Keywords

1 Introduction

Plants are forced to exposure to various abiotic stresses including salt, drought, extreme temperatures (cold and heat), UV radiations, etc., since they are unable to move to more favorable places. These abiotic stresses adversely affect the plant metabolic activities and are also responsible for crop loss to a greater extent (Mahajan and Tuteja 2005; Tuteja 2007a). During stress, the upregulation of many genes has been reported which helps the plant to withstand the stress conditions and lead to plant adaptation (Tuteja 2009a).

Plants perceive the external and internal signals and are used to regulate various responses for its development. Exposure to stimuli causes membrane depolarization of the plant cells with 10–30 s, which is coordinated with early Ca2+ influx (reviewed by Tuteja and Sopory 2008). Signals are perceived by the membranes and therefore membrane events are the likely routes for signal generation and transduction (Tuteja and Sopory 2008). Different signaling pathways can operate independently to each other and can modulate other pathways (Kaur and Gupta 2005). Sometimes, components of pathways are dependent on each other and can cross talk among them.

Different molecules have been reported to have a role in signal transduction. The present review throws light on various signaling molecules and pathways like calcium and MAPK signaling pathways during abiotic stress. We also tried to cover the role of phytohormones such as ABA, SA, JA, BR, etc., in abiotic stress signaling.

2 Calcium Signaling

Ca2+ regulates a range of activities within the cell such as cell division and elongation, cytoplasmic streaming, photomorphogenesis, and plant defense against environmental stresses (Song et al. 2008; Tuteja 2009b; Kader and Lindberg 2010). It functions as the central node in overall signaling web and has a promising role in stress tolerance (Tuteja and Sopory 2008). During abiotic stress, Ca2+ acts as a second messenger under various stress conditions (Knight 2000). Ca2+ signatures have a leading role in numerous physiological processes such as regulation of stomatal apertures in plants (Allen et al. 2001). Ca2+ signatures change according to the nature of stress (Kiegle et al. 2000), duration of stress (Plieth et al. 1999), earlier exposure to any stress (Knight et al. 1997), and the type of tissue exposed to stress (Kiegle et al. 2000). For studying the role of calcium and its dynamics, various pharmacological and transgenic approaches have been utilized. The calcium reporter protein aequorin has proved to be very useful for such studies in revealing calcium fluxes within one cell and in different tissues.

Calcium-binding proteins act as stress sensors, get conformationally transformed upon binding with Ca2+, which facilitates their interaction with downstream effector molecules (Clapham 2007; Gifford et al. 2007; Tuteja 2009b). Elongation factor (EF)-hand motif, which generally occurs in pairs, is the most common motif present in these Ca2+-binding proteins and helps in high-affinity binding of Ca2+. It has a helix-loop-helix structure and is reported in about 250 proteins of Arabidopsis (Day et al. 2002). In plants these EF-hand proteins comprise three different categories, namely, CDPKs (Ca2+-dependent protein kinases), CaMs (calmodulins) and CMLs (CaM-like proteins), and the CBLs (calcineurin B-like proteins) (Fig. 14.1). Of these, only CDPKs act as “responders,” as they are capable of directly transducing signals through their catalytic activity. CaMs/CMLs and CBLs are only sensors for regulating the downstream targets. CMLs, CDPKs, and CBLs are restricted to plants and some protists, whereas CaM is universal to all eukaryotes.

Fig. 14.1
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Calcium-dependent signaling. Environmental stress responses are perceived by cell surface which in turn increases cytosolic Ca2+ concentration. Ca2+ got activated by cytosolic Ca2+ and regulates downstream targets leading to physiological responses

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2.1 Calcium-Dependent Protein Kinases

CDPKs have been reported from several plants (Kawasaki et al. 2001; Seki et al. 2002; Ozturk et al. 2002; Asano et al. 2011) and are specific for a particular osmotic stress response. Studies on salt-tolerant and salt-sensitive rice varieties revealed specific CDPKs to be induced earlier with a sustained expression in the tolerant variety in comparison to the sensitive variety (Kawasaki et al. 2001). Arabidopsis CDPKs are also highly specific (Sheen 1996). Out of several CDPKs tested, only AtCDPK1 and AtCDPK1a were able to transcriptionally activate selected reporter genes. CDPKs are five domain-containing proteins, and range from ∼40 to 90 kDa in size. Their catalytic domain consists of a highly conserved serine/threonine kinase region, whereas, their N-terminal variable domain ranges from 21 to 185 amino acids in length (Klimecka and Muszynska 2007). Adjacent to the kinase domain is a pseudosubstrate-containing autoinhibitory junction domain that interacts with the active site and inhibits its kinase activity. Next to the autoinhibitory domain is the CaM-like domain (CLD) which is responsible for its Ca2+-binding activity. Their C-terminal domain is relatively short and variable.

Plants have a large family of Ca2+-dependent protein kinase (CDPKs) and have an important role in signaling during abiotic stresses like drought, wounding, and cold (Fig. 14.2). Induction of CDPK by osmotic stress has been shown by many workers in various plants (Kawasaki et al. 2001; Seki et al. 2002; Ozturk et al. 2002; Witte et al. 2010; Franza et al. 2011; Wurzinger et al. 2011). Induction of CDPK exists in longer duration in tolerant varieties of rice than in the salt-sensitive variety (Kawasaki et al. 2001). Martin and Busconi (2001) have reported that cold stress activates the membrane-associated CDPK in rice plants. Saijo et al. (2001) have also reported that overexpression of OsCDPK7 in rice provides cold and osmotic stress tolerance in these plants. Sheen (1996) has demonstrated that CDPK is involved in signal transduction. In his experiment, he demonstrated that in protoplast of maize leaf the expression of stress-responsive HVA1 was induced by active AtCDPK1. Sheen (1996) has also reported that AtCDPK1 activation is blocked by a protein phosphatase type 2C (AtPP2CA). Enhanced induction of CBF1, RAB18, RC12A, and LT178 (i.e., RD29A) gene expression has been observed in AtPP2CA-silenced Arabidopsis plants during cold and ABA treatment (Tahtiharju and Palva 2001). Romeis et al. (2001) also reported that pathogen infection also activated CDPK in plants.

Fig. 14.2
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Signaling through CDPK and protein phosphorylation by Ca2+ signatures

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2.2 Calmodulin and Other Calcium-Binding Proteins

Elevated levels of calcium within a cell activate various calcium-binding proteins which in turn induce specific kinases. On the basis of function, calcium-binding proteins are classified into two groups: (1) trigger proteins and (2) buffer proteins. The trigger proteins are activated when they bind with Ca2+ and after that they interact with other proteins and alter their activity. The trigger-type CaBPs are calmodulin (CaM), CaM-binding proteins, Ca2+-dependent protein kinase, and phosphatase (Reddy 2001).

