperture closure14,19. External Ca2+ or oxidative stress elicited Ca2+ oscillations followed by stomatal closure in the wild type, but cells of the det3 Arabidopsis mutant (which has impaired endomembrane energization) failed to show normal wild-type oscillations and did not close. However, det3 cells responded normally to cold and abscisic acid (ABA) stimulation, indicating that there are specific Ca2+-dependent pathways for different stresses. Concurrent addition of external Ca2+ and repeated depolarization of the plasma membrane induced artificial Ca2+ oscillations in det3 cells and initiated stomatal closure. These data suggest that Ca2+ oscillations are required for stomatal closure to occur in response to certain signals. Interestingly, the non-oscillatory stimulus-induced [Ca2+]cyt elevations seen in det3 mutants constituted a larger overall total increase in [Ca2+]cyt than did the wild-type oscillations. This might support the idea that the response is not achieved if the total [Ca2+]cyt elevation exceeds a certain level15. It should also be borne in mind that the det3 [Ca2+]cyt elevation might occur in the wrong cellular location. Various plant abiotic stress [Ca2+]cyt responses use Ca2+ from different subcellular sources, including the extracellular compartment, vacuole20 and mitochondria21. The Ca2+ signature reflects the source used22 and might encode information of specific relevance to the cellular machinery basedin those organelles. It is possible that ‘effective’ Ca2+ signatures only occur in those cell types that are required to respond. [Ca2+]cyt elevations occur globally in plants responding to cold but only in the root after drought22 (S. Scrase-Field and M.R. Knight, unpublished). Within the Arabidopsis root, the [Ca2+]cyt responses of epidermal, endodermal, pericycle and cortex cells differ from each other when challenged with cold, drought and salt23. These tissue-specific differences might correlate with different stress protein production or other responses. Although plants exhibit recognizable [Ca2+]cyt elevations in response to particular stresses, these are altered markedly after previous stress experiences8,24, indicating cross-talk between abiotic stress signal transduction pathways occurring at the level of Ca2+. Whereas an oxidative stress encounter abolished future responses to drought stimulus in Arabidopsis24, it increased the sensitivity of response to low temperature levels8. Drought pretreatment increased the magnitude of subsequent droughtinduced [Ca2+]cyt transients and increased the level of drought-inducible Ca2+-regulated gene expression and stress tolerance24. In summary, the Ca2+ signal is ubiquitous in abiotic stress signalling and it is therefore an important node at which cross-talk can occur.
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Calcium-regulated proteins

[Ca2+]cyt elevations achieve control of various processes via Ca2+-regulated effector proteins. Sometimes referred to as ‘calcium sensors’, these includecalmodulin, calcium-dependent protein kinases (CDPKs) and calcium-regulated phosphatases. Calmodulin has been implicated in plant responses to cold25, mechanical stimulation25,26 and oxidative stress27. The use of different isoforms could be involved in control of specificity between these pathways, as has been observed with calmodulin isoforms in specificity to biotic stresses such as salicylic-acid-mediated defence responses28. In animal cells, anchoring proteins located in different parts of the cell compartmentalize kinases to their site of action29. It is possible that specificity of response in plant systems, too, is introduced by localizing calcium sensors in this way. Only two out of eight Arabidopsis CDPK isoforms introduced into maize protoplasts induced expression of a specific stress-inducible gene, suggesting that there are specific CDPK isoforms for different stress signalling pathways17. However, these experiments involved the overexpression of constitutive versions of CDPK isoforms, which, lacking all but the kinase domains of the proteins, might not target appropriately. Also, because they are ectopically expressed, they might phosphorylate illegitimate substrates. Therefore, these results might not reflect the in vivo situation in Arabidopsis. There are many different CDPKs in Arabidopsis [~40 (Ref. 30)], therefore there is ample scope for partitioning of specific CDPK function between isoforms. The genes for some isoforms are induced in response to specific stresses, implicating these CDPKs in particular signallingpathways; thus, for example, AtCDPK1 and AtCDPK2 are implicated in salt and drought stress signalling17,31. In rice, expression of OsCDPK7 at the mRNA level is inducible by cold or salt stress32. However, overexpression of OsCDPK7 enhanced saltand drought-induced, but not cold-induced, expression of target genes, suggesting that the effect of this CDPK is specific to salt and drought signalling. Another group of proteins identified as interacting with Ca2+ to effect an end response include serine/threonine phosphatases33 (PPases). The type-2C PPases include a group of proteins with similarities to the calcium sensor protein calcineurin B (Refs 34,35) that are referred to in Arabidopsis as calcineurin-Blike (AtCBL) proteins. One of these, SOS3, is involved specifically in salt stress tolerance34 and the gene encoding another, AtCBL1 (but not those of other family members), is highly upregulated by cold and drought. Specificity through AtCBL isoforms might result from variation in the N-terminal sequence of AtCBL proteins, meaning that only some members have the potential to be targeted to membrane sites36. As well as achieving specificity through the use of different isoforms, calcium sensors might also serve as nodes at which cross-talk can occur. [Ca2+]cyt elevations


