Cold, salinity and drought stresses: An overview - UAMuam.es/personal_pdi/ciencias/solsa/revsaltstress.pdf · Cold, salinity and drought stresses: An overview Shilpi Mahajan, Narendra

Archives of Biochemistry and Biophysics 444 (2005) 139–158

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Cold, salinity and drought stresses: An overview

Shilpi Mahajan, Narendra Tuteja ¤

Plant Molecular Biology, International Centre for Genetic Engineering and Biotechnology, Aruna Asaf Ali Marg, New Delhi 110067, India

Received 31 August 2005, and in revised form 14 October 2005Available online 9 November 2005

Abstract

World population is increasing at an alarming rate and is expected to reach about six billion by the end of year 2050. On the otherhand food productivity is decreasing due to the eVect of various abiotic stresses; therefore minimizing these losses is a major area of con-cern for all nations to cope with the increasing food requirements. Cold, salinity and drought are among the major stresses, whichadversely aVect plants growth and productivity; hence it is important to develop stress tolerant crops. In general, low temperature mainlyresults in mechanical constraint, whereas salinity and drought exerts its malicious eVect mainly by disrupting the ionic and osmotic equi-librium of the cell. It is now well known that the stress signal is Wrst perceived at the membrane level by the receptors and then transducedin the cell to switch on the stress responsive genes for mediating stress tolerance. Understanding the mechanism of stress tolerance alongwith a plethora of genes involved in stress signaling network is important for crop improvement. Recently, some genes of calcium-signal-ing and nucleic acid pathways have been reported to be up-regulated in response to both cold and salinity stresses indicating the presenceof cross talk between these pathways. In this review we have emphasized on various aspects of cold, salinity and drought stresses. Variousfactors pertaining to cold acclimation, promoter elements, and role of transcription factors in stress signaling pathway have beendescribed. The role of calcium as an important signaling molecule in response to various stress signals has also been covered. In each ofthese stresses we have tried to address the issues, which signiWcantly aVect the gene expression in relation to plant physiology. 2005 Elsevier Inc. All rights reserved.

Keywords: Calcium; CBL; CIPK; Cold; Drought; Helicase; Plants; Salt; SOS pathway; Stress

Plant growth and productivity is adversely aVected bynature’s wrath in the form of various abiotic and bioticstress factors. Plants are frequently exposed to a plethora ofstress conditions such as low temperature, salt, drought,Xooding, heat, oxidative stress and heavy metal toxicity.Various anthropogenic activities have accentuated theexisting stress factors. Heavy metals and salinity havebegun to accumulate in the soil and water tables and maysoon reach toxic levels. Plants also face challenges frompathogens including bacteria, fungi, and viruses as well asfrom herbivores. All these stress factors are a menace forplants and prevent them from reaching their full geneticpotential and limit the crop productivity worldwide. Abi-otic stress in fact is the principal cause of crop failure worldwide, dipping average yields for most major crops by more

* Corresponding author. Fax: +91 11 26162316.E-mail address: [email protected] (N. Tuteja).

0003-9861/$ - see front matter 2005 Elsevier Inc. All rights reserved.doi:10.1016/j.abb.2005.10.018

than 50% [1]. Abiotic stresses cause losses worth hundredsof million dollars each year due to reduction in crop pro-ductivity and crop failure. In fact these stresses, threatenthe sustainability of agricultural industry.

In response to these stress factors various genes are up-regulated, which can mitigate the eVect of stress and lead toadjustment of the cellular milieu and plant tolerance. Innature stress does not generally come in isolation and manystresses act hand in hand with each other. In response tothese stress signals that cross talk with each other, naturehas developed diverse pathways for combating and tolerat-ing them. These pathways act in cooperation to alleviatestress.

In this review we have Wrst emphasized cold stressfollowed by salt and drought stresses and the reason forthese stresses being injurious for plants. Various genesinvolved in cold acclimation and their role towards mem-brane stabilization have been discussed. The physiological

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140 S. Mahajan, N. Tuteja / Archives of Biochemistry and Biophysics 444 (2005) 139–158

parameters pertaining to each stress, various promoter ele-ments, transcription factors, negative regulators and therole of calcium in relation to cold and salinity stress havealso been covered. Furthermore, the role of SOS pathwayin imparting salt tolerance to plants and the role of glycinebetaine as a major osmolyte in response to salt stress andWnally the role of abscisic acid (ABA)1 in stress have alsobeen discussed.

What is stress?

Stress in physical terms is deWned as mechanical forceper unit area applied to an object. In response to theapplied stress, an object undergoes a change in the dimen-sion, which is also known as strain. As plants are sessile, it istough to measure the exact force exerted by stresses andtherefore in biological terms it is diYcult to deWne stress. Abiological condition, which may be stress for one plant maybe optimum for another plant. The most practical deWni-tion of a biological stress is an adverse force or a condition,which inhibits the normal functioning and well being of abiological system such as plants [2].

Various stress elicitors

A cell is separated from its surrounding environment bya physical barrier, which is the plasma membrane. Thismembrane is permeable to only some small lipid moleculessuch as steroid hormones, which can diVuse through themembrane into the cytoplasm and is impermeable to thewater-soluble material including ions, proteins and othermacromolecules. The cellular responses are initiated pri-marily by interaction of the extracellular material with aplasma membrane protein. This extracellular molecule iscalled a ligand (or an elicitor) and the plasma membraneprotein, which binds and interacts with this molecule, iscalled a receptor. Various stress signals both abiotic as wellas biotic serve as elicitors for the plant cell (see Table 1).

Stress signaling pathways an overview

The stress is Wrst perceived by the receptors present onthe membrane of the plant cells (Fig. 1A), the signal is thentransduced downstream and this results in the generationof second messengers including calcium, reactive oxygenspecies (ROS) and inositol phosphates. These second mes-

1 Abbreviations used: ABA, abscisic acid; ROS, reactive oxygen species;LEA, late embryogenesis abundant; MAP, mitogen-activated protein;DRE, dehydration responsive elements; ABRE, ABA-responsive element;ICE1, inducer of CBF expression 1; CDPKs, calcium-dependent proteinkinases; GA, gibberellin; SOS, salt overly sensitive; SNF, sucrose non-fer-menting kinases; PS II, photosystem II; PQ, plastoquinone; GR, glutathi-one reductase; APX, ascorbate peroxidase; P5CS, pyrroline-5-carboxylatesynthase; HSPs, heat shock proteins; smHS, small HS; DAG, diacylglycer-ol; PA, phosphatidic acid; PLC, phospholipase C; PLD, phospholipase D;CaM, calmodulin; CBL, calcineurin B-like; CIPK, CBL-interacting pro-tein kinase.

sengers, such as inositol phosphates, further modulate theintracellular calcium level. This perturbation in cytosolicCa2+ level is sensed by calcium binding proteins, alsoknown as Ca2+ sensors. These sensors apparently lack anyenzymatic activity and change their conformation in a cal-cium dependent manner. These sensory proteins then inter-act with their respective interacting partners often initiatinga phosphorylation cascade and target the major stressresponsive genes or the transcription factors controllingthese genes. The products of these stress genes ultimatelylead to plant adaptation and help the plant to survive andsurpass the unfavorable conditions. Thus, plant responds tostresses as individual cells and synergistically as a wholeorganism. Stress induced changes in gene expression in turnmay participate in the generation of hormones like ABA,salicylic acid and ethylene. These molecules may amplifythe initial signal and initiate a second round of signalingthat may follow the same pathway or use altogether diVer-ent components of signaling pathway. Certain moleculesalso known as accessory molecules may not directly partici-pate in signaling but participate in the modiWcation orassembly of signaling components. These proteins includethe protein modiWers, which may be added cotranslation-ally to the signaling proteins like enzymes for myristoyla-tion, glycosylation, methylation and ubiquitination.

The various stress responsive genes can be broadly cate-gorized as early and late induced genes (Fig. 1B). Earlygenes are induced within minutes of stress signal perceptionand often express transiently. Various transcription factorsare included in the list of early genes as the induction ofthese genes does not require synthesis of new proteins andsignaling components are already primed. In contrast, mostof the other genes, which are activated by stress moreslowly, i.e. after hours of stress perception are included inthe late induced category. The expression of these genes isoften sustained. These genes include the major stressresponsive genes such as RD (responsive to dehydration)/KIN (cold induced)/COR (cold responsive), which encodesand modulate the proteins needed for synthesis, for

Table 1Various abiotic as well as biotic stress signals for plants

Abiotic stresses1. Cold (chilling and frost)2. Heat (high temperature)3. Salinity (salt)4. Drought (water deWcit condition)5. Excess water (Xooding)6. Radiations (high intensity of ultra-violet and visible light)7. Chemicals and pollutants (heavy metals, pesticides, and aerosols)8. Oxidative stress (reactive oxygen species, ozone)9. Wind (sand and dust particles in wind)

10. Nutrient deprivation in soil

Biotic stresses1. Pathogens (viruses, bacteria, and fungi)2. Insects3. Herbivores4. Rodents

S. Mahajan, N. Tuteja / Archives of Biochemistry and Biophysics 444 (2005) 139–158 141

Fig. 1. (A and B) Generic signal transduction pathway as well as the expres-sion of early and late genes in response to abiotic stress signaling. (A) Repre-sents the overview of signaling pathway under stress condition. Stress signal isWrst perceived by the membrane receptor, which activates PLC and hydrolysesPIP2 to generate IP3 as well as DAG. Following stress, cytoplasmic calciumlevels are up-regulated via movements of Ca2+ ions from apoplast or from itsrelease from intracellular sources mediated by IP3. This change in cytoplasmicCa2+ level is sensed by calcium sensors which interact with their down streamsignaling components which may be kinases and/or phosphatases. These pro-teins aVect the expression of major stress responsive genes leading to physio-logical responses. (B) Early and delayed gene expression in response to abioticstress signaling. Various genes are triggered in response to stress and can begrouped under early and late responsive genes. Early genes are induced withinminutes of stress perception and often express transiently. In contrast, variousstress genes are activated slowly, within hours of stress expression and oftenexhibit a sustained expression level. Early genes encode for the transcriptionfactors that activate the major stress responsive genes (delayed genes). Theexpression of major stress genes like RD/KIN/COR/RAB18/RAB29B resultin the production of various osmolytes, antioxidants, molecular chaperonesand LEA-like proteins, which function in stress tolerance.

Receptors

PLC

Ca2+ Sensors

PIP2 IP3 + DAG

Ca2 +

and othersecond messengers

[Ca2 +]ext

(InsP, ROS)

(CBLs/CaM…)

Kinases/PhosphatasesCIPKs/SOS2, CDPKs,MAPKs and variousprotein phosphatases

Transcription factors

~PO4/de~PO4

Major stress responsive genes

Physiological response

Stress

EARLY GENES DELAYED GENES(RD/KIN/COR/RAB18/RAB29B)

Encode proteins like transcription factors/

calcium sensors.

STRESS TOLERANCE EFFECTORSe.g. LEA like proteins, antioxidants

osmolyte synthesiszing enzymes

ACTIVATE

MODULATE

Stress Responsive Genes

A

B

example LEA-like proteins (late embryogenesis abundant),antioxidants, membrane stabilizing proteins and synthesisof osmolytes.

Cold stress

In this section, we have emphasized on various aspectsof cold stress, which includes aVect of cold on plants physi-ology, cold acclimation and its role in providing freeze-tol-erance, function of cold-regulated genes in coldacclimation, negative regulation of cold stress and the roleof calcium in relation to cold stress. All these topics wouldhelp in our better understanding of cold induced cellularchanges and its aVect on gene expression.

AVect of cold on plants physiology

Each plant has its unique set of temperature require-ments, which are optimum for its proper growth and devel-opment. A set of temperature conditions, which areoptimum for one plant may be stressful for another plant.Many plants, especially those, which are native to warmhabitat, exhibit symptoms of injury when exposed to lownon-freezing temperatures [3]. These plants including maize(Zea mays), soybean (Glycine max), cotton (Gossypiumhirsutum), tomato (Lycopersicon esculentum) and banana(Musa sp.) are in particular sensitive to temperatures below10–15 °C and exhibit signs of injury see [3–5]. The symp-toms of stress induced injury in these plants appear from 48to 72 h, however, this duration varies from plant to plantand also depend upon the sensitivity of a plant to coldstress. Various phenotypic symptoms in response to chillingstress include reduced leaf expansion, wilting, chlorosis(yellowing of leaves) and may lead to necrosis (death of tis-sue). Chilling also severely hampers the reproductive devel-opment of plants for example exposure of rice plants tochilling temperature at the time of anthesis (Xoral opening)leads to sterility in Xowers [6].

The major malicious eVect of freezing is that it inducessevere membrane damage [7,8]. This damage is largely dueto the acute dehydration associated with freezing. Mem-brane lipids are primarily composed of two kinds of fattyacids unsaturated as well as saturated fatty acids. Unsatu-rated fatty acids have one or more double bonds betweentwo carbon atoms (ACHBCHA) whereas saturated fattyacids are fully saturated with hydrogen atoms(ACH2ACH2A). It is a well-known fact that lipids con-taining saturated fatty acids solidify at temperatures higherthan those containing unsaturated fatty acids. Therefore,the relative proportion of unsaturated fatty acids in themembrane strongly inXuences the Xuidity of the membrane[8]. The temperature at which a membrane changes fromsemi Xuid state to a semi crystalline state is known as thetransition temperature. Chilling sensitive plants usuallyhave a higher proportion of saturated fatty acids and,therefore, a higher transition temperature. Chilling resistantspecies on the other hand are marked by higher proportion


of unsaturated fatty acids and correspondingly a lowertransition temperature.

The success of many crops rests on their ability to with-stand the freezing temperature of late spring or earlyautumn frost. Therefore tolerance to freezing temperaturesis in particular important for the sustainability of agricul-tural crops. As understanding the basics of a disease isessential for its cure, in the same way understanding of howfreezing induces its injurious eVects on plants is essential forthe development of frost tolerant crops. The real cause offreeze-induced injury to plants is the ice formation ratherthan low temperatures. It is noteworthy to mention herethat dehydrated tissues such as seeds and fungal spores cansurvive at very low temperatures without any symptoms ofinjury. Even cryopreservation is a common method forstorage of seeds and other biological materials, which isbased on the fact that water essentially solidiWes withoutthe formation of ice crystals.