Stress causes increase in inositol 1,4,5-triphosphate (IP3). Activation of phospholipase C (PLC) results in hydrolysis of PIP2 to IP3. IP3 is regarded as the activator of vacuolar calcium channels in plants. During stress, the increase in cytosolic Ca2+ is due to the activation of IP3-dependent calcium channels (Braam 2005; Boudsocq and Laurière 2005). Furthermore, calcium-binding proteins (CaBP) or calcium sensors recognize and translate the information provided in calcium signatures (Tuteja and Mahajan 2007) and pass the information downstream for regulation of gene expression.

These CaBPs are partially responsible in modulating the intracellular calcium levels. The calcium-binding proteins can be regulated either in a cell- or in a tissue-specific manner. NaCl-inducible Ca2+/calmodulin-dependent protein kinase in pea (PsCCaMK) was reportedly specific to roots (Pandey et al. 2002). A calmodulin-binding transcription activator family was shown to be specific only to the multicellular organisms (Bouche et al. 2002). Other examples of osmotic stress-activated calcium-binding proteins include Arabidopsis protein AtCP1, the membrane-associated rice protein OsEFA27, and the Arabidopsis counterpart RD20 (Frandsen et al. 1996; Jang et al. 1998). Some CaBPs can be negative regulators of osmotic stress also; one example is the calmodulin-binding protein in Arabidopsis, AtCaMBP25. Although it is upregulated by osmotic stress, its overexpression renders plants sensitive to osmotic or salt stresses and its antisense transgenics show improved tolerance (Perruc et al. 2004).

Calmodulin, an important CaBP, is a small acidic protein and is responsible for the regulation of intracellular Ca2+ levels. Increased Ca2+ concentration activates calmodulin which then induces specific kinases. Calmodulin is a very important calcium-binding protein in Ca2+ signaling and has been found to be involved in biotic and abiotic stresses (Fig. 14.3) (Reddy 2001; Tuteja and Mahajan 2007; Tuteja and Sopory 2008). Plants have been found to possess unique Ca2+ sensors like calmodulin-like proteins (CMLs), and Arabidopsis contains 50 such proteins (Tuteja and Sopory 2008). The calmodulin-like proteins differ from calmodulin in having more than 148 amino acids and also have one to six EF-hand motifs. These CMLs have been found to play a role as Ca2+ sensor during stress in plants (Vanderbeld and Snedden 2007). Certain Ca2+-binding proteins do not contain EF-hand motifs, such as calreticulin, annexins, calnexin, phospholipase D (PLD), and pistil-expressed Ca2+-binding proteins.

Fig. 14.3
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Ca2+ signatures induce transcriptional responses

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PLD activity has been reported to be involved in ethylene and ABA responses, synthesis of α-amylase in aleurone cells, closing of stomata, responses to pathogens, leaf senescence, and drought tolerance (reviewed by Tuteja and Sopory 2008).

Annexins have been reported to be involved in biological membrane organization and functions (Tuteja and Mahajan 2007). The exact function of annexins is not clear yet but are thought to play a role in secretary processes and they have ATPase and peroxidase activities (Tuteja and Mahajan 2007). Annexins have been regarded to have a role in stress responses (Gorecka et al. 2007a). Annexin At1 of Arabidopsis thaliana (AnnAt1) plays a vital role in pH-mediated cellular responses to environmental stress (Gorecka et al. 2007b).

Calnexin (CNX) is an important calcium-binding protein and is an endoplasmic reticulum (ER) type 1 integral membrane protein. CNX behaves as a molecular chaperone and has a leading role in the recognition of misfolded proteins, lectin-like activity, and Ca2+ binding (Sarwat and Tuteja 2007). Till date, the role of calnexin in stress has not been reported but it is considered that it has a role in ER stress response in plants (reviewed by Tuteja and Sopory 2008).

In plants, another group of Ca2+ sensors is the SOS (salt overly sensitive) family and is responsible for calcium-mediated pathway for salinity stress tolerance (Mahajan et al. 2008). Zhu (2003) has isolated sos mutants (sos1, sos2, and sos3) from Arabidopsis which are hypersensitive to salt. The sos mutants have been shown to accumulate more proline under salt stress which gives protection to the salt-stressed plants (Liu and Zhu 1998). Cloning and characterization of sos genes (SOS1–SOS3) has opened new doors for ion homeostasis and plant tolerance to salt. It has been reported that sos1, sos2, and sos3 function in a common pathway leading to salt tolerance (Halfter et al. 2000; Zhu 2000). Sos1 is activated by sos3–sos2 complex and has been reported by many workers. Shi and Zhu (2002) demonstrated that if sos1 alone was allowed to express in yeast cells, slight enhancement in salt tolerance was observed. However, if sos1 was expressed with sos3 and sos2 the yeast cells showed more tolerance to salt. Qiu et al. (2002) have reported very low Na+/H+ exchange activity in sos mutant plants in comparison with wild type in plasma membrane vesicles. Addition of activated sos2 protein to isolated membrane vesicles of mutant plants showed that the exchange activity increased in sos2 and sos3 but remains unaffected in sos1 mutants. The results lead to the conclusion that Na+/H+ exchange activity of sos1 was stimulated by sos3 and sos2 (reviewed by Xiong et al. 2002). sos3–sos2 also regulate Na+ transporter AtHKT1, which is a salt tolerance effector (Uozumi et al. 2000). Homologs of HKT1 in other plant species revealed that it can be either a K+ transporter or a Na+/K+ cotransporter (Horie et al. 2001; Liu et al. 2001). Rus et al. (2001) demonstrated that mutation in AtHKT1 of Arabidopsis suppressed the salt hepersensitivity phenotype of sos3, leading to the concept that activity of AtHKT1, the Na+ influx transporter, may be inhibited by SOS3 (reviewed by Xiong et al. 2002).

As compared to caltractin and calmodulin, sos3 binds with Ca2+ with low affinity and has three EF-hand motifs (Ishitani et al. 2000). In sos3 mutation, one of the EF-hand motifs gets mutated, thus preventing it to bind Ca2+ (Ishitani et al. 2000). sos4 and sos5 have recently been characterized (Mahajan et al. 2008). sos4 encodes a pyridoxal (PL) kinase which has a role in the biosynthetic pathway of pyridoxal-5-phosphate, which is an active form of vitamin B6. Sos5 is a putative cell surface adhesion protein and has a role in normal cell expansion. Under salt stress, sos5 has been found to have a role of maintenance of cell wall integrity (reviewed by Tuteja and Sopory 2008).

2.3 Calcineurin B-Like Proteins

These proteins possess four Ca2+-binding EF-hand domains and have significant identity to calcineurin B subunit and neural calcium sensor from yeast and animals (Kudla et al. 1999). The CBL protein family and their corresponding kinases (CIPKs) together form a complex and dynamic Ca2+ decoding signaling network (Fig. 14.4). Together they show Ca2+-binding functionality and kinase activity (Mahajan et al. 2006a).