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Cold Signal transduction

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Fig. 2.The DREB1 and DREB2 transcription factors, key components in cross-talk between cold and drought signalling in Arabidopsis. Cold and drought activate the expression of the DREB1 and DREB2 families of drought-responsive-element-binding (DRE-binding) transcription factors, respectively. Both sets of transcription factors alight on the same cis-acting element in the promoters of genes such as RD29A (also known as LTI78) called the DRE element. Therefore, the DRE element is an integration point for cross-talk between cold and drought signalling in Arabidopsis. For simplicity, the abscisicacid-responsive element is not shown on this diagram.

addition of a constitutively active ANP1 (via endogenous MAPKKs)45. Hydrogen peroxide (oxidative stress) increased the activation of MPK3 above the level achieved with the constitutively active ANP1. Further experiments showed that active ANP1 could activate hydrogen-peroxide-inducible promoters but not drought- or cold- or ABA-responsive promoters, implying primary signal-specificity encoded within ANP1. Reconstruction of the AtMEKK1 kinase cascade in yeast indicates that there is selectivity in the partners that associate with it. Using a combination of two-hybrid and yeast mutant complementation, it was found that AtMEKK1 preferentially associated with and activated the closely related MAPKKs, AtMKK2 and MEK1, which, in turn, associated with and activated the MAPK AtMPK4 (Refs 46,47). Thus, specificity could be encoded by only allowing certain MAPKKK–MAPKK–MAPK combinations in complexes, like‘chords’in a piece of music (H. Hirt, pers. commun.). There is evidence of cross-talk between these ‘chords’during abiotic stress, such as between MAPK cascades leading separately to ATMPK4 and ATMPK6 (Ref. 48).
Transcription factors

elicited by a specific stress could regulate phosphatases and kinases involved in the transduction of another stimulus. In alfalfa cells, cold-induced inactivation of protein phosphatase 2A (PP2A) is controlled by Ca2+ influx37, but it is conceivable that PP2A activity could also be modulated via other stress-induced [Ca2+]cyt elevations. The ABI1 and ABI2 proteins (identified through the abi1 and abi2 mutations) are homologous to type2C PPases38,39 and have been implicated as negative regulators in the early stages of ABA signal transduction40,41. These proteins are potential nodes for cross-talk between different signalling pathways involving ABA (e.g. cold and drought). Recently, a tobacco homologue of the Arabidopsis PP2C genes has been identified, NtPP2C1, transcript accumulation of which is upregulated by drought treatment but inhibited by oxidative stress or heat42. If NtPP2C1 functions as a negative regulator in a similar way to Arabidopsis ABI1 and ABI2, it is possible that downregulation of its expression by oxidative and heat stresses could increase cellular sensitivity to ABA during drought treatment.
MAPK cascades