Ice formation in plants, begins in the apoplastic space asit has relatively lower solute concentration. As the vaporpressure of ice is much lower than water at any given tem-perature, ice formation in the apoplast establishes a vaporpressure gradient between the apoplast and surroundingcells. The unfrozen cytoplasmic water migrates down thegradient from the cell cytosol to the apoplast, which con-tributes to the enlargement of existing ice crystals andcauses a mechanical strain on the cell wall and plasmamembrane leading to cell rupture [9,10]. Freeze induced cel-lular dehydration results in multiple forms of membranedamage including expansion-induced-cell lyses and fracturelesions [8,11] and lamellar-to-hexagonal-II phase transition.Although freeze exerts its eVect largely by membrane dam-age due to severe cellular dehydration, certain additionalfactors may also contribute to damage induced by freeze.

ROS produced in response to freeze stress contributes tomembrane damage. Chilling sensitive plants characteristi-cally exhibits structural injuries and may suVer from meta-bolic dysfunction when chilled [12]. Overall, chillingultimately results in loss in membrane integrity, which leadsto solute leakage. The integrity of intracellular organelles isalso disrupted leading to the loss of compartmentalization,reduction and impairing of photosynthesis, protein assem-bly and general metabolic processes. The primary environ-mental factors responsible for triggering increasedtolerance against freezing, is the phenomenon known as‘cold acclimation.’ It is the process where certain plantsincrease their freezing tolerance upon prior exposure to lownon-freezing temperatures.

Cold acclimation and its role in providing freeze-tolerance

The primary function of cold acclimation is to stabilizethe membranes against freeze injury. Acclimation results inincrease in proportion of unsaturated fatty acids andthereby a drop in transition temperature [13,14]. It func-tions to prevent the expansion-induced lyses and formationof hexagonal II phase lipids in rye and other plants [8,11].

Cold acclimation results in physical and biochemicalrestructuring of cell membranes through changes in thelipid composition and induction of other non-enzymaticproteins that alter the freezing point of water. Addition ofsolutes decreases the freezing point of water to a more neg-ative value, thus preventing ice formation.

Low temperatures induce a number of alterations in cel-lular components, including the extent of unsaturated fattyacids [15], the composition of glycerolipids [16], changes inprotein and carbohydrate composition and the activationof ion channels [17]. Accumulation of sucrose and othersimple sugars that occurs with cold acclimation also con-tributes to the stabilization of membrane as these moleculescan protect membranes against freeze-damage. Freezingtolerance is a multigenic trait. Low temperatures activate anumber of cold-inducible genes [18], such as those thatencode dehydrins, lipid transfer proteins, translation elon-gation factors and the late-embryogenesis-abundant pro-teins [19]. Moreover, intercellular ice formation can cause amechanical strain on cell wall and membrane leading to cellrupture [9,10]. There is also substantiation that proteindenaturation occurs in plants at low temperature whichcould also result in cellular damage [20].

Overall, cold acclimation results in protection and stabil-ization of the integrity of cellular membranes, enhancementof the antioxidative mechanisms, increased intercellularsugar levels as well as accumulation of other cryoprotec-tants including polyamines that protect the intracellularproteins by inducing the genes encoding molecular chaper-ones [21]. All these modiWcations help the plant to with-stand and surpass the severe dehydration associated withfreezing stress.

Function of cold-regulated genes in cold acclimation

Considerable eVorts have been directed towards deter-mining the nature of cold-inducible genes and establishingtheir role in freezing tolerance. The Arabidopsis FAD8 gene[22] encodes a fatty acid desaturase that contributes tofreezing tolerance by altering the lipid composition.

Cold-responsive genes encoding molecular chaperonesincluding a spinach hsp70 gene [23], and a Brassica napushsp90 gene [24], contribute to freezing tolerance by stabiliz-ing proteins against freeze-induced denaturation. Manycold-responsive genes encoding various signal transductionand regulatory proteins have been identiWed and this listincludes the mitogen-activated protein (MAP) kinase [25],MAP kinase, kinase, kinase (MAPKKK) [26] and the cal-modulin-related proteins [27]. These proteins might con-tribute to freezing tolerance as well as tolerance to otherstresses by controlling or regulating the expression andactivity of the major stress genes as well their proteins.

The largest class of cold induced genes encodes polypep-tides that are homologs of LEA proteins and the polypep-tides that are synthesized during the late embryogenesisphase, just prior to seed desiccation and also in the seed-lings in response to dehydration stress [28–30]. These LEA


like proteins are mainly hydrophilic, many have relativelysimple amino-acid composition, and are composed largelyof a few amino acids with repeated amino acid sequencemotifs. Many of these proteins are predicted to containregions capable of forming amphipathic � helices. Theexamples of cold responsive genes include: COR15a, [31],alfalfa Cas15 [32], and wheat WCS120 [33]. The expressionof COR genes has been shown to be critical for both chill-ing tolerance and cold acclimation in plants [34]. Arabidop-sis COR genes include: COR78/RD29, COR47, COR15a,COR6.6 and encode LEA like proteins [34]. These genes areinduced by cold, dehydration or ABA. COR15A polypep-tide is targeted to the chloroplast. Formation of hexagonalII phase lipids is a major cause of membrane damage innon-acclimated plants. COR15a expression decreases thepropensity of the membranes to form hexagonal II phaselipids in response to freezing [8,11].

The analysis of the promoter elements of COR genesrevealed that they contain DRE (dehydration responsiveelements) or CRT (C-repeats) and some of them containABRE (ABA-responsive element) as well [35,36]. Inductionof the COR genes was accomplished by over-expression oftranscription factor CBF (CRT/DRE binding factor) [36].CBF binds to the CRT/DRE elements present in the pro-moter of the COR genes and other cold-regulated genes.The over-expression of these regulatory elements not onlyresulted in increased freezing tolerance but also an increaseto drought tolerance [37]. This Wnding provides strong sup-port that a fundamental role of cold-inducible genes is toprotect the plant cells against cellular dehydration. Leeet al. [38] genetically analyzed HOS1 (high expression ofosmotically responsive genes) locus of Arabidopsis. Thehos1 mutation resulted in sustained and super induction ofCBF2, CBF3 and their target regulatory genes during coldstress. Therefore, HOS1 was identiWed as a negative regula-tor of COR genes by modulating the expression level ofCBFs. [39]. HOS1 gene encodes a ring Wnger protein and isconstitutively expressed but gets drastically down-regulatedwithin 10 min of cold stress. Genetic analysis led to theidentiWcation of ICE1 (inducer of CBF expression 1) as anactivator of CBF3 [39]. ICE1 encoded a transcription factorthat speciWcally recognized MYC sequence on the CBF3promoter. Transgenic lines overexpressing ICE1 did notexpress CBF3 at warm temperature but showed a higherlevel of expression for CBF3 as well as RD29 and COR15aat low temperatures. This study suggests that cold inducedmodiWcation of ICE1 is necessary for it to act as an activa-tor of CBF3 in planta.

Recently two CBF1-like cDNAs CaCBFIA and CaCB-FIB have been cloned and characterized [40] from hot pep-per. These were induced in response to low temperaturestress (4 °C) and not in response to wounding or ABA.Two-hybrid screening led to the isolation of a homeodo-main leucine zipper (4D-Zip) protein that interacts withCaCBFIB. The expression of 4D-Zip was elevated by lowtemperature and drought [40]. Calcium-dependent proteinkinases (CDPKs) play an important role in the signal trans-

duction and recently the function of OsCDPK13 (Oryzasativa CDPK 13) has been characterized [41]. The geneexpression as well as protein accumulation of OsCDPK13were up-regulated in response to cold and gibberellin (GA)but suppressed under salt and drought stress and also inresponse to ABA. The overexpressing transgenic lines ofOsCDPK13 had higher recovery rates following cold stressin comparison with the vector control rice. Cold-tolerantrice varieties exhibited higher expression of OsCDPK13than the cold sensitive ones. Antisense OsCDPK13 trans-genic lines were shorter in comparison with the vector con-trol lines. Moreover, dwarf mutants of rice also had lowerlevel of OsCDPK13 than in wild type [41]. However, therehas been no mention of the sensitivity of OsCDPK13 anti-sense lines in response to cold stress [41]. We howeverexpect that these antisense lines should be hypersensitive tocold stress as the gene has been shown to play an importantrole in mediating tolerance in response to cold stress whichis evident due to higher recovery rates following cold stressthan the vector control lines.

Negative regulation of cold stress

Mutagenesis study resulted in the identiWcation of agene, eskimo l (esk1), which has a major eVect on freezingtolerance. These plants were more freeze tolerant than thewild type plants without cold acclimation. The concentra-tion of free proline [42] in the esk1 mutant was found to be30-fold higher than in the wild-type plants. Proline has beenshown to be an eVective cryoprotectant and this is also oneof the major factors imparting freezing tolerance. In addi-tion to the total sugars, which were elevated, the expressionof RAB18 cold-responsive LEA II gene was also found tobe elevated three fold. This suggests that ESK1 may act as anegative regulator. SigniWcantly, the esk 1 mutation did notappear to aVect the expression of COR genes. This suggeststhat multiple signaling pathways are involved in responseto cold stress and they may cross talk with each other aswell as with genes involved in other stresses.

Role of calcium in relation to cold stress

Calcium is an important messenger in a low temperaturesignal transduction pathway. The change in cytosolic cal-cium levels is a necessary Wrst step in a temperature sensingmechanism, which enables the plant to withstand futurecold stress in a better way. In both Arabidopsis [17,27] andalfalfa [43] cytoplasmic calcium levels increase rapidly inresponse to low temperature, largely due to an inXux of cal-cium from extracellular stores. Through the use of pharma-cological and chemical reagents, it has been demonstratedthat calcium is required for the full expression of some ofthe cold induced genes including the CRT/DRE controlledCOR6 and KIN1 genes of Arabidopsis [17,32,43]. For exam-ple, Ca2+ chelators such as BAPTA and Ca2+ channelblockers such as La3+ inhibited the cold-induced inXux ofcalcium and resulted in the decreased expression of the cold


inducible Cas15 gene and blocked the ability of alfalfa toacclimate in cold. In addition Cas15 expression can beinduced at a much higher temperature, i.e., 25 °C by treatingthe cells with A23187, a Ca2+ ionophore that causes a rapidinXux of calcium [43].

Salinity stress

Salinity is a major environmental stress and is a substan-tial constraint to crop production. Increased salinization ofarable land is expected to have devastating global eVects,resulting in 30% land loss within next 25 years and up to50% by the middle of 21st century [44]. High salinity causesboth hyperionic and hyperosmotic stress and can lead toplant demise. Sea water contains approximately 3% ofNaCl and in terms of molarity of diVerent ions, Na+ isabout 460 mM, Mg2+ is 50 mM and Cl¡ around 540 mMalong with smaller quantities of other ions. Salinity in agiven land area depends upon various factors like amountof evaporation (leading to increase in salt concentration),or the amount of precipitation (leading to decrease in saltconcentration). Weathering of rocks also aVects salt con-centration. Inland deserts are marked by high salinity as therate of evaporation far exceeds the rate of precipitation.Agricultural lands that have been heavily irrigated arehighly saline. As drier areas in particular need intense irri-gation, there is extensive water loss through a combinationof both evaporation as well as transpiration. This process isknown as evapotranspiration and as a result, the saltdelivered along with the irrigation water gets concentrated,year-by-year in the soil. This leads to huge losses in terms ofarable land and productivity as most of the economicallyimportant crop species are very sensitive to soil salinity.These salt sensitive plants, also known as glycophytesinclude rice (Oryza sativa), maize (Zea mays), soybean (Gly-cine max) and beans (Phaseolus vulgaris). High salt concen-tration (Na+) in particular which deposit in the soil canalter the basic texture of the soil resulting in decreased soilporosity and consequently reduced soil aeration and waterconductance. The basic physiology of high salt stress anddrought stress overlaps with each other. High salt deposi-tions in the soil generate a low water potential zone in thesoil making it increasingly diYcult for the plant to acquireboth water as well as nutrients. Therefore, salt stress essen-tially results in a water deWcit condition in the plant andtakes the form of a physiological drought. The major ionsinvolved in salt stress signaling, include Na+, K+, H+ andCa2+. It is the interplay of these ions, which brings homeo-stasis in the cell.

In this section, we have emphasized on various aspectsof salinity stress, which includes the reasons why salinitystress is injurious to plant cells, generic function of K+, roleof Ca2+ and SOS pathway in relation to imparting saltstress tolerance, loss of water due to salinity stress and therole of glycine betaine as a major osmolyte. Moreover, therole of DNA unwinding enzymes, i.e., helicases, impartingsalinity stress tolerance have also been discussed.

Maladies caused by salt stress on plant cells arise from the following

(1) Disruption of ionic equilibrium: InXux of Na+ dissi-pates the membrane potential and facilitates theuptake of Cl¡ down the chemical gradient.

(2) Na+ is toxic to cell metabolism and has deleteriouseVect on the functioning of some of the enzymes [45].

(3) High concentrations of Na+ causes osmotic imbal-ance, membrane disorganization, reduction ingrowth, inhibition of cell division and expansion.

(4) High Na+ levels also lead to reduction in photosyn-thesis and production of reactive oxygen species [46–48].

Where sodium (Na+) is deleterious for plant growth, K+

is one of the essential elements and is required by the plantin large quantities.

Generic functions of K+

(1) K+ is required for maintaining the osmotic balance.(2) K+ has a role in opening and closing of stomata.(3) K+ is an essential co-factor for many enzymes like the

pyruvate kinase, whereas Na+ is not.

Movement of salt into roots and to shoots is a productof the transpirational Xux required to maintain the waterstatus of the plant [48,49]. As common proteins transportNa+ and K+, Na+ competes with K+ for intracellular inXux[45,50,51]. Many K+ transport systems have some aYnityfor Na+, i.e., Na+/K+ symporters. Thus external Na+ nega-tively impacts intracellular K+ inXux. Most cells maintainrelatively high K+ and low concentrations of Na+ in thecytosol. This is achieved through a coordinated regulationof transporters for H+, K+, Ca2+ and Na+.

The plasma membrane H+-ATPases serves as the pri-mary pump that generates a proton motive force drivingthe transport of other solutes including Na+ and K+.Increased ATPase-mediated H+ translocation across theplasma membrane is a component of the plant cell responseto salt imposition [52,53]. K+ and Na+ inXux can be diVer-entiated physiologically into two categories, one with highaYnity for K+ over Na+ and the other for which there islower K+/Na+ selectivity. The Na+/K+ transporter and K+

transporters with dual high and low aYnity may contributesubstantially to Na+ inXux.