Fig. 14.4
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Signaling through CBL/CIPK and protein phosphorylation by Ca2+ signatures

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CIPKs have a conserved N-terminal kinase domain and C-terminal regulatory domain, separated from the kinase domain by a variable junction domain. A conserved NAF domain present in the divergent regulatory domain is required for interaction with CBLs. CBL binds to the NAF domain of CIPKs, releases the C-terminal (autoinhibitory) domain from the kinase domain, in turn transforming the kinase into its active state (Guo et al. 2001; Mahajan et al. 2006a).

Bioinformatic analysis shows a number of components of both proteins in the families of CBLs and CIPKs; 10 CBLs and 26 CIPKs in Arabidopsis, 10 CBLs and 30 CIPKs in rice (Albrecht et al. 2001; Kolukisaoglu et al. 2004; Weinl and Kudla 2009). However, several species of green algae possess single CBL and CIPK genes, other plant species can have multiples, like Physcomitrella contains four CBLs and seven CIPKs and the fern Selaginella moellendorfii has five CBLs and five CIPKs (Batistic and Kudla 2009; Weinl and Kudla 2009); thus showing evolutionary complexity of these CBL and CIPK protein families from lower to higher organisms.

CBLs are found to be localized all through the cell. In Arabidopsis itself, four CBLs are present at the plasma membrane, four at the vacuolar membrane, and two in the cytoplasm and nucleus (Batistic et al. 2008; Cheong et al. 2007; D’ Angelo et al. 2006; Kim et al. 2007; Weinl and Kudla 2009). The pea CBL was reported to be exclusively localized in the cytosol whereas pea CIPK is localized in the cytosol and the outer membrane (Mahajan et al. 2006b). The pea CBL and CIPK were reported to be coordinately upregulated in response to high NaCl, cold, wounding and also in response to calcium and salicylic acid, whereas drought and abscisic acid had no effect on the expression of these genes (Mahajan et al. 2006b).

Our knowledge has been greatly facilitated by the reverse genetic approaches. A CIPK3 loss-of-function (LOF) mutant showed its involvement in regulating ABA-induced gene expression and ABA responses during seed germination (Kim et al. 2003). Through these studies, CBL1 has came out to function in an ABA-independent manner in controlling responses to drought, cold, and salinity (Albrecht et al. 2003; Cheong et al. 2003), whereas its closely related Ca2+ sensor CBL9 renders plants hypersensitive to ABA (Pandey et al. 2004). Interestingly, when CIPK1 complexes with CBL1, it mediates the ABA-dependent pathway (D’ Angelo et al. 2006). CBL1 and CBL9 activate CIPK23, and the complex regulates the activity of the shaker-like K+ channel ARABIDOPSIS K+ TRANSPORTER1 (AKT1) and thus contributes in K+ homeostasis within the cell (Kudla et al. 1999). This complex also has a role in stomatal regulation under drought conditions.

CBL–CIPK network is a central and critical system functioning in response to a broad variety of stimuli in order to decode Ca2+ signals. Each CBL and each CIPK can make alternative protein interactions and are part of multifunctional signaling component, thus determining the flow of information (Mahajan et al. 2006a).

3 Signaling Through MAPK Kinases

3.1 Historical Background

Sturgill and Ray (1986) for the first time discovered MAPK in animal cells and named it as microtubule-associated protein-2 kinase (MAP-2 kinase). It was renamed as mitogen-activated protein kinase (MAP kinase) 1 by Rossomando et al. (1987) as mitogen was found to activate the group of proteins and this kinase was found to be related to these proteins (reviewed by Sanan-Mishra et al. 2006; Sinha et al. 2011). In 1990, it was reported as serine/tyrosine kinase that belonged to a multigene family (Gotoh et al. 1990). MAP kinase genes (MsERK1) in plant system was for the first time reported from alfalfa in 1993 (Duerr et al. 1993) and D5 kinase in pea (Stafstrom et al. 1993). After that they were reported from various plants like tobacco, Arabidopsis, etc. (Jonak et al. 1994). Three major groups of MAPKs are found in yeast and animals: (1) extracellular signal-regulated kinases (ERK) (Cobb et al. 1994), (2) c-Jun amino (NH2)-terminal kinases or stress-activated protein kinases (JNS/SAPK) (Davis 1994), and (3) high osmolarity glycerol response or p38 kinases (Hog/p38) (Landry and Huot 1995). MAP kinase genes reported in plants belong to the ERK subfamily (Hirt 2000) and transmit a broader range of stimuli (Ligterink and Hirt 2001). Stafstrom et al. (1993) reported that the first MAP kinase gene isolated from pea has 41% identity with plant cdc2 kinases and other kinases involved in osmosensing. MAP kinases generally function as a cascade in which MAPKKK phosphoralates and activates MAPKK which in turn activates MAPK. All the three kinases are interlinked together and are also called extracellular receptor kinases (Hirt 2000; Sanan-Mishra et al. 2006). Different plant MAPKs recognize different substrates (Jonak et al. 2002) because of the high similarity in catalytic domain and little similarity in N-termini. The activation domain of most of the plant MAPKs contains TEY (Thu-Glu-Tyr) sequence and is similar to ERK/MAP kinases group of mammals and yeast (Hirt 2000). The activation domain of some MAPKs possesses TDY (Thr-Asn-Tyr) sequence and is closer to the p38/Hog group of mammals and yeast (Tena et al. 2001). No plant MAPK has been found which has TPY (Thr-Pro-Tyr) sequence at its activation domain. There are three functionally linked protein kinase viz.: MAP3K, MAP2K, and MAPKs. 60 MAP3Ks, 10 MAP2Ks, and 20 MAPK have been reported in Arabidopsis thaliana by Group et al. (2002).

3.2 MAP3Ks

It is also known as MAPKKKs or MEKKs. MAP3Ks constitute a diverse family of kinases and are divided into two subfamilies viz. MEKK1 and RAF-like kinases. The MEKK1-like subfamily members are similar to mammalian MEKK1 and to yeast STE11 and BCK1 and RAF-like kinases are similar to mammalian RAF1 MAPK (Group et al. 2002). About 8–10 algal MAP3Ks are found in Chlamydomonas and Volvox and about 40–60 in Sorghum and Populous. One of the important things in MEKK-like kinases is having a conserved catalytic domain and Arabidopsis has ten members in this group. Arabidopsis has been reported to have 80 putative MAPKKKs whereas rice has 75 members (Rao et al. 2010). Plant species having homologs of MAPKKKs have been identified, including the MEKK-like protein kinases, oxidative stress-activated MAP triplekinase 1 (OMTK1) from alfalfa (Nakagami et al. 2004), ANP1, ANP2, ANP3 (Kovtun et al. 2000), YDA (Lukowitz et al. 2004) from Arabidopsis, NPK1 (Nicotiana protein kinase 1) from tobacco (Nishihama et al. 2001), Raf-like protein kinase, EDR1 (enhanced disease resistance 1), and CTR1 (constitutive triple response 1) from Arabidopsis (Frye et al. 2001; Kieber et al. 1993). MEKK-like proteins have been found to participate in canonical MAP kinase cascades that activate downstream MAP2Ks (Rodriguez et al. 2010). The two RAF-like MAP3Ks are CTR1 and EDR1, and these two RAF-like MAP3Ks are found to participate in ethylene-mediated signaling and defense responses (Frye et al. 2001; Huang et al. 2003). It has also been reported that CTR1 and EDR1 do not participate in a canonical MAPK cascade (Rodriguez et al. 2010).