MAPK cascades are activated by numerous abiotic stresses43 but they can introduce specificity into the system. A MAPK kinase kinase (MAPKKK) phosphorylates a MAPK kinase(MAPKK), which in turn phosphorylates a MAPK. Three major types of MAPKKK have been identified in Arabidopsis: CTR1, ANP1-3 and the AtMEKK class. AtMEKK1 (Ref. 44) is expressed in response to abiotic stresses including cold, drought and mechanical stimulation. Of six target MAPKs, only two (AtMPK3 and AtMPK6) showed evidence of being phosphorylated in response to the
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Low positive temperatures increase the level of freezing tolerance in many plant species through cold acclimation49, but this state can also be achieved in response to drought or by application of the phytohormone ABA (Ref. 50). Many genes that are induced by cold are also induced by drought or ABA (Ref. 51), probably because many cold-inducible genes encode proteins to protect the plant from the consequences of freezing stress, which include dehydration. The gene RD29A (also known as LTI78 or COR78) has been used in several studies examining the convergence of these pathways. RD29A is one of many cold- and drought-regulated genes that have been found to contain the so-called DRE or CRT (drought-responsive or C-repeat element) in their promoters52. In Arabidopsis, two groups of transcription factors, DREB1 (also known as CBF) and DREB2, bind to this cis-acting element4,51,53. The DREB1 and DREB2 genes encode structurally different proteins and are induced specifically by low temperature and by salt or drought, respectively (Fig. 2). DREB2A and DREB2B are produced in the root only in response to salinity, but are produced in the stem and theroot after drought treatment54, offering a further level of specificity of response. Overproduction of either DREB1 or DREB2 proteins in protoplasts increased expression of an artificial RD29A-promoter GUS fusion gene4, indicating that the DRE promoter element is a point at which drought or salt and cold signal transduction pathways converge and that it can integrate information about these two stimuli (Fig. 2). It does seem strange that two entirely separate signalling pathways lead to the action of these two sets of transcription factors (Fig. 2) simply for them to result in the activation of the same cis-acting element in the


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of Arabidopsis57, which is deficient in the expression of cold-regulated genes that contain a DRE element but expresses non-DRE-regulated cold genes normally. Interestingly, in sfr6, DRE-containing genes also failed to be expressed fully in response to stimulation by ABA, suggesting that there is cross-talk between the ABRE and DRE. The interactions between these different pathways have been investigated in Arabidopsis through the analysis of mutants defective in the induction of RD29A by salt, cold, ABA or acombination of these58. The HOS1 and HOS2 loci encode signalling components that negatively regulate gene expression in response specifically to cold and not to other stress stimuli59,60. Conversely, HOS5 reduces expression in response to ABA and osmotic stresses but not to cold61. Combined cold and ABA or salt and ABA treatments have been shown to have a synergistic effect on RD29A–LUC expression62, in contrast with the reduced levels of expression in response to combined cold and salt treatment. The results of these studies have suggested that ABA-dependent and ABA-independent osmotic and cold stress pathways might converge at several hitherto unexpected points (Fig. 3). If they do, this increases opportunities for coordination between stress signals and ABA in the regulation of gene expression.
Perspectives

Fig. 3. Cold, osmotic stress and abscisic acid (ABA) signal transduction, as determined by the use of los, cos and hos mutants of Arabidopsis. The positions of HOS, COS and LOS gene products in the signalling pathways are indicated by pale blue arrows. The abscisic acid (ABA)-dependent pathway (red) interacts and eventually converges with ABA-independent pathways (dark-blue and green) to activate the expression of RD29A (also known as LTI78) and other genes containing the drought responsive and abscisic-acid-responsive promoter elements (DRE and ABRE) (black). Broken arrows indicate pathways (or parts of pathways) that cannot directly induce the expression of these genes but that require interaction with other pathways orbranches of pathways. (Modified from Ref. 58.)