Role of Ca2+ in relation to salt stress

For decades it has been shown that another ion, Ca2+

has role in providing salt tolerance to plant. Externallysupplied Ca2+ reduces the toxic eVects of NaCl, presum-ably by facilitating higher K+/Na+ selectivity [54–56].High salinity results in increased cytosolic Ca2+ that istransported from the apoplast as well as the intracellularcompartments [57]. This transient increase in cytosolic


Ca2+ initiates the stress signal transduction leading to saltadaptation.

The search to identify genes involved in providing salttolerance commenced in 1998, by Liu and Zhu [56] whereseveral mutants were screened and SOS (salt overly sensi-tive) genes were identiWed through positional cloning.BrieXy, SOS pathway results in the exclusion of excess Na+

ions out of the cell via the plasma membrane Na+/H+ anti-porter and helps in reinstating cellular ion homeostasis. Thediscovery of SOS genes paved the way for elucidation of anovel pathway linking the Ca2+ signaling in response to asalt stress [58,59].

SOS3 gene encodes a Ca2+ binding protein with 4 EFhand Ca2+ binding motifs and a myristoylation sequence(MGXXXST/K) at the N-terminus of the protein. Inresponse to Ca2+ perturbation SOS3 changes its conforma-tion and transduces the signal downstream by interactingwith an eVector kinase. Mutation in SOS3 (sos 3-1), whichresults in the reduction of its Ca2+ binding ability alsoimpairs the cellular ionic equilibrium and renders the planthypersensitive to salt stress [60]. This defect can be partiallyrescued by addition of high levels of Ca2+ in the growthmedium [56]. Ca2+ sensors diVer in their aYnity with whichthey bind Ca2+ and this diVerence is an important parame-ter in distinguishing and decoding various Ca2+ sensors. Incomparison with Ca2+ sensors like calmodulin and caltrac-tin, SOS3 binds Ca2+ with a relatively low aYnity.

SOS2 gene was isolated through the genetic screeningof mutants oversensitive to salt stress in Arabidopsis. ThemRNA level of SOS2 was shown to be up-regulated inresponse to salt stress in the roots [61]. SOS2/CIPK24encodes a novel serine/threonine protein kinase with anN terminal catalytic and C terminal regulatory domain.Whereas the N terminal domain shares sequence homol-ogy with sucrose non-fermenting kinases (SNF), the Cterminal domain is unique to this class of kinases andharbors a 21 amino acid FISL/NAF motif [62]. FISLmotif acts as an autoinhibitory domain and interactswith the catalytic domain thereby keeping the enzyme inan OFF state under normal conditions. SOS3 interactswith SOS2 via FISL motif and relieves the protein fromautoinhibition thereby making the kinase active. SOS3activates SOS2 protein kinase activity in a calcium-dependent manner [63]. SOS2 could be constitutivelyactivated by the deletion of FISL motif [64] and this dele-tion resulted in SOS2 acting independent of SOS3. Ara-bidopsis plants with double mutant genotype (sos3/sos2)showed no additive eVects towards salt sensitivity, thisindicates that SOS3 and SOS2 function in the same path-way [63]. Constitutively over-expressed SOS2 under thecontrol of CaMV35S promoter could rescue the salthypersensitive phenotype of both sos3 and sos2 mutants,thereby further supporting the functioning of SOS3 andSOS2 in the same Ca2+ mediated pathway during saltstress [59,65].

SOS1 gene was identiWed as the target of SOS3–SOS2pathway by genetic analysis of sos1 mutants of Arabidopsis.

Osmotic as well as ionic balance was impaired in sos1mutants and they exhibited hypersensitivity towards saltstress. SOS genes (SOS1, SOS2 and SOS3) were geneticallyconWrmed to function in a common pathway of salt toler-ance [58]. SOS1 gene was cloned and predicted to encode a127-kDa protein with a N terminal region composed of 12trans-membrane domains and a C terminal region with along hydrophilic cytoplasmic tail [66]. The trans-membraneregion of SOS1 shared substantial sequence homology tothe plasma membrane Na+/H+ antiporter isolated frombacteria and fungi [66].

The SOS pathway is depicted in Fig. 2. The perception ofsalt stress by an unknown hypothetical plasma membranesensor elicits cytoplasmic Ca2+ perturbations. This pertur-bation in the cytosolic Ca2+ levels is sensed by SOS3, whichtransduces the signal to the down stream components. Themyristoylation motif of SOS3 results in the recruitment ofSOS3–SOS2 complex to the plasma membrane, whereSOS2 phosphorylates and activates SOS1 (a plasma mem-brane Na+/H+ antiporter) [67]. The excess Na+ ions areexpelled out of the cell and cellular ion homeostasis isrestored. SOS pathway regulates Na+ ion homeostasis byinteracting with other regulatory proteins and seems tohave additional branches. AtHKT1 is a low aYnity Na+

transporter and seems to mediate Na+ entry into the rootcells of Arabidopsis during a salt stress [68]. Remarkably,mutation in Athkt1 also suppresses the sos3 mutation [69]suggesting that SOS3–SOS2 complex functions to downregulate HKT1 gene expression or inactivate the HKT1protein during salt stress, thereby preventing the Na+ entryand its build up in the cell [59]. SOS3 and SOS2 seem tonegatively regulate the activity of AtHKT1 under saltstress.

In addition to controlling SOS1 activity resulting ineZux of excess Na+ ions, SOS3–SOS2 complex also seemsto function in sequestration of excess Na+ ions in the intra-cellular compartments. SOS2 is shown to interact with vac-uolar Na+/H+ antiporter and inXuence the Na+/H+

exchange activity signiWcantly [70]. Recently, further crosstalk in the SOS pathway was explored and it was shownthat SOS2 interacted with the N terminus of CAX1 (H+/Ca2+) antiporter and regulated its activity [65]. This activa-tion of CAX1 via SOS2 was however independent of SOS3and resulted in maintenance of Ca2+ homeostasis. SOSpathway may also inXuence the functioning of other mem-brane proteins in sequestration of excess Na+ ions in othersub-cellular compartments.

Overall, osmotic homeostasis after salt stress is mediatedby Na+ eZux across the plasma membrane and/or by itscompartmentalization into the vacuoles. The energy forthese reactions is provided by H+-ATPases that serve asprimary pumps. Plant cDNAs encoding NHE (Na+/H+

exchanger)-like proteins similar to mammalian sodium/proton exchangers were isolated and can functionally com-plement a yeast mutant deWcient for the endomembraneNa+/H+ transporter, NHX1 [71,72]. The AtNHX1 geneencodes a tonoplast Na+/H+ antiporter and functions in


compartmentalizing excess Na+ into the vacuole [72]. Over-expression of AtNHX1 antiporter substantially enhancedsalt tolerance of Arabidopsis [71].

Loss of water due to salinity stress

A major consequence of NaCl stress is the loss of intra-cellular water. To prevent this water loss from the cell andprotect the cellular proteins, plants accumulate manymetabolites that are also known as “compatible solutes.”These solutes do not inhibit the normal metabolic reactions[73,74]. Frequently observed metabolites with an osmolytefunction are sugars, mainly fructose and sucrose, sugaralcohols and complex sugars like trehalose and fructans. Inaddition charged metabolites like glycine betaine prolineand ectoine are also accumulated. The accumulation ofthese osmolytes, facilitate the osmotic adjustment [75–77].Water moves from high water potential to low water poten-tial and accumulation of these osmolytes make the waterpotential low inside the cell and prevent the intracellularwater loss.

Role of glycine-betaine

Glycine betaine (N,N,N-trimethylglycine-betaine) is amajor osmolyte [78,79] and is synthesized by many plantsin response to abiotic stresses. Biosynthetic pathway ofbetaine is a two-step oxidation of choline. Recently, a bio-synthetic pathway of betaine from glycine, catalyzed bytwo N-methyl transferase enzymes, was found [80]. Thepotential role of N-methyl transferase gene for betainesynthesis has been examined in Synechococcus sp. a freshwater cyanobacteria, and in Arabidopsis. It has beenfound that the co-expression of N-methyl transferase genein cyanobacteria caused accumulation of betaine in sig-niWcant amounts and conferred salt tolerance to a freshwater cyanobacterium suYcient for it to become capableof growth in seawater [80]. Arabidopsis plants expressingN-methyltransferase gene also accumulated betaine tohigh levels and improved seed yield under stress condi-tions [80].

On the whole, plants possess speciWc mechanisms toovercome the hypersaline environment and thrive in such

Fig. 2. Regulation of ion homeostasis by SOS and related pathways in relation to salt stress adaptation. Salt stress is perceived by an unknown receptor (?)present at the plasma membrane (PM) of the cell. This induces a cytosolic calcium perturbation, which is sensed by SOS3 and accordingly changes its con-formation in a Ca2+-dependent manner and interacts with SOS2. This interaction relieves SOS2 of its auto-inhibition and results in activation of theenzyme. Activated SOS2, in complex with SOS3 phosphorylates SOS1, a Na+/H+ antiporter resulting in eZux of excess Na+ ions. SOS3–SOS2 complexinteracts with and inXuences other salt mediated pathways resulting in ionic homeostasis. This complex inhibits HKT1 activity (a low aYnity Na+ trans-porter) thus restricting Na+ entry into the cytosol. SOS2 also interacts and activates NHX (vacuolar Na+/H+ exchanger) resulting in sequestration ofexcess Na+ ions, further contributing to Na+ ion homeostasis. CAX1 (H+/Ca+ antiporter) has been identiWed as an additional target for SOS2 activity rein-stating cytosolic Ca2+ homeostasis.

Salt Sensor?

Ca2+ Increase

SOS3 (CBL)

SOS3 + SOS2

NHX HKT SOS1

Na+ in vacuoles Na+ entry blocked Na+ efflux-PM

SOS2 (CIPK)

?

S A L I N I T Y T O L E R A N C E

TRANSPORTERS

SALT STRESS(excess Na+)

LOW CYTOPLASMIC Na+

(PM-Na+/H+ anti-porter)(V-Na+/H+ exchanger) (Low affinity Na+ transporter)

(Ca2+ Sensor)

Motor proteins?

The ionic aspect of salt stress signaled via SOS pathway

?

CAX1(V-Ca2+/H+ antiporter)

Ca2+ homeostasis


conditions by adjusting their internal osmotic status. Thesemechanisms have already been discussed brieXy andinclude: exclusion of Na+ from cell by plasma membraneNa+/H+ antiporter, sequestration of excess Na+ in vacuolesby tonoplast Na+/H+ antiporters and accumulation oforganic, compatible solutes such as sugars, certain aminoacids and glycine betaine.

Role of helicases in imparting salinity stress tolerance

Abiotic stress condition often aVects the cellular gene-expression machinery. Therefore, the molecules that areinvolved in the processing of nucleic acids including heli-cases are also likely to be aVected. Multiple DNA heli-cases are present in the cell and are involved in generegulation at various developmental stages as well as instress conditions. These DNA unwinding enzymes mayhave diVerent substrates as well as structural require-ments [81,82]. Though a number of diVerent helicaseshave been reported from E. coli, bacteriophages, viruses,yeast, calf thymus and humans the biological role of onlya few DNA helicases have been explored [83–85]. More-over, our knowledge about plant DNA helicases has alsobeen limited with only 6 helicase proteins having beenpuriWed [82]. The role of helicases and the underlyingmolecular mechanisms is only beginning to be under-stood.

Recently the potential role of PDH45 (pea DNA heli-case 45) in overcoming salinity stress was explored [86]. Theauthors have proved that PDH45 overexpressing trans-genic lines showed high salinity tolerance and the T1 trans-genic plants were able to grow to maturity and set normalviable seeds under continuous salinity stress without anyreduction in plant yield in terms of seed weight. Theauthors have proposed a dual mode of action for PDH45.(i) PDH45 may act at the translation level to stabilize orenhance protein synthesis. As a support to this hypothesis itwas earlier proved that antibodies against PDH45 inhibitedthe protein synthesis in vitro, suggesting its role in transla-tion [87]. mRNA and protein synthesis machinery are sensi-tive to stress and may be potential targets to salt toxicity inplants. (ii) PDH45 may associate with DNA multi-subunitprotein complexes to alter gene expression. This hypothesiswas supported by the demonstration of the interaction ofPDH45 with topoisomerase I. This interaction was pro-posed to play an important role at the level of transcrip-tional regulation by the authors.

Recently, a novel DNA helicase gene PDH47 (peaDNA helicase 47) was isolated and shown to be inducedunder cold as well as salinity stress [88]. The enzyme con-tained a bi-directional DNA helicase activity (both 3�–5�and 5�–3�) and was involved in translation initiation ofthe proteins. This enzyme showed a dual localization innucleus as well as in the cytoplasm. Another reportproved that a DEAD box RNA helicase, LOS4, is essen-tial for mRNA export and is important for developmentand stress response in Arabidopsis [89].

Drought stress

Water stress may arise as a result of two conditions,either due to excess of water or water deWcit. Flooding is anexample of excess of water, which primarily results inreduced oxygen supply to the roots. Reduced O2 results inthe malfunctioning of critical root functions including lim-ited nutrient uptake and respiration. The more commonwater stress encountered is the water deWcit stress known asthe drought stress. Removal of water from the membranedisrupts the normal bilayer structure and results in themembrane becoming exceptionally porous when desic-cated. Stress within the lipid bilayer may also result in dis-placement of membrane proteins and this contributes toloss of membrane integrity, selectivity, disruption of cellu-lar compartmentalization and a loss of activity of enzymes,which are primarily membrane based. In addition to mem-brane damage, cytosolic and organelle protein may exhibitreduced activity or may even undergo complete denatur-ation when dehydrated. The high concentration of cellularelectrolytes due to the dehydration of protoplasm may alsocause disruption of cellular metabolism.

The components of drought and salt stress cross talkwith each other as both these stresses ultimately result indehydration of the cell and osmotic imbalance. Virtuallyevery aspect of plants physiology as well cellular metabo-lism is aVected by salt and drought stress. Drought and saltsignaling encompasses three important parameters [56].

(1) Reinstating osmotic as well as ionic equilibrium ofthe cell to maintain cellular homeostasis under thecondition of stress.

(2) Control as well as repair of stress damage by detoxiW-cation signaling.

(3) Signaling to coordinate cell division to meet therequirements of the plant under stress.