3.3 MAP2Ks

It is also known as MEKs and MKKs and is divided into four groups viz. Groups A, B, C, and D (Hamel et al. 2006). MKK1 and MKK2 belong to Group A and act upstream of the MAPK MAK4 (Ichimura et al. 1998). There are several reports about the involvement of MKK2 in response to cold and salinity stress and apart from this both MKK1 and MKK2 mediate innate immunity responses (Meszaros et al. 2006; Qiu et al. 2008). Group B includes MKK3. One of the distinguishing features of this group is nuclear transfer factor (NTF) domain (Hamel et al. 2006). Steggerda and Paschal (2002) reported that NTF enhances the nuclear import of cargo proteins, which suggests that plant MAP2Ks with NTF domains are involved in cytoplasmic nuclear trafficking. MKK3 has been found to participate in cascades that are elicited by pathogens and are dependent on jasmonic acid (JA) signaling (Doczi et al. 2007; Takahashi et al. 2007). Group C has MKK4 and MKK5 and Group D has rest of the kinases from MKKs 7–10. In general, all plant phyla appear to use a more limited number of MKKs compared to other MAPK components. Rodriguez et al. (2010) have reported a single MKK each for the algae Chlamydomonas and Volvox. This indicates that the same MAP2K may function in several different MAPK modules. Genetic analysis has shown that closely related pairs of plant MAP2Ks have similar functions, e.g., MKK1 and MKK2 are proposed to activate MAPK MPK4 (Qiu et al. 2008), whereas MKK4 and MKK5 act upstream of MPK3 and MPK6, apparently in a redundant manner (Asai et al. 2002). In rice system, MKK genes exhibit differential regulation under different abiotic stresses (Kumar et al. 2008).

3.4 MAPKs

These are also known as MPKs and have been divided into four groups (A–D) (Group et al. 2002). The activation domain of Groups A, B, and C contains TEY (Thu-Glu-Tyr) sequence similar to ERK kinases of animals. The activation domain of Group D contains TDY (Thr-Asn-Tyr) sequence (Rodriguez et al. 2010). MPK3 and MPK6 belong to Group A and have been reported to have a role in developmental processes and also shows responses against biotic and abiotic stresses (Zhang and Klessig 2001; Seo et al. 2007; Sinha et al. 2011). MPK4 belongs to Group B and has a role in pathogen defense and abiotic stress responses (Andreasson et al. 2005; Brodersen et al. 2006; Qiu et al. 2008). The characteristic feature of Group D MAPKs is a C-terminal docking domain that may act as a docking site for MAP2Ks (Yoo et al. 2008).

3.5 MAPKs and Abiotic Stress Signaling

The mitogen-activated protein kinase (MAPK/MPKs) cascades transducer environmental cues into intracellular responses. The stimulated plasma membrane receptors activate MAP kinase kinase kinase and then the sequential phosphorylation ensues as MAP3Ks activate downstream MAP kinase kinase that in turn activates MAPKs (Sinha et al. 2011) (Fig. 14.5). MAPKs then target various effector proteins in the cytoplasm or nucleus, which include other kinases, enzymes, or transcription factors (Khokhlatchev et al. 1998; Sinha et al. 2011; Wurzinger et al. 2011).

Fig. 14.5
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The MAP kinase signal transduction pathway (Adopted from Ahmad et al. 2008)

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Abiotic stress is responsible for the activation of MAPK genes and increased MAPK activity. MEKK1, MKK2, MPK4, and/or MPK6 have been activated during salt, drought, and cold stress (Teige et al. 2004; Sinha et al. 2011). Temperature stress (26–38°C) induces 50 kDa MAP kinase in tomatoes. Induction of MPK3 during cold and salinity in Arabidopsis has also been reported by different workers. Ichimura et al. (2000) reported that MPK4 and MPK6 are activated during low-temperature and osmotic stress. The role of several MAP kinases in response to salinity has also been reported by Sanan-Mishra et al. (2006). Matsuoka et al. (2002) demonstrated the role of MAPK kinase (MKK1) in abiotic stress signaling. Multiple abiotic stresses such as wounding, cold, drought, and salinity activated MKK1 which activates downstream MPK4 (Matsuoka et al. 2002). In Arabidopsis a specific MAPKK kinase, ANP1, was activated by H2O2. This ANP1 activates MPK3 and MPK6 and its positive regulator nucleoside diphosphate kinase (NDP) 2 (Moon et al. 2003). Overexpression of ANP1 shows tolerance to heat shock, freezing, and salt stress tin transgenic plants (Kovtun et al. 2000). Plants expressing AtNDPK2 show reduction in H2O2 accumulation and tolerance to multiple stresses like cold, salt, and oxidative stress. Munnik et al. (1999) demonstrated that a 46 kDa MAP kinase named SIMK activated during moderate hyperosmotic stress in alfalfa. At severe hyperosmotic stress, a smaller kinase gets activated and the activation of SIMK was not observed at severe hyperosmotic stress suggesting that the two kinases function at different stress levels. Mikolajczyk et al. (2000) showed that a salicylic acid-induced protein kinase (SIPK) was activated in tobacco cells under hyperosmotic stress. In tobacco, MAP kinase is activated by multiple stresses like hyperosmotic, hypoosmotic, salicylic acid, and fungal elicitors. A 40 kDa kinase was activated in Arabidopsis by salt stress in a calcium and ABA-independent manner (Hoyos and Zhang 2000). MPK4 acts as a negative regulator in plant defense mechanisms. Tang et al. (2005) demonstrated that Arabidopsis EDR1 acts as negative regulator of disease resistance and drought. The edr1 mutant containing a kinase-deficient form of the EDR1 gene exhibits enhanced resistance to pathogens. The edr1 mutants could also enhance stress responses and spontaneous necrotic lesions under drought conditions (Tang et al. 2005). Accumulation of H2O2, superoxide anions, and hydroxyl radicals during abiotic stress causes oxidative burst in cells (Samuel and Ellis 2000). Plants can withstand this oxidative stress by production of antioxidant enzymes like catalase, which decompose H2O2 in the cells. Xing et al. (2008) demonstrated that AtMKK1 mediates ABA-induced CAT1 expression in Arabidopsis thaliana. The Arabidopsis mutants mkk1 and mpk6 were altered in their responses to ABA and desiccation stress and the results showed that MKK1–MPK6 regulate H2O2 metabolism by CAT1 (Xing et al. 2008). Nakagami et al. (2004) reported that MEKK1–MPK4 cascades are an important part of ROS metabolism. During abiotic stress, two signaling events must occur to induce defense responses in plant cells: one is the inhibition of negative regulators such as EDR1 and the other is activation of positive regulators (Tena et al. 2001). LeMPK3 isolated from Lycopersicon peruvianum is homologous to AtMPK3 and is activated by UV-B radiations (Holley et al. 2003). Mayrose et al. (2004) showed that LeMPK3 is involved in mechanical stress and wounding in tomatoes. CbMAPK3 isolated from Chorispora bungeana shows identity of MPK3 and is activated by cold, salt, and ABA. H2O2 activated a novel MAPKKK OMTK1 in alfalfa which activates the MMK3 pathway. In tobacco, the overexpression of NtMEK2 stimulates the gene expression for defense and the generation of reactive oxygen species, which are led by the stimulation of two endogenous MAPKs, SIPK and WIPK (Yang et al. 2001; Ren et al. 2002). In rice system, Rao et al. (2011) showed activation of OsMPK3 and OsMPK4 by arsenic stress. While OsMPK3 was activated only in leaves, both OsMPK3 and MPK4 showed activation in roots. Recently, MPK3 from rice and pea were reported to function as effector molecules of the stress-regulated beta subunit of pea heterotrimeric G-proteins (Bhardwaj et al. 2011).