same genes. It might well be that, in other species, the DREB1 and DREB2 factors control the expression of two different sets of genes and that Arabidopsis has, during evolution, rationalized these two functions. An ABA-responsive promoter element (ABRE) has been identified and a family of basic leucine zipper (bZIP) DNA-binding protein interact with it55,56. Genes such as RD29A contain both DRE and ABRE elements in their promoters and can be activated by ABAdependent and ABA-independent pathways. A single ABRE element cannot function independently56, for instance, RD29B has two copies. RD29A has both an ABRE and a DRE, suggesting that, in promoters containing both elements, the ABRE requires the DRE for ABA-induced expression. This suggestion is supported by data from the sfr6 cold acclimation mutant
References 1 Bowler, C. and Chua, N.H. (1994) Emerging themes of plant signal transduction. Plant Cell 6, 1529–1541 2 Bowler, C. et al. (1994) Phytochrome signaltransduction pathways are regulated by reciprocal control mechanisms. Genes Dev. 8, 2188–2202
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Studying abiotic stress signalling pathways in isolation is valuable but it can be misleading because they form part of complex networks. In future, the onus will be on taking this fact into account, both intellectually and in terms of technology development. A perfect example of this is the availability of microarray technology. This enables researchers to examine the expression of not only all their particularstress-induced genes of interest but also thousands of others, without prejudice and without extra effort63. This will lead to a greater understanding of the effect that abiotic stress pathways have on each other as well as on pathways and processes that were not known to be connected. The recent completion of the Arabidopsis genome project means that identifying the genes involved in specificity and cross-talk will be more rapid. This will then lead into work on the proteins themselves, to ask how exactly these proteins operate to encode specificity or to act as communicators or nodes in cross-talk. Finally, signalling components whose genes are induced by abiotic stress, and hence implied in abiotic stress signalling, can be directly tested for specificity or cross-talk (problems of redundancy aside) by phenotypic analysis of knockout mutants.
transduction pathways in drought- and lowtemperature-responsive gene expression, respectively, in Arabidopsis. Plant Cell 10, 1391–1406 5 Bowler, C. and Fluhr, R. (2000) The role of calcium and activated oxygens as signals for controlling cross-tolerance. Trends Plant Sci. 5, 241–246

3 Foyer, C.H. et al. (1997) Hydrogen peroxide- and glutathione-associated mechanisms of acclimatory stress tolerance and signalling. Physiol. Plant. 100, 241–254 4 Liu, Q. et al. (1998) Two transcription factors, DREB1 and DREB2, with an EREBP/AP2 DNA binding domain separate two cellular signal