As a consequence of drought stress many changes occurin the cell and these include change in the expression levelof LEA/dehydrin-type genes, synthesis of molecular chaper-ones, which help in protecting the partner protein fromdegradation and proteinases that function to remove dena-tured and damaged proteins. This stress also leads to acti-vation of enzymes involved in the production and removalof ROS [59,90]. The over-expression of barley group 3 LEAgene HVA1 in leaves and roots of rice and wheat lead toimproved tolerance against osmotic stress as well asimproved recovery after drought and salinity stress [91].Dehydrins, also known as group 2 LEA proteins accumu-late in response to both dehydration as well as low temper-ature [28].

Other physiological eVects of drought on plants are thereduction in vegetative growth, in particular shoot growth.Reduced cyclin-dependent kinase activity results in slowercell division as well as inhibition of growth under waterdeWcit condition [92]. Leaf growth is generally more sensi-tive than the root growth. Reduced leaf expansion is


beneWcial to plants under water deWcit condition, as lessleaf area is exposed resulting in reduced transpiration. Inaccordance, many mature plants, for example cotton sub-jected to drought respond by accelerating senescence andabscission of the older leaves. This process is also known asleaf area adjustment. Regarding root, the relative rootgrowth may undergo enhancement, which facilitates thecapacity of the root system to extract more water fromdeeper soil layers.

Under drought stress, we have focused on variousaspects, which include response of stomata to drought con-dition, eVect of drought on photosynthetic machinery, roleof sugars and other osmolytes, and the role of MAPKinases in mediating osmotic stress tolerance. Moreover,the role of phospholipids signaling under an osmotic stresscondition and a generic pathway in response to salt,drought and cold stress is also described in this section.

Response of stomata to drought condition

Increase in temperature or a rapid drop in humidityoften results in acute water deWcit condition in plants.Moreover, dry air mass, which moves into the environment,can also add to rapid and acute water losses from plants.Such atmospheric changes result in a dramatic increase inthe vapor pressure gradient between leaf and the ambientair. This results in increased rate of transpiration. More-over, increase in vapor pressure gradient enhances waterloss from the soil.

The Wrst response of virtually all the plants to acutewater deWcit is the closure of their stomata to prevent thetranspirational water loss [93]. Closure of stomata mayresult from direct evaporation of water from the guard cellswith no metabolic involvement. This process of stomatalclosure is referred to as hydropassive closure. Stomatal clo-sure may also be metabolically dependent and involve pro-cesses that result in reversal of the ion Xuxes that causestomatal opening. This process of stomatal closure, whichrequires ions and metabolites, is known as hydroactive clo-sure. This process seems to be ABA regulated.

Plant growth and response to a stress condition islargely under the control of hormones. Hormones, in par-ticular ABA along with cytokinins and ethylene, havebeen implicated in the root–shoot signaling. This long dis-tance signaling may be mediated particularly via ABA aswell as ROS [94]. Recent studies have implicated that thetransport of ABA into root xylem is modulated by envi-ronmental factors such as xylem pH and the duration ofthe day see [95]. Under the water deWcit condition the pHof xylem sap increases therefore promoting the loading ofABA into the root xylem and its transport to the shoot[96]. Environmental conditions that increase the rate oftranspiration also result in an increase in the pH of leafsap, which can promote ABA accumulation and lead toreduction in stomatal conductance [95,97]. Increased cyto-kinin concentration in the xylem sap was shown to pro-mote stomatal opening directly as well as decrease the

sensitivity of stomata towards ABA see [95]. It has beenproposed by Sharp [98] that the concentration of varioushormones may govern their mode of action. For instance,as ethylene inhibits growth, an insuYcient amount ofABA accumulation in the shoot would result in ethylenemediated growth inhibition, whereas, higher accumula-tion of ABA in the root would prevent the ethylene medi-ated growth inhibition.

ABA promotes the eZux of K+ ions from the guardcells, which results in the loss of turgor pressure leading tostomata closure. Stomata closure does not always dependupon the perception of water deWcit signals arising fromleaves. In fact, stomata closure also responds directly to thesoil desiccation even before there is any signiWcant reduc-tion in leaf mesophyll turgor pressure. The fact that ABAcan act as a long distance communication signal betweenwater deWcit roots and leafs, inducing the closure of sto-mata is about two decades old [99].

AVect of drought on photosynthetic machinery

As stresses co-exist in nature with each other, a croptherefore may have to survive a stress episode of droughtaccompanied by high temperature. Plants respond quicklyto prevent the photosynthetic machinery from suVeringfrom irreversible damages. Stomatal closure in response toa water deWcit stress primarily results in decline in the rateof photosynthesis. Very severe drought conditions results inlimited photosynthesis due to decline in Rubisco activity[100]. The activity of photosynthetic electron chain is Wnelytuned to the availability of CO2 in the plant and photosys-tem II (PS II) often declines in parallel under drought con-ditions [101]. It has been shown that the decline in the rateof photosynthesis in drought stress is primarily due to CO2deWciency, as the photochemical eYciency could be broughtback to normal after a fast transition of leaves to an envi-ronment enriched in CO2 [102]. Decline in intracellular CO2levels results in the over-reduction of components withinthe electron transport chain and the electrons get trans-ferred to oxygen at photosystem I (PS I). This generatesROS including superoxide, hydrogen peroxide (H2O2) andhydroxyl radicals. These ROS need to be scavenged by theplant as they may lead to photo-oxidation. Redox signalsare like a forewarning for the plant, controlling the energybalance of the leaves. Some of the key electron carriers suchas plastoquinone (PQ), or the electron acceptors such asferredoxin/thioredoxin system as well as ROS are includedin the redox signaling molecules. Whereas a reduced PQpool activates the transcription of PS I reaction centre, theoxidized pool activates the transcription of PS II reactioncentre [103]. Plant detoxifying systems, which include ascor-bate and glutathione pools control the intracellular concen-tration of ROS. These ROS acts as second messengers inredox signal transduction and are implicated in hormonalmediated events [104]. H2O2 acts as a signal for the closureof leaf stomata, acclimation of leaf to high irradiation andthe induction of heat shock proteins [105]. In Arabidopsis


application of ABA to guard cells was shown to induce aburst of H2O2 that resulted in stomatal closure [106].

In a situation where water deWcit becomes too intenseor prolonged, plants can wilt, cells can undergo shrinkageand this may lead to mechanical constraint on cellularmembranes. The strain on membrane is one of the severeeVects of drought implicated on a plants’ physiology. Thisin particular impairs the functioning of ions and trans-porters as well as membrane associated enzymes. Chloro-plast membranes are in particular sensitive to oxidationstress damage caused by the generation of excessiveamount of ROS in these membranes. ROS can causeextensive peroxidation and de-esteriWcation of membranelipids, as well as lead to protein denaturation and muta-tion of nucleic acids [107]. Dehydration results in cellshrinkage and consequently a decline in cellular volume.This results in cellular content becoming viscous, there-fore increasing the probability of protein–protein interac-tion leading to their aggregation and denaturation [108].Increased concentration of solutes may also exceed toxiclevels, which may be deleterious for the functioning ofsome of the enzymes including the enzymes required forphotosynthetic machinery [108]. The transcript of some ofthe antioxidant genes such as glutathione reductase (GR)or the ascorbate peroxidase (APX) is higher during therecovery of water deWcit period and may play a role in theprotection of cellular machinery against photo-oxidationby ROS [109].

Role of sugars and other osmolytes in response to drought stress

Plants tend to cope with water deWcit stress by a processknown as osmotic adjustment. In this process, plantsdecrease their cellular osmotic potential by the accumula-tion of solutes. Certain metabolic processes are triggered inresponse to stress, which increase the net solute concentra-tion in the cell, thereby helping the movement of water intothe leaf resulting in increase in leaf turgor. Large numbersof compounds are synthesized, which play a key role inmaintaining the osmotic equilibrium and in the protectionof membranes as well as macromolecules. These com-pounds include proline, glutamate, glycine-betaine, carni-tine, mannitol, sorbitol, fructans, polyols, trehalose,sucrose, oligosaccharides and inorganic ions like K+. Thesecompounds help the cells to maintain their hydrated stateand therefore function to provide resistance againstdrought and cellular dehydration [108,110]. The hydroxylgroup of sugar alcohols substitutes the OH group of waterto maintain the hydrophilic interactions with the mem-brane lipids and proteins. Thus, these molecules help tomaintain the structural integrity of the membranes. Themost striking property of these stress-accumulated solutesis that they do not intervene with cells normal metabolicprocesses. The species, which synthesize large quantities ofsolutes, are known as osmotic adjusters for example Vignaunguiculata.

In response to a stress, the carbohydrate status of a leafgets altered and this might serve as a metabolic signal inresponse to stress [111,112]. Whereas the starch synthesis isnormally under strong inhibition even under moderatewater deWcit condition [113], the concentration of solublesugars in general increases or at least remains constantunder a stress condition [114]. Recent studies report theaccumulation of simple sugars such as glucose and fructosefollowing an increase in the invertase activity in the leavesof the drought challenged plants [114,115].

There was a direct correlation between the activities ofacid vacuolar invertase with the concentration of ABA inthe xylem sap [115]. ABA has been implicated in enhancingthe activity and expression of vacuolar invertase [115].There may also be a direct control of ABA biosynthesis byglucose as the transcript of several genes responsible forABA synthesis was increased by glucose in Arabidopsisseedlings [116]. A signaling pathway, which is initiated bydiVerent elicitors such as light, water and CO2 may con-verge down stream and be integrated as sugar signals[117,118]. Whereas, a decline in the sugar level triggers anincrease in the plants photosynthetic activity due to a de-repression of sugar control on transcription, the accumula-tion of sugar due to its low utilization have opposite eVecton photosynthetic activity [117].

There may exist cross talk between the sugars and planthormones such as ABA and ethylene. Glucose and ABAsignaling act in coordination regulating plants growth anddevelopment. Whereas high concentration of ABA andsugars act to inhibit growth in a severe drought stress, lowconcentration can promote growth. The inhibitory inXu-ence of glucose on growth could be overcome by ethylene[119]. These interactions appear to be dependent on theconcentration as well as tissue speciWc localization of thesehormones.

Osmolytes in low accumulation function in protectingmacromolecules either by stabilizing the tertiary structureof protein or by scavenging ROS produced in response todrought [120]. However, higher accumulation of osmolytesin transgenic plants can cause impaired growth in theabsence of any stress probably due to plants adaptationstrategy to conserve water in acute stress [121,122]. There-fore, controlled synthesis of osmolytes is the main concernin designing transgenic strategies for crop improvement.

Oligosaccharides such as raYnose and galactinol areamong the sugars synthesized in response to drought. Thesecompounds seem to function as osmoprotectants ratherthan providing osmotic adjustment [123]. Mannitol is oneof the most widely distributed sugar alcohol in nature andfunctions to scavenge the ROS, hydroxyl radicals and italso stabilizes the macro molecular structure of enzymes[124,125]. These osmolytes form hydrogen bonds with mac-romolecules under water deWcit condition and prevent theformation of intramolecular hydrogen bonds, which couldirreversibly damage the 3-dimensional structure of protein.Trehalose is a non-reducing disaccharide of glucose andhas been shown to exert its positive inXuence during


drought by stabilizing membranes and macromolecules.Trehalose over-expression helps in the maintenance of anelevated capacity for photosynthesis primarily due toincreased protection of PS II against photooxidation [126].

Some of the compatible solutes such as betaines, ectoineand proline accumulate in plants in response to variousenvironmental stresses [127,128]. Proline is one of theamino acids, which appear most commonly in response tostress. Plants synthesize proline from glutamine in theirleaves. Some of the crop plants for instance wheat ismarked by low level of these compounds and correspond-ingly the accumulation and mobilization of proline wasfound to increase tolerance towards water deWcit stress[129]. The over-expression of P5CS (pyrroline-5-carboxyl-ate synthase) gene from Vigna aconitifolia in tobacco, leadto increased levels of proline and consequently improvedgrowth under drought stress [130].

The maintenance of membrane Xuidity is an importantparameter against stress injury. Rehydration, after a longperiod of dehydration can also cause disruption of mem-brane integrity and leakage of solutes. During rehydration,water replaces sugar at the membrane surface leading to atransient membrane leakage [108]. Heat shock proteins(HSPs) are synthesized in response to cold as well as dehy-dration. These HSPs act as molecular chaperones and pro-tect the associated protein both during dehydration as wellas rehydration process. These HSPs help the protein tomaintain its tertiary structure and minimize the aggrega-tion and degradation of proteins [131]. The small HS(smHS) for instance the over-expression of AtHSP17.6Aclass from Arabidopsis could increase salt and droughtstress due to its chaperone-like activity [132].

Role of MAP kinases in osmotic stress

In plants several MAPKs (mitogen activated proteinkinase) are activated in response to hyperosmotic stress. Inalfalfa, a 46 kDa MAP kinase named SIMK (salt stressinducible MAPK) became activated in response to moder-ate hyperosmotic stress [133]. In tobacco cells, a SIMK-likeMAP kinase named SIPK (salicylic acid-induced proteinkinase) was activated by hyperosmotic stress [134]. Tran-script level for a number of protein kinases including a two-component histidine kinase MAPKKK, MAPKK andMAPK increases in response to osmotic stress [134]. Thisultimately results in the accumulation of osmolytes thathelps reestablish the osmotic balance, protection fromstress damage or repair mechanisms by induction of LEA/dehydrin-type stress genes.

Osmotic stress activates phospholipids signaling

Membrane phospholipids constitute a dynamic systemthat generates a multitude of signaling molecules like inositol1,4,5-triphosphate (IP3), diacylglycerol (DAG), phosphatidicacid (PA), etc. [59]. Phospholipase C (PLC) catalyzes thehydrolysis of phosphatidylinositol 4,5-bisphosphate (PIP2)

into IP3 and DAG, which acts as second messengers. IP3releases Ca2+ from internal stores. Several studies haveshown that in various plants systems IP3 levels rapidlyincrease in response to hyperosmotic stress [135–137]. IP3 lev-els also increased upon treatment with exogenous ABA inVicia faba guard cell protoplast [138] and in Arabidopsisseedlings [139]. An Arabidopsis PLC gene, AtPLC, is alsoinduced by salt and drought stress [140]. In guard cells, IP3induced Ca2+ increase in the cytoplasm lead to stomatal clo-sure and thus retention of water in the cells [141]. Microinjec-tion as well as pharmacological experiments suggested thatincrease in the cytoplasmic Ca2+ could lead to the expressionof osmotic stress responsive genes [142].