H2O2 induces 2MAPK-like activities in Arabidopsis and are independent of ethylene and jasmonic acid (Grant et al. 2000). It is not clear that these MAPKs belong to AtMPK3 and AtMPK6. Jonak et al. (1999) have reported that AtMPK3 and AtMPK6 have similarity to wound and SA-induced protein kinases (WIPK and SIPK) of tobacco, respectively. It was also found that only SIPK is activated in tobacco by ozone and H2O2.

4 Nitric Oxide

Nitric oxide (NO) is an important endogenous plant signaling molecule that is responsible for several developmental and physiological processes (Neill et al. 2003, 2008; Tuteja et al. 2004; Delledonne 2005; Lamotte et al. 2005; Erdei and Kolbert 2008; Molassiotis et al. 2010). Extensive work on mammalian system has revealed that NO is a crucial signaling molecule in animals. The physiological functions in plants that are influenced by NO include reduction in seed dormancy (Libourel et al. 2006; Bethke et al. 2007), plant growth regulation and senescence (Mishina et al. 2007), floral transition suppression (He et al. 2004), stomatal movements (Bright et al. 2006; Garcia-Mata and Lamattina 2007), and tolerance to abiotic and biotic stress responses (Uchida et al. 2002; Modolo et al. 2005; Zhao et al. 2007; Floryszak-Wieczorek et al. 2007; Molassiotis et al. 2010). It has also been reported that externally applied NO or NO donors enhance plant tolerance to environmental stresses (Uchida et al. 2002; Garcia-Mata and Lamattina 2007; Zhao et al. 2007a, b).

4.1 Biosynthesis of NO in Plants

In animals, NO is synthesized via a pathway where l-citrulline is formed from l-arginine with the help of nitric oxide synthase (NOS) (Fig. 14.6). The electron donor in this reaction is NADPH and cofactors FMN, FADH, and tetrahydropterin are also used in this reaction. For the biosynthesis of NO in plants, two enzymes NOS and nitrate reductase (NR) are involved (Crawford 2006). There may be other sources for NO biosynthesis apart from these two enzymes (Arnaud et al. 2006). In higher plants, the genes responsible for the upregulation of NOS proteins are yet to be identified. A unique plant NOS (AtNOS1) was isolated from Arabidopsis which encodes a protein associated with NO synthesis (Guo et al. 2003). Overexpression of AtNOS1 increases NO synthesis in E. coli. Reduced root growth and guard cell NO synthesis has been found in the Atnos1 mutant of Arabidopsis plant in response to ABA (Guo et al. 2003). Low amount of NO accumulation in Atnos1 mutants was also reported by many workers (Zhao et al. 2007b; Bright et al. 2006; Zottini et al. 2007). Recently, AtNOS1 was found not to have NOS activity and was not required for the normal synthesis of NO, and it now appears that it is not in fact an NOS at all (Crawford et al. 2006; Zemojtel et al. 2006). AtNOS1 in the biosynthesis of NO is either indirect or regulatory and thus AtNOS1 was renamed as Arabidopsis thaliana nitric oxide-associated 1 (ATNOA1).

Fig. 14.6
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Synthesis of NO from l-arginine catalyzed by nitric oxide synthase

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Another enzymatic source of NO generation is NAD(P)H-nitrate reductase (NR) that converts nitrite to nitric oxide (NO) (Yamasaki 2000; Rockel et al. 2002). The preliminary function of NR in plants is nitrogen assimilation, but in an NAD(P)H-dependent reaction NR can also convert nitrite to NO (Neill et al. 2003; Bright et al. 2006; Crawford 2006). Generation of NO by NR activity has been reported in different plants by different workers, e.g., cucumber (Haba et al. 2001), sunflower, maize (Rockel et al. 2002), and wheat (Xu and Zhao 2003). NR is encoded by two genes NIA1 and NIA2 in Arabidopsis. Double mutants (nia1nia2) deficient in NIA genes showed low NR activity and low NO production in guard cells while their stomata do not close in response to ABA or nitrite (Desikan et al. 2002). Modolo et al. (2006) have reported that due to the lack of NR activity, the nia1nia2 double mutants showed reduced levels of l-arginine and the exogenous l-arginine can restore NO generation in this mutant. Reduction in NOS-mediated NO production may be due to reduced levels of arginine. It is still to be identified how cooperation of two pathways of NO generation controls production of NO in plants.

Another source of NO is a plasma membrane-bound root-specific enzyme, nitrite:NO oxidoreductase (Ni-NOR). The electron donor in this enzyme is not NAD(P)H but cytochrome c and its optimum pH is more acidic than that of NR. The physiological role and genetic identity of this enzyme are not clear yet (Stohr and Stremlau 2006).