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6 Urao, T. et al. (1999) A transmembranehybridtype histidine kinase in Arabidopsis functions as an osmosensor. Plant Cell 11, 1743–1754 7 Sanders, D. et al. (1999) Communicating with calcium. Plant Cell 11, 691–706 8 Knight, H. (2000) Calcium signaling during abiotic stress in plants. Int. Rev. Cytol. 195, 269–324 9 Ding, J.P. and Pickard, B.G. (1993) Modulation of mechanosensitive calcium-selective cation channels by temperature. Plant J. 3, 713–720 10 Minorsky, P.V. (1989) Temperature sensing by plants: a review and hypothesis. Plant Cell Environ. 12, 119–135 11 Plieth, C. et al. (1999) Temperature sensing by plants: the primary characteristics of signal perception and calcium response. Plant J. 18, 491–497 12 Murata, N. and Los, D.A. (1997) Membrane fluidity and temperature perception. Plant Physiol. 115, 875–879 13 Orvar, B.L. et al. (2000) Early steps in cold sensing by plant cells: the role of actin cytoskeleton and membrane fluidity. Plant J. 23, 785–794 14 McAinsh, M.R. and Hetherington, A.M. (1998) Encoding specificity in Ca2+ signalling systems. Trends Plant Sci. 3, 32–36 15 Dolmetsch, R.E. et al. (1997) Differential activation of transcription factors induced by Ca2+ response amplitude and duration. Nature 386, 855–858 16 Monroy, A.F. and Dhindsa, R.S. (1995) Lowtemperature signal transduction: induction of cold acclimation-specific genes of alfalfa by calcium at 25ºC. Plant Cell 7, 321–331 17 Sheen, J. (1996) Ca2+-dependent protein kinases and stress signal transduction in plants. Science 274, 1900–1902 18 Clayton, H. et al. (1999) Dissection of theozoneinduced calcium signature. Plant J. 17, 575–579 19 Allen, G.J. et al. (2000) Alteration of stimulusspecific guard cell calcium oscillations and stomatal closing in Arabidopsis det3 mutant. Science 289, 2338–2342 20 Knight, H. et al. (1996) Cold calcium signaling in Arabidopsis involves two cellular pools and a change in calcium signature after acclimation. Plant Cell 8, 489–503 21 Subbaiah, C.C. et al. (1998) Mitochondrial contribution to the anoxic Ca2+ signal in maize suspension-cultured cells. Plant Physiol. 118, 759–771 22 Knight, H. and Knight, M.R. (2000) Imaging spatial and cellular characteristics of low temperature calcium signature after cold acclimation in Arabidopsis. J. Exp. Bot. 51, 1679–1686 23 Kiegle, E. et al. (2000) Cell-type-specific calcium responses to drought, salt and cold in the Arabidopsis root. Plant J. 23, 267–278 24 Knight, H. et al. (1998) A history of stress alters drought calcium signalling pathways in Arabidopsis. Plant J. 16, 681–687 25 Braam, J. and Davis, R.W. (1990) Rain-, wind-, and touch-induced expression of calmodulin and calmodulin- related genes in Arabidopsis. Cell 60, 357–364 26 Botella, J.R. and Arteca, R.N. (1994) Differential expression of two calmodulin genes in response to physical and chemical stimuli. Plant Mol. Biol. 24, 757–766 27 Harding, S.A. et al. (1997) Transgenic tobacco expressing a foreign calmodulin gene shows an enhanced production of active oxygen species. EMBO J. 16, 1137–1144
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28 Heo, W.D. et al. (1999) Involvement of specific calmodulinisoforms in salicylic acid-independent activation of plant disease resistance responses. Proc. Natl. Acad. Sci. U. S. A. 96, 766–771 29 Mochly-Rosen, D. (1995) Localization of protein kinases by anchoring proteins: a theme in signal transduction. Science 268, 247–251 30 Harmon, A.C. et al. (2000) CDPKs – a kinase for every Ca2+ signal? Trends Plant Sci. 5, 154–159 31 Urao, T. et al. (1994) Two genes that encode Ca2+dependent protein kinases are induced by drought and high-salt stresses in Arabidopsis thaliana. Mol. Gen. Genet. 244, 331–340 32 Saijo, Y. et al. (2000) Over-expression of a single Ca2+-dependent protein kinase confers both cold and salt/drought tolerance on rice plants. Plant J. 23, 319–327 33 Luan, S. (1998) Protein phosphatases and signaling cascades in higher plants. Trends Plant Sci. 3, 271–275 34 Liu, J. and Zhu, J-K. (1998) A calcium sensor homolog required for plant salt tolerance. Science 280, 1943–1945 35 Kudla, J. et al. (1999) Genes for calcineurin B-like proteins in Arabidopsis are differentially regulated by stress signals. Proc. Natl. Acad. Sci. U. S. A. 96, 4718–4723 36 Shi, J. et al. (1999) Novel protein kinases associated with calcineurin B-like calcium sensors in Arabidopsis. Plant Cell 11, 2393–2406 37 Monroy, A.F. et al. (1998) Low temperature signal transduction during cold acclimation: protein phosphatase 2A as an early target for coldinactivation. Plant J. 13, 653–660 38 Meyer, K. et al. (1994) A protein phosphatase 2C involved in ABA signal transduction in Arabidopsis thaliana. Science 264,1452–1455 39 Leung, J. et al. (1994) Arabidopsis ABA response gene ABI1: features of a calcium-modulated protein phosphatase. Science 264, 1448–1452 40 Gosti, F. et al. (1999) ABI1 protein phosphatase 2C is a negative regulator of abscisic acid signaling. Plant Cell 11, 1897–1910 41 Sheen, J. (1998) Mutational analysis of protein phosphatase 2C involved in abscisic acid signal transduction in higher plants. Proc. Natl. Acad. Sci. U. S. A. 95, 975–980 42 Vranova, E. et al. (2000) Oxidative stress, heat shock and drought differentially affect expression of a tobacco protein phosphatase 2C. J. Exp. Bot. 51, 1763–1764 43 Ligterink, W. and Hirt, H. (2001) Mitogen-activated protein (MAP) kinase pathways in plants: versatile signaling tools. Int. Rev. Cytol. 201, 209–275 44 Mizoguchi, T. et al. (1996) A gene encoding a mitogen-activated protein kinase kinase kinase is induced simultaneously with genes for a mitogenactivated protein kinase and an S6 ribosomal protein kinase by touch, cold, and water stress in Arabidopsis thaliana. Proc. Natl. Acad. Sci. U. S. A. 93, 765–769 45 Kovtun, Y. et al. (2000) Functional analysis of oxidative stress-activated mitogen-activated protein kinase cascade in plant. Proc. Natl. Acad. Sci. U. S. A. 97, 2940–2945 46 Mizoguchi, T. et al. (1998) Identification of a possible MAP kinase cascade in Arabidopsis thaliana based on pairwise yeast two-hybrid analysis and functional complementation tests of yeast mutants. FEBS Lett. 437, 56–60 47 Ichimura, K. et al. (1998) Isolation of ATMEKK1 (a MAP kinase kinasekinase)-interacting