Osmotic stress activates Phospholipase D (PLD) activityin the suspension cells of Chlamydomonas, tomato, andalfalfa [143]. PLD cleaves membrane phospholipids to pro-duce PA and free head groups. PLD was rapidly activatedin response to drought stress in two plant species, i.e., Crat-erostigma plantagineum and Arabidopsis [144,145]. Whendrought stress-induced PLD activity was comparedbetween drought-resistant and sensitive cultivars of cow-pea, it was found that activity was higher in the drought-sensitive cultivars [146]. Consistent with this observation,blocking PLD activity resulted in reduced stress injury andimproved freezing tolerance. This suggests that PLD acti-vation results in lipolitic membrane disintegration duringstress injuries. Interestingly, the PLD product, PA hasemerged as a molecule to mitigate the eVect of stress injury.The application of PA mimics the eVect of ABA in inducingthe closure of stomata [147].

A generic pathway in response to salt, drought and coldstress is described in Fig. 3. Salt and drought exert their inXu-ence on a cell by disrupting the ionic and osmotic equilibriumresulting in a stress condition. Thus excess of Na+ ions andosmotic changes in the form of turgor pressure are the initialtriggers of this pathway. This leads to a cascade of events,which can be grouped under ionic and osmotic signalingpathway, the out come of which is ionic and osmotic homeo-stasis, leading to stress tolerance. These stresses are marked bysymptoms of stress injury including chlorosis and necrosisand may also exert its negative inXuence on cell divisionresulting in growth retardation of plant. Reduction in shootgrowth, especially, leaves is beneWcial for plant as it reducesthe surface area exposed for transpiration hence minimizingwater loss. Plants may also sacriWce or shed their older leaves,which is another adaptation in response to drought. Stressinjury may occur through denaturation of cellular proteins/enzymes or through the production of ROS, Na+ toxicity anddisruption of membrane integrity. In response to a stressinjury plants trigger a detoxiWcation process, which mayinclude change in the expression of LEA/dehydrin type genesynthesis of molecular chaperones, proteinases, enzymes forscavenging ROS and other detoxiWcation proteins. This pro-cess functions in the control and repair of stress induced dam-age and results in stress tolerance. Cold stress mainly results indisruption of membrane integrity leading to severe cellulardehydration and osmotic imbalance. Cold acclimation results


in the triggering of various genes, which result in restructuringof the cellular membranes by change in the lipid compositionand generation of osmolytes, which prevent cellular dehydra-tion therefore leading to stress tolerance.

ABA and abiotic stress signaling

ABA is an important phytohormone and plays a criticalrole in response to various stress signals. The application ofABA to plant mimics the eVect of a stress condition. Asmany abiotic stresses ultimately results in desiccation of thecell and osmotic imbalance, there is an overlap in theexpression pattern of stress genes after cold, drought, highsalt or ABA application. This suggests that various stresssignals and ABA share common elements in their signalingpathways and these common elements cross talk with eachother, to maintain cellular homeostasis [34,148–150]. Func-tions of ABA include:

Fig. 3. A generic pathway under salt, drought and cold stress. Salt anddrought disrupt the ionic and osmotic equilibrium of the cell resulting in astress condition. This triggers the process, which functions to reinstateionic and osmotic homeostasis leading to stress tolerance. Stress imposesinjury on cellular physiology and result in metabolic dysfunction. Thisinjury imposes a negative inXuence on cell division and growth of a plant.This is an indirect advantage to the plant as reduction of leaf expansionreduces the surface area of leaves exposed for transpiration and therebyreducing water loss. Stress injury and ROS generated in response to stressalso triggers a detoxiWcation signaling by activating genes responsible fordamage control and repair mechanism therefore leading to stress toler-ance. Cold stress mainly exerts its malicious eVect by disruption of mem-brane integrity and solute leakage. Moreover, other physiological factorssuch as rate of photosynthesis, protein assembly and general metabolicprocesses are severely hampered. Cold acclimation results in the restruc-turing of cellular membranes and synthesis of various osmolytes, whichfunction towards reinstating the normal cellular metabolism and stresstolerance.

Ioni

cst

ress

Osmoti

c stres

s

Ionic and osmotic homeostasis via SOS

pathway or related pathways

STRESS TOLERANCE

Activation ofStress genes

Stressinduced injury

Regulation of celldivision andexpansion

Growth inhibition

Detoxificationsignaling

Damage controland repair

Disruption of membraneintegrity, dehydration,solute leakage and

metabolic dysfunction

Activation ofStress genes

Restructuring of cellmembrane and

synthesis of osmolytes

SALT DROUGHT COLD
(1) ABA causes seed dormancy and delays its germina-
tion.(2) ABA promotes stomatal closure.

ABA levels are induced in response to various stress sig-nals. ABA actually helps the seeds to surpass the stress con-ditions and germinate only when the conditions areconducive for seed germination and growth. ABA also pre-vents the precocious germination of premature embryos.Stomatal closure under drought conditions prevents theintracellular water loss and thus ABA is aptly called as astress hormone.

The main function of ABA seems to be the regulation ofplant water balance and osmotic stress tolerance. SeveralABA deWcient mutants namely aba1, aba2 and aba3 havebeen reported for Arabidopsis [151]. ABA deWcient mutantsfor tobacco, tomato and maize have also been reported[152]. Without any stress treatment the growth of thesemutants is comparable to wild type plants. Under droughtstress, ABA deWcient mutants readily wilt and die if thestress persists. Under salt stress also ABA deWcient mutantsshow poor growth [139]. In addition, ABA is required forfreezing tolerance, which also involves the induction ofgenes in response to dehydration stress [139,153].

Processes that trigger activation of ABA synthesis andinhibition of its degradation result in ABA accumulation.Several ABA biosynthesis genes have been cloned whichincludes zeathanxin epoxidase (known as ABA1 in Arabid-opsis), [154], 9-cis-epoxycarotenoid dioxygenase (NCED)[155], ABA aldehyde oxidase and ABA3 also known asLOS5 [139].

Studies suggest that osmotic stress imposed by high saltor drought is transmitted through at least two pathways;one is ABA-dependent and the other ABA independent.Cold exerts its eVects on gene expression largely through anABA-independent pathway [150]. ABA induced expressionoften relies on the presence of cis acting element calledABRE [34,149,150,156]. Genetic analysis indicates thatthere is no clear line of demarcation between ABA-depen-dent and ABA-independent pathways and the componentsinvolved may often cross talk or even converge in the sig-naling pathway [157,158]. Calcium, which serves as a sec-ond messenger for various stresses, represents a strongcandidate, which can mediate such cross talk. Several stud-ies have demonstrated that ABA, drought, cold and highsalt result in rapid increase in calcium levels in plant cells[141,159,160]. The signaling pathway results in the activa-tion of various genes, which play signiWcant role towardsthe maintenance of cellular homeostasis.

Role of transcription factors in the activation of stress responsive genes

The promoters of stress responsive genes have typicalcis-regulatory elements like DRE/CRT, ABRE, MYCRS/MYBRS and are regulated by various upstream transcrip-tional factors (Fig. 4). These transcription factors fall in the


category of early genes and are induced within minutes ofstress. The transcriptional activation of some of the genesincluding RD29A has been well worked out. The promoterof this gene family contains both ABRE as well as DRE/CRT elements [36]. Transcription factors, which can bindto these elements were isolated and were found to belong toAP2/EREBP family and were designated as CBF1/DREB1B, CBF2/DREB1C, and CBF3/DREB1A[36,161,162]. These transcription factors (CBF1, 2 and 3)are cold responsive and in turn bind CRT/DRE elementsand activate the transcription of various stress responsivegenes. A novel transcription factor responsive to cold aswell as ABA was isolated from soybean and termed asSCOF-1 (soybean zinc Wnger protein). This transcriptionfactor, however, was not responsive to drought or salinitystress [163]. SCOF1 was a zinc Wnger nuclear localized pro-tein but failed to bind directly to either CRT/DRE orABRE elements. Yeast 2-hybrid study revealed that SCOF-1 interacted strongly with SGBF-1 (Soybean G-box bind-ing bZip transcription factor) and in vitro DNA bindingactivity of SGBF-1 to ABRE elements was greatlyimproved by the presence of SCOF-1. This study supportedthat protein–protein interaction is essential for the activa-tion of ABRE-mediated cold responsive genes [163]. Tran-scription factors like DREB2A and DREB2B getsactivated in response to dehydration and confer toleranceby induction of genes involved in maintaining the osmoticequilibrium of the cell [37]. Several basic leucine zipper(bZip) transcription factors (namely ABF/AREB) havebeen isolated which can speciWcally bind to ABRE elementand activate the expression of stress genes [156,164]. These

AREB genes (AREB1 and AREB2) are ABA responsiveand need ABA for their full activation. These transcriptionfactors exhibited reduced activity in the ABA-deWcientmutant aba2 as well as in ABA insensitive mutant aba1-1.Some of the stress responsive genes for example RD22 lackthe typical CRT/DRE elements in their promoter indicat-ing their regulation by other mechanisms. Transcriptionfactor RD22BP1 (a MYC transcription factor) andAtMYB2 (a MYB transcription factor) could bindMYCRS (MYC recognition sequence) and MYBRS (MYBrecognition sequence) elements, respectively, and couldcooperatively activate the expression of RD22 gene [165].As cold, salinity and drought stress ultimately impair theosmotic equilibrium of the cell it is likely that these tran-scription factors as well as the major stress genes may crosstalk with each other for their maximal response and help inreinstating the normal physiology of the plant.

Role of CBL and CIPK genes in response to abiotic stresses in plants

Signaling pathway is complex as it involves the coordi-nated action of various genes in a single pathway or diversepathways. Calcium is a prime candidate, which functions asa central node in mediating the coordination and synchro-nization of diverse stimuli into speciWc cellular responses.Thus, the proteins, which sense cytoplasmic Ca2+ perturba-tions and relay this information to downstream molecules,serve as an important component of signaling. In plant cellmany calcium sensors have been recognized which includecalmodulin (CaM) and calmodulin-related proteins

Fig. 4. Involvement of diVerent transcription factors in response to cold, salinity and drought pathways in the induction of stress genes. Various transcrip-tion factors such as CBF1, 2 and 3 are triggered in response to cold stress and mediate their inXuence on stress genes mainly via ABA-independent path-way. Transcription factors such as SCOF-1 and SGBF-1 seem to follow ABA dependent pathway for expression of genes involved in imparting coldtolerance. Osmotic stress signaling generated via salinity and drought stress seems to be mediated by transcription factors such as DREB2A, DREB2B,bZip and MYC and MYB transcription activators, which interacts with CRT/DRE, ABRE or MYCRE/MYBRE elements in the promoter of stressgenes. Salinity mainly works through SOS pathway reinstating cellular ionic equilibrium.

ColdCold DroughtDrought SaltSalt

CBF1,2,3/DREB1B,1C,1A

ABRE

mRNA

DRE/CRT MYCRS/

MYBRS

TATA

ICEinactive

ICE activeMYB

MYC G BOX

SCOFSCOF

SGBFSGBF

DREB2A, 2BDREB2A, 2B bZIP

ABA

MYC/MYB

SOS pathwaySOS3/SOS2

~ PO4

SOS1

activates

HOS1

AB

A

ABA

Ca2+

ABA


[166,167], Ca2+-dependent protein kinases (CDPKs)[159,168,169] and the relatively recently discovered sensorCBL (calcineurin B-like) protein [56]. CBLs are character-ized by 4 helix-loop-helix calcium binding domains termedas EF hands. Currently, 10 isoforms of CBL have been dis-covered in Arabidopsis and named as CBL due to their sig-niWcant sequence similarity to animal calcineurin B. Despitethis sequence similarity, Arabidopsis lacks calcineurin in itsdata bank [170]. Various isoforms of CBL are up-regulatedin stress condition (refer Table 2). CBLs speciWcally interactwith a class of kinases known as CBL-interacting proteinkinase (CIPKs) to transduce the signal via phosphorylationof downstream signaling components.

Presently the direction of research is more towards isola-tion of master switches, which can control these stressgenes. As cytosolic calcium upregulation is more or less auniversal phenomenon associated with stress signaling, thusthe calcium sensors, which decode these Ca2+ signaturesand relay the information down stream, may act as masterswitches in controlling various stress genes. Moreover,mutations in these calcium sensors like AtCBL1 and theirinteracting protein kinases have been shown to cause aber-rations in the expression of some of the major stressresponsive genes like RD29A, KINI, KIN2 and RD22 indi-cating their immense signiWcance in stress signaling [171–173].

Conclusion and future prospects

Abiotic stress signaling is an important area with respectto increase in plant productivity. Therefore, the basicunderstanding of the mechanisms underlying the function-ing of stress genes is important for the development oftransgenic plants. Each stress is a multigenic trait andtherefore their manipulation may result in alteration of alarge number of genes as well as their products. A deeperunderstanding of the transcription factors regulating thesegenes, the products of the major stress responsive genes andcross talk between diVerent signaling components shouldremain an area of intense research activity in future.

The knowledge generated through these studies shouldbe utilized in making transgenic plants that would be ableto tolerate stress condition without showing any growthand yield penalty. In the improvement of crops it is veryimportant to perturb the natural machinery as minimum aspossible and activate the stress genes at a correct time.Therefore, it is desirable that appropriate stress induciblepromoters should drive the stress genes as well as transcrip-tion factors, which will minimize their expression under anon-stressed condition thereby reducing yield penalty. Theproduct of these genes should also be targeted to thedesired tissue as well as cellular location to control the tim-ing as well as intensity of expression.

Attempts should be made to design suitable vectors forstacking relevant genes of one pathway or complementarypathways to develop durable tolerance. These genes shouldpreferably be driven by a stress inducible promoter to havemaximal beneWcial eVects. Additionally, due importanceshould be laid on the physiological parameters such as therelative content of diVerent ions present in the soil as well asthe water status of the crop in designing transgenic plantsfor the future.

Acknowledgments

We thank Dr. Renu Tuteja for critically reading themanuscript and the Department of Biotechnology, Govern-ment of India grant for partial support and Council of Sci-entiWc and Industrial Research, New Delhi for fellowship toS.M. We apologize if some references could not been citeddue to space constraint.

References

[1] E.A. Bray, J. Bailey-Serres, E. Weretilnyk, Responses to abioticstresses, in: W. Gruissem, B. Buchannan, R. Jones (Eds.), Biochemis-try and Molecular Biology of Plants, American Society of Plant Biol-ogists, Rockville, MD, 2000, pp. 158–1249.