4.2 NO Signal Transduction

NO signaling involves cyclic GMP (cGMP)-dependent and -independent pathways such as protein nitrosylation. Pfeiffer et al. (1994) demonstrated that NO stimulated cGMP formation in spruce needles. It is also reported that in tobacco cGMP synthesis is required for NO signaling (Durner et al. 1998). NO activates the cGMP-dependent pathway leading to adventitious root formation in cucumber (Pagnussat et al. 2003). NO increases the cGMP content and inhibitors of cGMP synthesis through guanylate cyclase. It has also been observed that NO induces PCD in Arabidopsis and cGMP synthesis is also involved there. cGMP was suggested to be the likely target of NO signaling in guard cells. The inhibitor of cGMP synthesis ODQ attenuated ABA and NO-induced stomatal closure. Clarke et al. (2000) reported that the addition of cell-permeable cGMP analogue, 8-bromo cGMP relieved this inhibition. However, 8-bromo cGMP alone did not promote closing of stomata, suggesting that synthesis of cGMP is involved but is insufficient for stomotal closure (Neill et al. 2002). Desikan et al. (2004) reported that stomatal closure by H2O2 was not suppressed by ODQ which indicates that H2O2 and NO are different signaling pathways in terms of cGMP signaling. Neither guanylate cyclase nor a cGMP-dependent protein kinase has yet been isolated and cloned from plants (Neill et al. 2003). NO activates intracellular Ca2+ channels through cGMP/cADPR-dependent signaling pathway. Besson-Bard et al. (2008) demonstrated that calcium and MAP kinase can mediate NO signaling. K/Na ratios in Populus eupharatica have been regulated by NO and H2O2 (Zhang et al. 2007). Activation of MAP kinases by NO has been reported in Arabidopsis (Clarke et al. 2000) and tobacco (Kumar and Klessig 2000). Other signaling molecules like H2O2 (Samuel et al. 2000) and SA (Kumar and Klessig 2000) have been reported to activate MAPK in tobacco which indicates that MAPK cascades should be a focal point of convergence of both H2O2 and NO signaling pathway activated in response to various stresses.

5 Sugar as Signaling Molecule

In plants, the biomolecules are very much essential for plant growth and development. Among biomolecules, sugars serve as energy source and as structural component in plants. Koch (2004) reported that 80% of the CO2 assimilated during photosynthesis is used for synthesis of sucrose. Sucrose is the major transport form of organic carbon exported from the photosynthetic source to sink organs. Extensive losses in agricultural production have been observed due to environmental stresses (Bohnert et al. 2006; Mittler 2006). Sucrose is performing dual functions, one as transported carbohydrate in vascular part and the other as signaling molecule (Baier et al. 2004). Sugars are responsible for gene expression involved in plant metabolism viz. photosynthesis, glycolysis, N2 metabolism, cell cycle regulation, etc. Soluble sugars like hormones can act as primary messengers and regulate signals that control the expression of different genes involved in plant growth and metabolism (Rolland et al. 2006; Chen 2007). Gupta and Kaur (2005) have reported that HXK is a sugar sensor in plants. Evolutionary conserved glucose sensor hexokinase 1 (HXK1) mixes different signals (nutrient and hormone signals) and use them for the expression of genes and plant growth in response to environmental stress (Cho et al. 2006). It is not fully understood how the HXK1 controls the gene expression for encoding proteins involved in photosynthesis. Rolland et al. (2006) reported two glucose signal transduction pathways in plants: hexokinase (HXK)-dependent and -independent. HXK-dependent system requires the phosphorylation of glucose while HXK-independent system do not need (Smeekens 2000). Recently, expression of specific photosynthetic genes by HXK1 nuclear complex has been observed and the process is glucose metabolism independent and requires two partners VHAB1 and RPT5B (Chen 2007). This leads to the conclusion that enzymes involved in metabolic activities can play a part in signal transduction by expression of genes in the nucleus. Arabidopsis thaliana mutant hsr8 (high sugar response 8) exhibited increased sugar-responsive growth and expression of genes (Li et al. 2007). Li et al. (2007) observed that hsr8 plants grown under light showed lower chlorophyll content and higher levels of starch and anthocyanin in response to glucose treatment. The hsr8 plant grown under dark showed glucose hypersensitivity. HXK transgenic Arabidopsis plants (AtHXK) have three distinct glucose signal transduction pathways. First is AtHXK1-dependent pathway, where the expression of gene was correlated with the AtHXK1-mediated signaling functions. The second pathway is glycolysis dependent and is regulated by catalytic activity of both AtHXK1 and the heterologous yeast HXK2. The third is AtHXK1-independent pathway, where the expression of gene was independent of AtHXK1 (Xiao et al. 2000). Hence, the role of HXK in sensing the sugar status is still under discussion (reviewed by Rosa et al. 2009).

Kempa et al. (2007) reported that MsK4 (Medicago sativa glycogen synthase kinase 3-like kinases) have been involved in stress signaling with carbon metabolism. MsK4 is located in plastids and is associated with starch granules. Kempa et al. (2007) have also reported that kinase activity of MsK4 is induced rapidly under high salt concentration. Overexpression of MsK4 in transgenic plants showed tolerance to salinity stress accompanied with more starch accumulation and modified carbohydrate content (Kempa et al. 2007). The protein kinases KIN10 and KIN11 connect abiotic stress signals, sugar signals, and developmental signals to regulate plant metabolism (Baena-Gonzalez et al. 2007).

Sucrose nonfermenting-1 (SNF-1)-related proteins, analogues of the protein kinase (SNF-1) yeast signaling pathway, have been reported in plants (Loreti et al. 2001). SNF-1-related proteins have a role in sugar sensing (Purcell et al. 1998).

6 Abscisic Acid in Signaling

Ohkuma et al. in 1963 for the first time purified and crystallized abscisic acid (ABA) from cotton fruits and named it as Abscisin II. It was also isolated from sycamore leaves and was named as dormin. Chemical characterization of both revealed identical nature of the two compounds and it was later named as abscisic acid. Apart from being having an inhibitory role in the plant system, the hormone is also known to possess a stress-protective function. ABA plays an important role in plant responses to drought and salt stresses (Tuteja 2007b; Bansal et al. 2011).

ABA has been shown to regulate many agronomical aspects of plant development like synthesis of proteins and lipids, seed desiccation tolerance and dormancy, and germinative, vegetative, and reproductive growth (Leung and Giraudat 1998; Rock 2000; Rohde et al. 2000). In addition, it mediates the responses of plants to many abiotic stresses like drought, salt, cold stress, and biotic stress like pathogen (Leung and Giraudat 1998; Rock 2000; Rohde et al. 2000; Shinozaki and Yamaguchi-Shinozaki 2000). This implies that ABA is involved in both long-term developmental processes as well as short-term physiological effects. Long-term processes involve changes in pattern of gene expression whereas short-term responses involve changes in the activity of various signaling molecules and fluxes of ion channels across the membranes. Both set of responses require the action of signaling elements which amplify the primary signal generated when the hormone binds to its receptors.