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proteins and analysis of a MAP kinase cascade in Arabidopsis. Biochem. Biophys. Res. Commun. 253, 532–543 Ichimura, K. et al. (2000) Various abiotic stresses rapidly activate Arabidopsis MAP kinases ATMPK4 and ATMPK6. Plant J. 24, 655–665 Thomashow, M.F. (1998) Role of cold-responsive genes in plant freezing tolerance. Plant Physiol. 118, 1–8 Mäntylä, E. et al. (1995) The role of abscisic acid in drought-induced freezing tolerance, cold acclimation, and accumulation of LTI78 and RAB18 proteins in Arabidopsis thaliana. Plant Physiol. 107, 141–148 Shinozaki, K. and Yamaguchi-Shinozaki, K. (2000) Molecular responses to dehydration and low temperature: differences and cross-talk between two stress signaling pathways. Curr. Opin. Plant Biol. 3, 217–223 Yamaguchi-Shinozaki, K. and Shinozaki, K. (1994) A novel cis-acting element in an Arabidopsis gene is involved in responsiveness to drought, low-temperature, or high-salt stress. Plant Cell 6, 251–264 Stockinger, E.J. et al. (1997) Arabidopsis thaliana CBF1 encodes an AP2 domain-containing transcriptional activator that binds to the Crepeat/DRE, a cis-acting DNA regulatory element that stimulates transcription in response to low temperature and water deficit. Proc. Natl. Acad. Sci. U. S. A. 94, 1035–1040 Nakashima, K. et al. (2000) Organization and expression of two Arabidopsis DREB2 genes encoding DRE-binding proteins involved in dehydration- and high-salinity-responsive geneexpression. Plant Mol. Biol. 42, 657–665 Shinozaki, K. and Yamaguchi-Shinozaki, K. (1996) Molecular responses to drought and cold stress. Curr. Opin. Biotechnol. 7, 161–167 Uno, Y. et al. (2000) Arabidopsis basic leucine zipper transcription factors involved in an abscisic acid-dependent signal transduction pathway under drought and high-salinity conditions. Proc. Natl. Acad. Sci. U. S. A. 97, 11632–11637 Knight, H. et al. (1999) The sfr6 mutation in Arabidopsis suppresses low-temperature induction of genes dependent on the CRT/DRE sequence motif. Plant Cell 11, 875–886 Ishitani, M. et al. (1997) Genetic analysis of osmotic and cold stress signal transduction in Arabidopsis: interactions and convergence of abscisic acid-dependent and abscisic acidindependent pathways. Plant Cell 9, 1935–1949 Lee, H. et al. (1999) Cold-regulated gene expression and freezing tolerance in an Arabidopsis thaliana mutant. Plant J. 17, 301–308 Ishitani, M. et al. (1998) HOS1, a genetic locus involved in cold-responsive gene expression in Arabidopsis. Plant Cell 10, 1151–1161 Xiong, L. et al. (1999) HOS5 – a negative regulator of osmotic stress-induced gene expression in Arabidopsis thaliana. Plant J. 19, 569–578 Xiong, L. et al. (1999) Interaction of osmotic stress, temperature, and abscisic acid in the regulation of gene expression in Arabidopsis. Plant Physiol. 119, 205–212 Seki, M. et al. (2001) Monitoring the expression pattern of 1300 Arabidopsis genes under drought and cold stresses using a full-length cDNA microarray. Plant Cell 13, 61–72