[2] H.G. Jones, M.B. Jones, Introduction: some terminology and commonmechanisms, in: H.G. Jones, T.J. Flowers, M.B. Jones (Eds.), PlantsUnder Stress, Cambridge university Press, Cambridge, 1989, pp. 1–10.

Table 2Expression of CBL and CIPK genes in response to abiotic stresses

CBL and CIPK genes Responsive to stress/hormones/sugars Species Reference

CBL1 Salt, drought and cold. Negative regulator of cold signaling Arabidopsis [172,174]CBL2 Light and G.A Arabidopsis & Oryza sativa [175,176]CBL3 Late induced under cold, salt, S.A and proposed to be involved

in maintenance of stress responsePisum sativum (Unpublished data)

CBL4/SOS3 Salt stress Arabidopsis [56,61,70,177,178]CBL5 ABA Arabidopsis [64]CBL9 Negative regulator of ABA signaling and promotes seed germination. Arabidopsis [173]CIPK3 Expression induced strongly in response to cold followed by

drought salinity, wounding and ABA signaling. May act as a cross talk node between ABA-dependent and independent signaling

Arabidopsis [171]

CIPK8/PKS11 Transgenic plants overexpressing CIPK8 were resistant to high level of glucose indicating its role in sugar signaling

Arabidopsis [179]

CIPK14 mRNA level was up-regulated in response to sugars such as sucrose, glucose and fructose

Arabidopsis [180]


[3] D.V. Lynch, Chilling injury in plants: the relevance of membrane lip-ids, in: F. Katterman (Ed.), Environmental Injury to Plants, Academicpress, New York, 1990, pp. 17–34.

[4] .L. Guy, cold acclimation and freezing stress tolerance: role of proteinmetabolism, Annu. Rev. Plant. Physiol. 41 (1990) 187–223.

[5] W.G. Hopkins, The physiology of plants under stress, in: Introductionto Plant Physiology, second ed., Wiley, New York, 1999, pp. 451–475.

[6] Q.W Jiang, O. Kiyoharu, I. Ryozo, Two novel mitogen-activated pro-tein signaling components, OsMEK1 and OsMAP1, are involved in amoderate low-temperature signaling pathway in Rice1, Plant Physiol.129 (2002) 1880–1891.

[7] P.L. Steponkus, Role of the plasma membrane in freezing injury andcold acclimation, Annu. Rev. Plant. Physiol. 35 (1984) 543–584.

[8] P.L. Steponkus, M. Uemura, M.S. Webb, A contrast of the cryostabil-ity of the plasma membrane of winter rye and spring oat-two speciesthat widely diVer in their freezing tolerance and plasma membranelipid composition, in: P.L. Steponkus (Ed.), Advances in Low-Tem-perature Biology, vol. 2, JAI Press, London, 1993, pp. 211–312.

[9] B.D. McKersie, S.R. Bowley, Active Oxygen and Freezing Tolerancein Transgenic Plants, Plant Cold Hardiness, Molecular Biology, Bio-chemistry and Physiology, Plenum, New York, 1997, pp. 203–214.

[10] C.R. Olien, M.N. Smith, Ice adhesions in relation to freeze stress,Plant Physiol. 60 (1997) 499–503.

[11] M. Uemura, P.L. Steponkus, EVect of Cold Acclimation on Mem-brane Lipid Composition and Freeze Induced Membrane Destabili-zation, Plant Cold Hardiness. Molecular Biology, Biochemistry andPhysiology, Plenum, New York, 1997, pp. 171–79.

[12] A. Kacperska, Metabolic consequences of low temperature stress inchilling-insensitive plants, in: P.H. Li (Ed.), Low TemperatureStress Physiology in Crops, CRC Press, Boca Raton, FL, 1989, pp.27–40.

[13] J.K. Raison, G.R. Orr, Phase transition in liposomes formed frompolar lipids of mitochondria from chilling sensitive plants, PlantPhysiol. 81 (1986) 807–811.

[14] J.P. Williams, M.U. Kahn, K. Mitchell, G. Johnson, The eVect of tem-perature on the level and biosynthesis of unsaturated fatty acids indiaglycerols of Brassica napus leaves, Plant Physiol. 87 (1988) 904–910.

[15] A.R. Cossins, Homeoviscous adaptation of biological membranes andits functional signiWcance, in: A.R. Cossins (Ed.), temperature Adap-tation of Biological membranes, Portland Press, London, 1994, pp.63–76.

[16] D.V. Lynch, G.A. Thompson Jr., Low temperature-induced altera-tions in the chloroplast and microsomal membrane of Dunaliellasalina, Plant Physiol. 69 (1982) 1369–1375.

[17] H. Knight, A.J. Trewavas, M.R. Knight, Cold calcium signaling inArabidopsis involves two cellular pools and a change in calcium signa-ture after acclimation, Plant Cell 8 (1996) 489–503.

[18] P.G. Jones, M. Inouye, The cold shock response—a hot topic, Mol.Microbiol. 11 (1994) 811–818.

[19] I. Nishida, N. Murata, Chilling sensitivity in plants and Cyanobacte-ria: the crucial contribution of membrane lipid, Annu. Rev. PlantPhysiol. Plant Mol. Biol. 47 (1996) 541–568.

[20] C. Guy, D. Haskell, Q.B. Li, Association of proteins with the stress 70molecular chaperones at low temperature evidence for the existenceof cold labile proteins in spinach, Cryobiology 36 (1998) 301–314.

[21] C.L. Guy, Q.B. Li, The organization and evolution of the spinach stress70 molecular chaperone gene family, Plant Cell 10 (1998) 539–556.

[22] S. Gibson, V. Arondel, K. Iba, C. Somerville, Cloning of a tempera-ture-regulated gene encoding a chloroplast Omega-3 desaturase fromArabidopsis thaliana, Plant Physiol. 106 (1994) 1615–1621.

[23] Anderson, Q.B. Li, D.W. Haskell, C.L. Guy, Structural organizationof the spinach endoplasmic reticulum-luninal 70-kilodalton heat-shock cognate gene and expression of 70-kilodalton heat shock genesduring cold acclimation, Plant Physiol. 104 (1994) 1359–1370.

[24] P. Krishna, M. Sacco, J.F. Cherutti, S. Hill, Cold-induced accumula-tion of hsp 90 transcripts in Brasscia napus, Plant Physiol. 107 (1995)915–923.

[25] T. Mizoguchi, N. Hayashida, K. Yamaguchi-Schinozaki, H. Kamada,K. Shinozaki, AtMPKs: a gene family of plant MAP Kinases in Ara-bidopsis thaliana, FEBS Lett. 336 (1993) 440–444.

[26] T. Mizoguchi, K. Irie, T. Hirayama, N. Hayashida, K. Yamaguchi-Shinozaki, et al., A gene encoding a mitogen activated protein kinasekinase kinase is induced simultaneously with genes for a mitogen-acti-vated protein kinase and an S6 ribosomal protein kinase by touch,cold and water stress in Arabidopsis thaliana, Proc. Natl. Acad. Sci.USA 93 (1996) 765–769.

[27] D.H. Polisensky, J. Braam, Cold shock regulation of the ArabidopsisTCH genes and the eVects of modulating intracellular calcium levels,Plant Physiol. 111 (1996) 1271–1279.

[28] TJ. Close, Dehydrins: a commonality in the response of plants todehydration and low temperature, Physiologia Plantarum 100 (1997)291–296.

[29] L. Dure III, A repeating 11-mer amino acid motif and plant desicca-tion, Plant J. 3 (1993) 363–369.

[30] J. Ingram, D. Bartels, The molecular basis of dehydration tolerance inplants, Annu. Rev. Plant Physiol. Plant Mol. Biol. 47 (1996) 377–403.

[31] N.N. Artus, M. Uemura, P.L. Steponkus, S.J. Gilmour, C.T. Lin, M.F.Thomashow, Constitutive expression of the cold regulated Arabidop-sis thaliana COR 15a gene aVects both chloroplast and protoplastfreezing tolerance, Proc. Natl. Acad. Sci. USA 93 (1996) 13404–13409.

[32] A.F. Monroy, Y. Castonguay, S. Laberge, F. Sarhan, L.P. Vezina, R.S.Dhindsa, A new cold-induced alfalfa gene is associated with enhancedhardening at sub zero temperature, Plant Physiol. 102 (1993) 873–879.

[33] M. Houde, R.S. Dhindsa, F. Sarhan, A molecular marker to select forfreezing tolerance in Gramineae, Mol. Gen. Genet. 234 (1992) 43–48.

[34] M.F. Thomashow, Plant cold acclimation: freezing tolerance genesand regulatory mechanism, Annu. Rev. Plant Physiol. Plant Mol. Biol.50 (1999) 571–599.

[35] K. Yamaguchi-Shinozaki, K. Shinozaki, A novel cis-acting element inan Arabidopsis gene is involved in responsiveness to drought, low-temperature, or high-salt stress, Plant Cell 6 (1994) 251–264.

[36] E.J. Stockinger, S.J. Gilmour, M.F. Thomashow, Arabidopsis thalianaCBF1 encodes an AP2 domain-containing transcription activatorthat binds to the C-repeat/DRE, a cis-acting DNA regulatory elementthat stimulates transcription in response to low temperature andwater deWcit, Proc. Natl. Acad. Sci. USA 94 (1997) 1035–1040.

[37] Q. Liu, M. Kasuga, Y. Sakuma, H. Abe, S. Miura, et al., Two tran-scription factors, DREBI and DREB2, with an EREBP/AP2 DNAbinding domain separate two cellular signal transduction pathways indrought and low temperature responsive gene expression respectively,in Arabidopsis, Plant Cell 10 (1998) 1391–1406.

[38] H. Lee, L. Xiong, Z. Gong, M. Ishitani, B. Stevenson, J.K. Zhu, TheArabidopsis HOS1 gene negatively regulates cold signal transduc-tion and encodes a RING Wnger protein that displays cold-regu-lated nucleo-cytoplasmic partitioning, Genes Dev. 15 (2001) 912–924.

[39] V. Chinnusamy, M. Ohta, S. Kanrar, B.H. Lee, X. Hong, M. Agarwal,J.K. Zhu, ICE1: a regulator of cold-induced transcriptome and freez-ing tolerance in Arabidopsis, Genes Dev. 17 (2003) 1043–1054.

[40] S. Kim, C.S. An, Y.N. Hong, K.W. Lee, Cold-inducible transcriptionfactor, CaCBF, is associated with a homeodomain Leucine Zipperprotein in hot pepper, Mol. Cells 18 (2004) 300–30836.

[41] F. Abbasi, H. Onodera, S. Toki, H. Tanaka, S. Komatsu, OsCDPK 13,a calcium-dependent protein kinase gene from rice, is induced by coldand gibberellin in rice leaf sheath, Plant Mol. Biol. 55 (2004) 541–552.

[42] Z. Xin, J. Browse, Eskimo1 mutants of Arabidopsis are constitutivelyfreezing tolerant, Proc. Natl. Acad. Sci. USA 95 (1998) 7799–7804.

[43] A.F. Monroy, R.S. Dhindsa, Low temperature signal transduction:induction of cold acclimation-speciWc genes of alfalfa by calcium at25 °C, Plant Cell 7 (1995) 321–331.

[44] W. Wang, B. Vinocur, A. Altman, Plant responses to drought, salinityand extreme temperatures: towards genetic engineering for stress tol-erance, Planta 218 (2003) 1–14.

[45] X. Niu, R.A. Bressan, P.M. Hasegawa, J.M. Pardo, Ion homeostasis inNaCl stress environments, Plant Physiol. 109 (1995) 735–742.


[46] T.J. Flowers, P.F. Troke, A.R. Yeo, The mechanisms of salt tolerancein halophytes, Annu. Rev. Plant Physiol. 28 (1977) 89–121.

[47] H. Greenway, R. Munns, Mehcnaisms of salt tolerance in non hal-phytes, Annu. Rev. Plant Physiol. 31 (1980) 149–190.

[48] A.R. Yeo, Molecular biology of salt tolerance in the context of whole-plant physiology, J. Exp. Bot. 49 (1998) 915–929.

[49] T.J. Flowers, A.R. Yeo, Solute Transport in plants, Glasgow, Scot-land, Blackie, vol. 176, 1992, pp. 45.

[50] A. Amtmann, D. Sanders, Mechanism of Na+ uptake by plant cells,Adv. Bot. Res. 29 (1999) 75–112.

[51] E. Blumwald, G.S. Aharaon, M.P. Apse, Sodium transport in plantcells, Biochem. Biophys. Acta (2000).

[52] Y. Braun, M. Hassidim, H.R. Lerner, L. Reinhold, Studies on H+-translocating ATPase in plants of varying resistance to salinity. I.Salinity during growth modulates the proton pump in the halophyteAtriplex nummularia, Plant physiol. 81 (1986) 1050–1056.

[53] A.A. Watad, M. Reuveni, R.A. Bressan, P.M. Hasegawa, Enhancednet K+ uptake capacity of NaCl-adapted cells, Plant Phyisol. 95(1991) 1265–1269.

[54] G.R. Cramer, J. Lynch, A. Lauchli, E. Epstein, InXux of Na+, K+, andCa2+ into roots of salt-stressed cotton seedlings, Plant Physiol. 83(1987) 510–516.

[55] A. Lauchli, S. Schubert, The role of calcium in the regulation of mem-brane and cellular growth processes under salt stress, NATO ASI Ser.G. 19 (1989) 131–137.

[56] J. Liu, J.K. Zhu, A calcium sensor homolog required for plant salt tol-erance, Science 280 (1998) 1943–1945.

[57] H. Knight, A.J. Trewavas, M.R. Knight, Calcium signalling in Arabid-opsis thaliana responding to drought and salinity, Plant J. 12 (1997)1067–1078.

[58] J.K. Zhu, J. Liu, L. Xiong, Genetic analysis of salt tolerance in Arabid-opsis. Evidence for a critical role of potassium nutrition, Plant Cell 10(1998) 1181–1191.

[59] J.K. Zhu, Salt and drought stress signal transduction in plants, Annu.Rev. Plant Physiol. Plant Mol. Biol. 53 (2002) 247–273.

[60] M. Ishitani, J. Liu, U. Halfter, C.S. Kim, W. Shi, J.K. Zhu, SOS3 func-tion in plant salt tolerance requires N-myristoylation and calciumbinding, Plant Cell 12 (2000) 1667–1677.

[61] J. Liu, M. Ishitani, U. Halfter, C.S. Kim, J.K. Zhu, The Arabidopsisthaliana SOS2 gene encodes a protein kinase that is required for salttolerance, Proc. Natl. Acad. Sci. USA 97 (2000) 3730–3734.