Signaling through ABA causes the production and accumulation of second messengers like Ca2+, phosphatidic acid (PA), or reactive oxygen species (ROS) in the cell which play an important role in ABA signal transduction (Bansal et al. 2011). Reversible protein phosphorylation is an early and central mechanism that occurs in ABA signal transduction (Leung et al. 1997; Himmelbach et al. 2003; Sokolovski et al. 2005). This mechanism involves several protein kinases and phosphatases (Leung and Giraudat 1998; Finkelstein et al. 2002). For example, ABA-activated serine–threonine protein kinase (AAPK) which is a guard cell-specific protein kinase in Vicia faba or orthologous OPEN STOMATA 1/SNF1-RELATED PROTEIN KINASE 2.6 (OST1/SnRK2.6) which regulates ABA-stimulated closure of stomata (Li et al. 2000; Mustilli et al. 2002). Fujii et al. (2007) reported other SnRK2, SnRK2.2, and SnRK2.3 that regulate ABA response in germination, growth, and gene expression. PKABA1 is another protein kinase involved in suppressing the gibberellin (GA)-inducible gene expression in barlet aleurone layers (Gomez-Cadenas et al. 1999). Apart from the above-mentioned calcium-independent protein kinases, the role of several calcium-independent protein kinases has also been revealed in ABA signaling. These either belong to the CDPK or to the SnRK3 family. CDPK1 and CDPK1a have been reported to activate ABA-dependent promoters (Sheen 1996). Also, CDPK3 and CDPK6 possess a role in controlling the aperture of guard cells under abiotic stress (Mori et al. 2006). Zhu et al. (2007) reported the involvement of CPK4 and CPK11 which positively regulate ABA signal transduction in seed germination, seedling growth, and stomatal movements. The mechanism involved is probably the phosphorylation of abscisic acid responsive element-binding factor 1 (ABF1) and abscisic acid responsive element-binding factor 4 (ABF4). Finally, several reports confirm the involvement of mitogen-activated protein kinases (MAPKs) in ABA-dependent response to different stresses (Boudsocq and Laurière 2005; Zhang et al. 2006) and germination (Lu et al. 2002). ABA, in case of stress, regulates gene expression in both positive and negative manner (Chandler and Robertson 1994). Under stress conditions, the gene expression results in the production of transcripts that are responsible for hardening or stress tolerance.

7 Phytohormones as Signaling Molecules

7.1 Brassinosteroids

Brassinosteroids (BRs) are plant steroidal hormones having growth-promoting activities (Hacham et al. 2011). Grove et al. (1979) discovered the brassinolide (BL) (the most active form of BR) from the pollens of Brassica napus. BRs play significant role in seed germination, photomorphogenesis, root and stem elongation, vascular differentiation, senescence, flowering, and resistance to biotic and abiotic stresses (Clouse and Sasse 1998; Divi and Krishna 2010; Divi et al. 2010). The biosynthetic pathway of BRs was elucidated through chemical analysis and isolation of additional BR-biosynthetic mutants, defective in genes encoding proteins which catalyze the plant steroid conversion to BR precursors (Asami et al. 2005). First BR biosynthesis inhibitor, brassinazole, is another powerful tool for elucidation of BR signaling pathway (Asami et al. 2000). Genetic, genomic, and proteomic approaches lead to the establishment of BR signaling pathway by providing an important role in the mechanism of receptor activation and regulating components by process of phosphorylation (Tang et al. 2010).

In Arabidopsis, extensive genetic screens for LOF BR signaling mutants are detected in one locus, BRI1 encoding LRR RLK (Clouse et al. 1996; Kauschmann et al. 1996; Li and Chory 1997; Noguchi et al. 1999). Phenotypes of BRI1 mutants are similar as that of BR-deficient mutants, but these are not rescued by the addition of BRs. Components of BR signaling pathway have been characterized in additional suppressor and gain-of-function screens, which involve second LRR RLK, the BRI1-associated receptor kinase-1 (BAK1) (Li et al. 2002); the glycogen synthase kinase-3 (GSK3)-like kinase, BR insensitive-2 (BIN2) (Li et al. 2001b; Li and Nam 2002), the serine/carboxypeptidase BRI1 suppressor-1 (BRS1) (Li et al. 2001a), the phosphatase BRI1 suppressor-1 (BSU1) (Mora-Garcia et al. 2004), and the transcription factors brassinazole-resistant 1 (BZR1) (Wang et al. 2002) and BZR2 (BRI1-EMS suppressor 1 (BES1)) (Yin et al. 2002).

Recently, BR signaling model has been refined by proteomic studies by identifying the components like BR signaling kinases (BSKs), which are not found in previous screens, generating a complete signaling pathway from an RLK to transcription factors in plants (Tang et al. 2008).

BRs regulate the signaling pathway identical to that of classic receptor tyrosine kinases (RTKs) and transform growth factor-β (TGF-β)-mediated signaling in plants (Feng and Derynck 1997; Schlessinger 2000, 2002). In Arabidopsis, genome sequence has more than 600 RLK members (Shiu et al. 2004) leading to identical signaling mechanisms. Plant RLKs and signaling pathways provide activation to signaling networks, which are controlled by plant hormones (Smet et al. 2009).

7.2 Ethylene

Ethylene is a gaseous plant hormone, plays significant role in developmental processes like seed germination, senescence, fruit ripening, root nodulation, leaf abscission, programmed cell death, stress, and pathogen attack (Johnson and Ecker 1998; Bleecker and Kende 2000; Binder et al. 2010). Ethylene has “triple response” effect on plant growth of etiolated dicotyledonous seedlings. This response leads to radial swelling of hypocotyl, inhibition of hypocotyls, and root cell elongation and exaggerated curvature of the apical hook. Genetic screens of Arabidopsis are based on the triple-response phenotype. More than dozen of mutants are divided into three distinct categories. Constitutive triple-response mutants, i.e., ethylene-insensitive overproduction (eto1), eto2, eto3, constitutive triple-response1 (ctr1) and responsive to antagonist1 (ran1)/ctr2; ethylene-insensitive2 (ein2), ein3, ein4, ein6 and tissue-specific ethylene-insensitive mutants, i.e., hookless1 (hls1), ethylene insensitive root1 (eir1) and various auxin-resistant mutants (Johnson and Ecker 1998; Bleecker and Kende 2000; Stepanova and Ecker 2000). Ethylene belongs to the family of membrane-associated receptors, which include ETR1/ETR2, ethylene response Sensor1 (ERS1)/ERS2 and EIN4 in Arabidopsis (Chang et al. 1993; Hua et al. 1995, 1998; Sakai et al. 1998). Ethylene attaches to its receptor by copper transporter RAN1-delivered copper cofactor. Functions of receptor are inactivated by the hormone binding (Hua and Meyerowitz 1998). EIN2, EIN3, EIN5, and EIN6 act downstream of CTR1 and positively regulate the ethylene response. EIN2 acts as an integral membrane protein, EIN3 acts as transcription factor and expression of intermediate target gene like ethylene response factor1 is regulated.