[62] V. Albrecht, O. Ritz, S. Linder, K. Harter, J. Kudla, The NAF domaindeWnes a novel protein–protein interaction module conserved in Ca2+-regulated kinases, EMBO J. 20 (2001) 1051–1063.

[63] U. Halfter, M. Ishitani, J.K. Zhu, The Arabidopsis SOS2 proteinkinase physically interacts with and is activated by the calcium-bind-ing protein SOS3, Proc. Natl. Acad. Sci. USA 97 (2000) 3735–3740.

[64] Y. Guo, L. Xiong, C.P. Song, D. Gong, U. Halfter, J.K. Zhu, A cal-cium sensor and its interacting protein kinase are global regulators ofabscisic acid signaling in Arabidopsis, Dev. Cell 3 (2002) 233–244.

[65] N.H. Cheng, J.K. Pittman, J.K. Zhu, K.D. Hirschi, The protein kinaseSOS2 activates the Arabidopsis H+/Ca+ antiporter CAX1 to integrateCa transport and salt tolerance, J. Biol. Chem. 279 (2004) 2922–2926.

[66] H. Shi, M. Ishitani, C. Kim, J.-K. Zhu, The Arabidopsis thaliana salttolerance gene SOS1 encodes a putative Na+/H+ antiporter, Proc.Natl. Acad. Sci. USA 97 (2000) 6896–6901.

[67] F.J. Quintero, M. Ohta, H. Shi, J.K. Zhu, J.M. Pardo, Reconstitutionin yeast of the Arabidopsis SOS signaling pathway for Na+ homeosta-sis, Proc. Natl. Acad. Sci. USA 99 (2002) 9061–9066.

[68] N. Uozumi, E.J. Kim, F. Rubio, T. Yamaguchi, S. Muto, et al., TheArabidopsis HKT1 gene homolog mediates inward Na currents inXenopus laevis oocytes and Na uptake in Saccharomyces cerevisiae,Plant Physiol. 122 (2000) 1249–1260.

[69] A. Rus, S. Yokoi, A. Sharkhuu, M. Reddy, B.H. Lee, et al., AtHKT1 isa salt tolerance determinant that controls Na entry into plant roots,Proc. Natl. Acad. Sci. USA 98 (2001) 14150–14155.

[70] Q.S. Qiu, Y. Guo, M. Dietrich, K.S. Schumaker, J.K. Zhu, Regulationof SOS1, a plasma membrane Na/H exchanger in Arabidopsis thali-

ana, by SOS2 and SOS3, Proc. Natl. Acad. Sci. USA 99 (2002) 8436–8441.

[71] M.P. Apse, G.S. Aharon, W.A. Snedden, E. Blumwald, Salt toleranceconferred by over expression of a vacuolar Na+/H+ antiport in Ara-bidopsis, Science 285 (1999) 1256–1258.

[72] R.A. Gaxiola, R. Rao, A. Sherman, P. GrisaW, S. Alper, G.R. Fink,The Arabidopsis thaliana proton transporters AtNhx1 and Avp1, canfunction in cation detoxiWcation in yeast, Proc. Natl. Acad. Sci. USA96 (1999) 1480–1485.

[73] C.W. Ford, Accumulation of low molecular weight solutes in waterstress tropical legumes, Phytochemistry 22 (1984) 875–884.

[74] R.A. Breesan, P.M. Hasegawa, J.M. Pardo, Plants use calcium toresolve salt stress, Trends Plant Sci. 3 (1998) 411–412.

[75] A.J. Delauney, D.P.S. Verma, Proline biosynthesis and osmoregula-tion in plants, Plant J. 4 (1993) 215–223.

[76] P. Louis, E.A. Galinski, Characterization of genes for the biosynthesisof the compatible solute ectoine from Marinococcus halophilus andosmoregulated expression in E. coli, Microbiology 143 (1997) 1141–1149.

[77] K.F. McCue, A.D. Hanson, Drought and salt tolerance: towardsunderstanding and application, Biotechnology 8 (1990) 358–362.

[78] D. Rhodes, A.D. Hanson, Quaternary ammonium and tertiary sul-phonium compounds in higher plants, Annu. Rev. Plant Physiol. Mol.Biol. 44 (1993) 357–384.

[79] A.D. Hanson, B. Rathinasabapathi, J. Rivoal, M. Burnet, M.O. Dil-lon, D.A. Gage, Osmoprotective compound in the Plumbaginaceae: anatural experiment in metabolic engineering of stress tolerance, Proc.Natl. Acad. Sci. USA 91 (1994) 306–310.

[80] R. Waditee, N.H. Bhuiyan, V. Rai, K. Aoki, Y. Tanaka, T. Hibino, S.Suzuki, J. Takano, A.T. Jagendorf, T. Takabe, T. Takabe, Genes fordirect methylation of glycine provide high levels of glycine betaineand abiotic-stress tolerance in Synechococcus and Arabidopsis, Proc.Natl. Acad. Sci. USA 102 (2005) 1318–1323.

[81] S.W. Matson, D. Bean, J.W. George, DNA helicases; enzymes withessential roles in all aspects of DNA metabolism, BioEssays 16 (1994)13–21.

[82] N. Tuteja, R. Tuteja, DNA helicases: the long unwinding road, Nat.Genet. 13 (1996) 11–12.

[83] T.M. Lohman, K.P. Bjornson, Mechanism of helicase-catalyzed DNAunwinding, Ann. Rev. Biochem. 65 (1996) 169–214.

[84] N. Tuteja, R. Tuteja, Prokaryotic and eukaryotic DNA helicases.Essential molecular motor proteins for cellular machinery, Eur. J.Biochem. 271 (2004) 1835–1848.

[85] N. Tuteja, R. Tuteja, Unraveling DNA helicases. Motif, structure,mechanism and function, Eur. J. Biochem. 271 (2004) 1849–1863.

[86] N. Sanan-Mishra, X.H. Phan, S.K. Sopory, N. Tuteja, Pea DNA heli-case 45 overexpression in tobacco confers high salinity tolerance with-out aVecting yield, Proc. Natl. Acad. Sci. USA 102 (2005) 509–514.

[87] X.H. Pham, M.K. Reddy, N.Z. Ehtesham, B. Matta, N. Tuteja, A DNAhelicase from Pisum sativum is homologous to translation initiation fac-tor and stimulates topoisomerase I activity, Plant J. 24 (2000) 219–229.

[88]A.A. Vashisht, A. Pradhan, R. Tuteja, N. Tuteja, Cold and salinitystress-induced bipolar pea DNA helicase 47 is involved in proteinsynthesis and stimulated by phosphorylation with protein kinase C,Plant J. 44 (2005) 76–87.

[89] Z. Gong, C.H. Dong, H. Lee, J. Zhu, L. Xiong, D. Gong, B. Stevenson,J.K. Zhu, A DEAD BOX RNA helicase is essential for mRNA exportand important for development and stress responses in Arabidopsis,Plant Cell 17 (2005) 256–267.

[90] J.C. Cushman, H.J. Bohnert, Genomic approaches to plant stress tol-erance, Curr. Opin. Plant Biol. 3 (2000) 117–124.

[91] E. Sivamani, A. Bahieldin, J.M. Wraith, T. Al-Niemi, W.E. Dyer,T.H.D. Ho, R. Qu, Improved biomass productivity and water useeYciency under water deWcit conditions in transgenic wheat constitu-tively expressing the barley HVA1 gene, Plant Sci. 155 (2000) 1–9.

[92] U. Schuppler, P.H. He, P.C.L. John, R. Munns, EVects of water stresson cell division and cell-division-cycle-2-like cell-cycle kinase activityin wheat leaves, Plant Physiol. 117 (1998) 667–678.


[93] T.J. MansWeld, C.J. Atkinson, Stomatal behaviour in water stressedplants, in: R.G. Alscher, J.R. Cumming (Eds.), Stress Responses inPlants: Adaptation and Acclimation Mechanisms, Wiley-Liss, NewYork, 1990, pp. 241–264.

[94] J.A. Lake, F.I. Woodward, W.P. Quick, Long-distance CO2 signallingin plants, J. Exp. Bot. 53 (2002) 183–193.

[95] S. Wilkinson, W.J. Davies, ABA-based chemical signalling: the co-ordination of responses to stress in plants, Plant Cell Environ. 25(2002) 195–210.

[96] W. Hartung, A. Sauter, E. Hose, Abscisic acid in the xylem: wheredoes it come from, where does it go to? J. Exp. Bot. 53 (2002) 27–37.

[97] W.J. Davies, S. Wilkinson, B. Loveys, Stomatal control by chemicalsignalling and the exploitation of this mechanism to increase wateruse eYciency in agriculture, The New Phytol. 153 (2002) 449–460.

[98] R.E. Sharp, Interaction with ethylene: changing views on the role ofabscisic acid in root and shoot growth responses to water stress, PlantCell Environ. 25 (2002) 211–222.

[99] P.G. Blackman, W.J. Davies, Root-to-shoot communication in maizeplants of the eVects of soil drying, J. Exp. Bot. 36 (1985) 39–48.

[100] J. Bota, J. Flexas, H. Medrano, Is photosynthesis limited bydecreased Rubisco activity and RuBP content under progressivewater stress? New Phytol. 162 (2004) 671–681.

[101] F. Loreto, D. Tricoli, G. Di Marco, On the relationship between elec-tron transport rate and photosynthesis in leaves of the C4 plant Sor-ghum bicolor exposed to water stress, temperature changes andcarbon metabolism inhibition, Aust. J. Plant Physiol. 22 (1995) 885–892.

[102] S. Meyer, B. Genty, Mapping intercellular CO2 mole fraction (Ci) inRosa rubiginosa leaves fed with abscisic acid by using chlorophyllXuorescence imaging: signiWcance of Ci estimated from leaf gasexchange, Plant Physiol. 116 (1998) 947–957.

[103] H. Li, L.A. Sherman, A redox-responsive regulator of photosynthesisgene expression in the cyanobacterium Synechocystis sp. strain PCC6803, J. Bacteriol. 182 (2000) 4268–4277.

[104] C.H. Foyer, G. Noctor, Redox sensing and signalling associated withreactive oxygen in chloroplasts, peroxisomes and mitochondria,Physiol. Plantarum 119 (2003) 355–364.

[105] B. Karpinska, G. Wingsle, S. Karpinski, Antagonistic eVects ofhydrogen peroxide and glutathione on acclimation to excess excita-tion energy in Arabidopsis, IUBMB Life 50 (2000) 21–26.

[106] R. Desikan, M.K. Cheung, J. Bright, D. Henson, J. Hancock, S. Neill,ABA, hydrogen peroxide and nitric oxide signaling in stomatalguard cells, J. Exp. Bot. 55 (2004) 205–212.

[107] C. Bowler, M.V. Montagu, D. Inze, Superoxide dismutase and stresstolerance, Ann. Rev. Plant Physiol. Plant Mol. Biol. 43 (1992) 83–116.

[108] F.A. Hoekstra, E.A. Golovina, J. Buitink, Mechanisms of plant des-iccation tolerance, Trends Plant Sci. 6 (2001) 431–438.

[109] H.H. Ratnayaka, W.T. Molin, T.M. Sterling, Physiological and anti-oxidant responses of cotton and spurred anoda under interferenceand mild drought, J. Exp. Bot. 54 (2003) 2293–2305.

[110] S. Ramanjulu, D. Bartels, Drought- and desiccation-induced modu-lation of gene expression in plants, Plant Cell Environ. 25 (2002)141–151.

[111] J.C. Jang, J. Sheen, Sugar sensing in higher plants, Trends Plant Sci. 2(1997) 208–214.

[112] M.M. Chaves, J.P. Maroco, J.S. Pereira, Understanding plantresponse to drought: from genes to the whole plant, Funct. PlantBiol. 30 (2003) 239–264.

[113] M.M. Chaves, EVects of water deWcits on carbon assimilation, J. Exp.Bot. 42 (1991) 1–16.

[114] C. Pinheiro, M.M. Chaves, C.P. Ricardo, Alterations in carbon andnitrogen metabolism induced by water deWcit in the stems and leavesof Lupinus albus L., J. Exp. Bot. 52 (2001) 1063–1070.

[115] J. Trouverie, C. The’venot, J.P. Rocher, B. Sotta, J.L. Prioul, Therole of abscisic acid in the response of a speciWc vacuolar invertaseto water stress in the adult maize leaf, J. Exp. Bot. 54 (2003) 2177–2186.

[116] W.H. Cheng, A. Endo, L. Zhou, et al., A unique short-chain dehy-dro-genase/reductase in Arabidopsis glucose signaling and abscisicacid biosynthesis and functions, The Plant Cell 14 (2002) 2732–2743.

[117] J.V. Pego, A.J. Kortstee, C. Huijser, S.C.M. Smeekens, Photosynthe-sis, sugars and the regulation of gene expression, J. Exp. Bot. 51(2000) 407–416.

[118] S.L. Ho, Y.C. Chao, W.F. Tong, S.M. Yu, Sugar co-ordinately anddiVerentially regulates growth- and stress-related gene expression viaa complex signal transduction network and multiple control mecha-nisms, Plant Physiol. 125 (2001) 877–890.

[119] P. Leon, J. Sheen, Sugar and hormone connections, Trends Plant Sci.8 (2003) 110–116.

[120] J.K. Zhu, Plant salt tolerance, Trends Plant Sci. 6 (2001) 66–71.[121] A. Maggio, S. Miyazaki, P. Veronese, T. Fujita, J.I. Ibeas, B. Damsz,

M.L. Narasimhan, P.M. Hasegawa, R.J. Joly, R.A. Bressan, Doesproline accumulation play an active role in stress-induced growthreduction? Plant J. 31 (2002) 699–712.

[122] T. Abebe, A.C. Guenzi, B. Martin, J.C. Chushman, Tolerance ofmannitol-accumulating transgenic wheat to water stress and salinity,Plant Physiol. 131 (2003) 1748–1755.

[123] T. Taji, C. Ohsumi, S. Iuchi, M. Seki, M. Kasuga, M. Kobayashi, K.Yamaguchi-Shinozaki, K. Shinozaki, Important roles of drought-and cold-inducible genes for galactinol synthase in stress tolerance inArabidopsis thaliana, Plant J. 29 (2002) 417–426.

[124] B. Shen, R.G. Jensen, H.J. Bohnert, Increased resistance to oxidativestress in transgenic plants by targeting mannitol biosynthesis tochloroplasts, Plant Physiol. 113 (1997) 1177–1183.

[125] B. Shen, R.G. Jensen, H.J. Bohnert, Mannitol protects against oxida-tion by hydroxyl radicals, Plant Physiol. 115 (1997) 527–532.

[126] A.K. Garg, J.K. Kim, T.G. Owens, A.P. Ranwala, Y.D. Choi, L.V.Kochian, R.J. Wu, Trehalose accumulation in rice plants confershigh tolerance levels to diVerent abiotic stresses, Proc. Natl. Acad.Sci. USA 99 (2002) 15898–15903.

[127] T.H.H. Chen, N. Murata, Enhancement of tolerance of abiotic stressby metabolic engineering of betaines and other compatible solutes,Curr. Opin. Plant Biol. 5 (2002) 250–257.

[128] D. Rontein, G. Basset, A. Hanson, Metabolic engineering of osmo-protectant accumulation in plants, Metab. Eng. 4 (2002) 49–56.

[129] H. Nayyar, D.P. Walia, Water stress induced proline accumulationin contrasting wheat genotypes as aVected by calcium and abscisicacid, Biol. Plantarum 46 (2003) 275–279.

[130] P.B. Kavi Kishor, Z. Hong, G.H. Miao, C.A.A. Hu, D.P.S. Verma,Overexpression of D1-pyrroline-5-carboxylate synthetase increasesproline production and confers osmotolerance in transgenic plants,Plant Physiol. 108 (1995) 1387–1394.

[131] M.E. Feder, G.E. Hofmann, Heat-shock proteins, molecular chaper-ones, and the stress response: evolutionary and ecological physiol-ogy, Ann. Rev. Physiol. 61 (1999) 243–282.

[132] W. Sun, C. Bernard, B.V.D. Cotte, M.V. Montagu, N. Verbruggen,At-HSP17.6A, encoding a small heat-shock protein in Arabidopsis,can enhance osmotolerance upon overexpression, Plant J. 27 (2001)407–415.

[133] T. Munnik, W. Ligterink, I. Meskiene, O. Calderini, J. Beyerly, et al.,Distinct osmosensing protein kinase pathways are involved in sig-naling moderate and severe hyperosmotic stress, Plant J. 20 (1999)381–388.

[134] M. Mikolajczyk, S.A. Olubunmi, G. Muszynska, D.F. Klessig, G.Dobrowolska, Osmotic stress induces rapid activation of a salicylicacid-induced protein kinase and a homolog of protein kinase ASK1in tobacco cells, Plant Cell 12 (2000) 165–178.

[135] D.B. DeWald, J. Torabinejad, C.A. Jones, J.C. Shope, A.R. Cange-losi, et al., Rapid accumulation of phosphatidylinositol 4,5-bisphos-phate and inositol 1,4,5-trisphosphate correlates with calciummobilization in salt-stressed Arabidopsis, Plant Physiol. 126 (2001)759–769.

[136] I. Heilmann, I.Y. Perera, W. Gross, W.F. Boss, Change in phosphoino-sitide metabolism with days in culture aVect signal transduction path-ways in Galdieria suphuraria, Plant Physiol. 119 (1999) 1331–1339.


[137] S. Takahashi, T. Katagiri, T. Hirayama, K. Yamaguchi-Shinozaki,K. Shinozaki, Hyperosmotic stress induced a rapid and transientincrease in inositol 1,4,5-trisphosphate independent of abscisic acidin Arabidopsis cell culture, Plant Cell Physiol. 42 (2001) 214–222.

[138] Y. Lee, Y.B. Choi, J. Suh, J. Lee, S.M. Assmann, et al., Abscisic acid-induced phosphoinositide turnover in guard cell protoplasts of Viciafaba, Plant Physiol. 110 (1996) 987–996.

[139] L. Xiong, M. Ishitani, H. Lee, J.K. Zhu, The Arabidopsis LOS5/ABA3 locus encodes a molybdenum cofactor sulfurase and modu-lates cold stress- and osmotic stress-responsive gene expression,Plant Cell 13 (2001) 2063–2083.

[140] T. Hirayama, C. Ohto, T. Mizoguchi, K. Shinozaki, A gene encodinga phosphatidylinositol-speciWc phospholipase C is induced by dehy-dration and salt stress in Arabidopsis thaliana, Proc. Natl Acad. Sci.USA 92 (1995) 3903–3907.

[141] D. Sanders, C. Brownlee, J.F. Harper, Communicating with calcium,Plant Cell 11 (1999) 691–706.

[142] Y. Wu, J. Kuzma, E. Marechal, R. GraeV, H.C. Lee, et al., Abscisicacid signaling through cyclic ADP-ribose in plants, Science 278(1997) 2126–2130.

[143] T. Munnik, H.J.G. Meijer, B. ter Riet, W. Frank, D. Bartels, A. Mus-grave, Hyperosmotic stress stimulates phospholipae D activity andelevates the levels of phosphatidic acid and diacylglycerol pyrophos-phate, Plant J. 22 (2000) 147–154.

[144] W. Frank, T. Munnik, K. Kerkmann, F. Salamini, D. Bartels, WaterdeWcit triggers phospholipase D activity in the resurrection plantCraterostigma plantagineum, Plant Cell 12 (2000) 111–123.

[145] T. Katagiri, S. Takahashi, K. Shinozaki, Involvement of a novel Ara-bidopsis phospholipase D, AtPLD delta, in dehydration-inducibleaccumulation of phosphatidic acid in stress signaling, Plant J. 26(2001) 595–605.

[146] H. El Maarouf, Y. Zuily-Fodil, M. Gareil, A. d’Arcy-Lameta, A.T.Pham-Thi, Enzymatic activity and gene expression under waterstress of phospholipase D in two culitivars of Vigna unguiculata L.Walp. DiVering in drought tolerance, Plant Mol. Biol. 39 (1999)1257–1265.

[147] T. Jacob, S. Ritchie, S.M. Assmann, S. Gilroy, Abscisic acid signaltransduction in guard cells is mediated by phospholipase D activity,Proc. Natl. Acad. Sci. USA 96 (1999) 12192–12197.

[148] J. Leung, J. Giraudat, Abscisic acid signal transduction, Annu. Rev.Plant Physiol. Plant Mol. Biol. 49 (1998) 199–222.

[149] K. Shinozaki, K. Yamaguchi-Shinozaki, Molecular responses todehydration and low temperature: diVerences and cross-talkbetween two stress signaling pathways, Curr. Opin. Plant Biol. 3(2000) 217–223.

[150] R.R. Finkelstein, S.S.L. Gampala, C.D. Rock, Abscisic acid signalingin seeds and seedlings, Plant Cell 14 (suppl.) (2002) S15–S45.

[151] M. Koornneef, K.M. Leon-Kloosterziel, S.H. Schwartz, J.A.D.Zeevaart, The genetic and molecular dissection of abscisic acid bio-synthesis and signal transduction in Arabidopsis, Plant Physiol. Bio-chem. 36 (1998) 83–89.

[152] S. Liotenberg, H. North, A. Marion-Poll, Molecular biology and reg-ulation of abscisic acid biosynthesis in plants, Plant Physiol. Bio-chem. 37 (1999) 341–350.

[153] F. Llorente, J.C. Oliveros, J.M. Martinez-Zapater, J. Salinas, A freez-ing-sensitive mutant of Arabidopsis, frs1, is a new aba3 allele, Planta211 (2000) 648–655.

[154] E. Marin, L. Nussaume, A. Quesada, M. Gonneau, B. Sotta, et al.,Molecular identiWcation of zeaxanthin expoxidase of Nicotianaplumbaginifolia, a gene involved in abscisic acid biosynthesis andcorresponding to the ABA locus of Arabidopsis thalian, EMBO J. 15(1996) 2331–2342.

[155] B.C. Tan, S.H. Schwartz, J.A.D. Zeevaart, D.R. Mc-Carty, Geneticcontrol of abscisic acid biosynthesis in maize, Proc. Natl. Acad. Sci.USA 94 (1997) 12235–12240.

[156] Y. Uno, T. Furihata, H. Abe, R. Yoshida, K. Shinozaki, K. Yamagu-chi Shinozaki, Arabidopsis basic leucine zipper transcription factorsinvolved in an abscisic acid-dependent signal transduction pathway

under drought and high-salinity conditions, Proc. Natl. Acad. Sci.USA 97 (2000) 11632–11637.

[157] H. Knight, M.R. Knight, Abiotic stress signaling pathways: speciWc-ity and cross talk, Trends Plant Sci. 6 (2001) 262–267.

[158] L. Xiong, J.K. Zhu, Abiotic stress signal transduction in plants: molec-ular and genetic perspectives, Physiol. Plant 112 (2001) 152–166.

[159] D. Sanders, J. Pelloux, C. Brownlee, J.F. Harper, Calcium at thecrossroads of signaling, Plant Cell 14 (suppl.) (2002) S401–S417.

[160] H. Knight, M.R. Knight, Imaging spatial and cellular characteristicsof low temperature calcium signature after cold acclimation in Ara-bidopsis, J. Exp. Bot. 51 (2000) 1679–1686.

[161] S.J. Gilmour, D.G. Zarka, E.J. Stockinger, M.P. Salazar, J.M. Hough-ton, M.F. Thomashow, Low temperature regulation of the ArabidopsisCBF family of AP2 transcriptional activators as an early step in cold-induced COR gene expression, Plant J. 16 (1998) 433–443.

[162] J. Medina, M. Bargues, J. Terol, M. Perez-Alonso, J. Salinas, TheArabidopsis CBF family is composed of three genes encoding AP2domain containing proteins whose expression is regulated by lowtemperature but not by abscisic acid or dehydration, Plant Physiol.119 (1999) 463–470.

[163] J.C. Kim, S.H. Lee, Y.H. Cheng, et al., A novel cold-inducible zincWnger protein from soybean, SCOF-1, enhances cold tolerance intransgenic plants, Plant J. 25 (2001) 247–259.

[164] H.I. Choi, J.H. Hong, J.O. Ha, J.Y. Kang, S.Y. Kim, ABFs a familyof ABA-responsive element binding factors, J. Biol. Chem. 275(2000) 1723–1730.

[165] H. Abe, K. Yamaguchi-Shinozaki, T. Urao, T. Iwasaki, D. Hosak-awa, K. Shinozaki, Role of Arabidopsis MYC and MYB homologsin drought- and abscisic acid regulated gene expression, Plant Cell 9(1997) 1859–1868.

[166] R.E. Zielinski, Calmodulin and calmodulin-binding proteins inplants, Ann. Rev. Plant Physiol. Plant Mol. Biol. 49 (1998) 697–725.

[167] S. Luan, J. Kudla, M. Rodríguez-Concepción, S. Yalovsky, W. Gru-issem, Calmodulins and calcineurin B-like proteins: calcium sensorsfor speciWc signal response coupling in plants, Plant Cell 14 (2002)S389–S400.

[168] S.K. Sopory, M. Munchi, Protein kinases and phosphatases andtheir role in cellular signaling in plants, Crit. Rev. Plant Sci. 17(1998) 245–318.

[169] S. Pandey, S.B. Tiwari, K.C. Upadhyaya, S.K. Sopory, Calcium sig-naling: linking environmental signals to cellular functions, Crit. Rev.Plant Sci. 19 (2000) 219–318.

[170] D. Gong, Y. Guo, K.S. Schumaker, J.K. Zhu, The SOS3 family ofcalcium sensors and SOS2 family of protein kinases in Arabidopsis,Plant Physiol. 134 (2004) 919–926.

[171] K.N. Kim, Y.H. Cheong, J.J. Grant, G.K. Pandey, S. Luan, CIPK3, acalcium sensor-associated protein kinase that regulates abscisic acidand cold signal transduction in Arabidopsis, Plant Cell 15 (2003)411–423.

[172] Y.H. Cheong, K.N. Kim, G.K. Pandey, R. Gupta, J.J. Grant, S. Luan,CBL1, a calcium sensor that diVerentially regulates salt, drought,and cold responses in Arabidopsis, Plant Cell 15 (2003) 1833–1845.

[173] G.K. Pandey, Y.H. Cheong, K.N. Kim, J.J. Grant, L. Li, W. Hung, C.D‘Angelo, S. Weinl, J. Kudla, S. Luan, The calcium sensor calcineu-rin B-Like 9 modulates ABA sensitivity and biosynthesis in Arabid-opsis, Plant Cell 16 (2004) 1912–1924.

[174] V. Albrecht, S. Weinl, D. Blazevic, C. D’Angelo, O. Batistic, U.Kolukisaoglu, R. Bock, B. Schulz, K. Harter, J. Kudla, The calciumsensor CBL1 integrates plant responses to abiotic stresses, Plant J. 36(2003) 457–470.

[175] A. Nozawa, N. Koizumi, H. Sano, An Arabidopsis SNF1-relatedprotein kinase, AtSR1, interacts with a calcium-binding protein,AtCBL2, of which transcripts respond to light, Plant Cell Physiol. 42(2001) 976–981.

[176] Y.S. Hwang, P.C. Bethke, Y.H. Cheong, H.S. Chang, T. Zhu, R.L.Jones, A gibberellin-regulated calcineurin B in rice localizes to thetonoplast and is implicated in vacuole function, Plant J. 138 (2005)1347–1358.


[177] S.J. Wu, D. Lee, J.K. Zhu, SOS1, a genetic locus essential forsalt tolerance and potassium acquisition, Plant Cell 8 (1996)617–627.

[178] M.J. Sanchez-Barrena, M. Martinez-Ripoll, J.K. Zhu, A. Albert, Thestructure of the Arabidopsis thaliana SOS3: molecular mechanism ofsensing calcium for salt stress response, J. Mol. Biol. 345 (2005)1253–1264.

[179] D. Gong, Z. Gong, Y. Guo, X. Chen, J.K. Zhu, Biochemical andfunctional characterization of PKS11, a novel Arabidopsis proteinkinase, J. Biol. Chem. 277 (2002) 28340–28350.

[180] E.J. Lee, Hiromichi, H. Sano, N. Koizumi, Sugar responsible and tis-sue speciWc expression of a gene encoding AtCIPK14, an Arabidop-sis CBL-interacting protein kinase, Biosci. Biotechnol. Biochem. 69(2005) 242–245.

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Cold, salinity and drought stresses: An overview - UAMuam.es/personal_pdi/ciencias/solsa/revsaltstress.pdf · Cold, salinity and drought stresses: An overview Shilpi Mahajan, Narendra