Ethylene belongs to the family of five receptors (ETR1, ETR2, ETS1, ERS2, and EIN4) and is divided into two subfamilies on the basis of structural similarities. Type-I subfamily contains ETR1 and ERS1 having amino-terminal ethylene-binding domain (which is also known as sensor domain) and carboxy-terminal histidine (His) kinase domain, whereas type-II subfamily receptors contain ETR2, ERS2, and EIN4, which involve an amino-terminal ethylene binding domain and a degenerate His kinase domain. Receptors of ethylene negatively regulates the ethylene responses (Bleecker and Kende 2000; Chang and Stadler 2001). Dominant ethylene in sensitivity mutations in receptor ETR1 leads to signaling (Schaller and Bleecker 1995). LOF mutants have no ethylene response phenotypes. Recently, LOF mutations were isolated in ERS1 gene (Zhao et al. 2002; Wang et al. 2003) with etr1, etr2, ers2, and ein4 mutants. Double LOF etr1ers1 mutants possess strong constitutive-ethylene response phenotypes (Wang et al. 2003). These phenotypes are present in plants containing strong allele of ran1, which cause loss-of-function of all receptors of ethylene (Woeste and Kieber 2000). ETR possesses His kinase activity in vitro, which is important for receptor function (Gamble et al. 1998). For other aspects of receptor functionality like localization, protein stability or interaction with other factors, His kinase activity is essential.

In the mechanism of ethylene signaling, ethylene perception and signaling occurs at endoplasmic reticulum (Chen et al. 2002; Gao et al. 2003). For ER association, the amino-terminal membrane-spanning sensor domain of ETR1 is essential. ER localization of ETR1 is not affected by the introduction of etr1-1 mutations or BR application. CTR is found at ER (Gao et al. 2003). CTR1 contains an amino-terminal domain and carboxy-terminal kinase domain that is linked with Raf-like mitogen-activated protein kinase (MAPK). CTR1 interacts with His kinase domains of ETR1 and ERS1 (Clark et al. 1998). ER-associated CTR1 level inhibits due to removal of ethylene receptors and distribution of CTR1 and receptor interactions. CTR1–ETR1 interaction depends on two lines of evidences in vivo. Co-purification of ETR1 leads to affinity purification of CTR1 from the Arabidopsis ER-membrane fraction, which describes the ETR1 and CTR1 presence in the protein complex (Gao et al. 2003). Overexpression of amino-terminal domain of CTR1 causes LOFctr1 mutant phenotype. Type-I receptors, i.e., ETR1 and ERS1 play significant role in ethylene signaling. This role is not due to His kinase activity of type-I receptors.

7.3 Jasmonates

Jasmonates (JA) regulate plant growth and development. In the reproductive development of plants, jasmonate signaling plays an important role (Stintzi and Browse 2000; Avanci et al. 2010) by giving protection to plants from abiotic stresses (Traw and Bergelson 2003; Huang et al. 2004; Avanci et al. 2010; Lackman et al. 2011) and from pathogens and insects (Farmer and Ryan 1990; Engelberth et al. 2004; Smith et al. 2009; Ma et al. 2010). In Arabidopsis, three mutants namely jar1, coi1, and jin1, which are defective in JA response and one triple mutant defective in JA biosynthesis (fad3-2fad-72fad8) help in understanding the functioning of JA in plants (Staswick et al. 1992; Feys et al. 1994; Berger et al. 1996; McConn and Browse 1996). Disruption of biosynthetic pathway of JA causes susceptibility of plants to various insects and pathogens (Engelberth et al. 2004; Lewsey et al. 2010); for example, susceptibility of coil to Alternaria brassicicola and Pythium mastophorum (Feys et al. 1994). Oxo-phytodienoic acid, JA-amino acids, and JA-glucosyl are the intermediates of JA biosynthesis which act as the signaling molecule in JA pathway (Staswick et al. 2002).

7.4 Salicylic Acid

Salicylic acid (SA) is a naturally occurring phenolic compound having carboxylic acid group attached to the benzene ring. SA has important role in various aspects of plant development (Hayat et al. 2007, 2010). In mung bean, SA helps in the increase in yield and pod number (Singh and Kaur 1980). It also possesses tuber-inducing capacity in potato (Koda et al. 1992). It has positive influence on productivity and nitrogen content in maize (Singh and Srivastava 1978; Asthana and Srivastava 1978), flowering, and helps in reducing transpiration by regulation of stomata (Khurana and Maheswari 1978; Larque 1979) and alleviation of abiotic stress (Ahmad et al. 2011).

SA signaling has been evaluated in case of plants exposed to abiotic stress. Plant tissues when exposed to abiotic stress release more superoxide anions which further increase the level of hydrogen peroxide (Doke et al. 1994; Ahmad et al. 2010). The increased level of hydrogen peroxide has the ability to stimulate the accumulation of SA (Ahmad et al. 2011). Hence, there is a connection between increase in H2O2 level and SA accumulation (Rao et al. 1997). Role of SA has also been described by many workers during cold tolerance in plants like maize, rice, wheat, cucumber, tomato, banana, pea, and mung bean (Janda et al. 1999; yDing et al. 2002; Kang and Saltveit 2002; Kang et al. 2003; Tasgin et al. 2003, Krantev et al. 2009; Popova et al. 2009; Khan et al. 2010). Joseph et al. (2010) have reviewed the exogenous application of SA ant its protective role in different plants under salt stress. Scott et al. (2004) showed the inhibitory effect of SA on the growth of Arabidopsis exposed to chilling conditions. Under low temperatures, the salicylate is reported to accumulate as free and glucosyl SA. Based on studies on various wild species and mutants in Arabidopsis, it was proposed that SA induces low-temperature growth inhibition. Wang and Li (2006) showed increase in cytoplasmic Ca2+ levels after the pretreatment of grape plants with SA. This increased Ca2+ helps in maintaining the integrity of plasma membrane during the stress conditions. Also, it was shown that SA-treated plants had higher levels of antioxidants like glutathione and ascorbic acid.

7.5 Auxins

Auxin, the dynamic plant hormone, controls the growth and developmental processes by modulating the levels of auxin/indole acetic acid proteins (Mockaitis and Estelle 2008; Iglesias et al. 2011). Exogenous application of auxin to plants causes alteration in the transcription of gene families, changes in the rate of cell division and cell elongation, range of electrophysiological responses, and changes of tissue pattern and differentiation (Berleth and Sachs 2001; Perrot-Rechenmann 2010). Auxin signaling initiates with the interaction of auxin receptors. Auxin is considered as a multifunctional hormone and the signal is transduced through several signaling pathways. Iglesias et al. (2010) reported that auxin signaling participates in the adaptive response against oxidative stress and salinity in Arabidopsis. For wild-type auxin response, a large screen for mutants with changed auxin sensitivity was used to define genes for normal functioning. AXR1, AXR2, AXR3, AXR4, and AXR6 are five different loci and TIR1 is the sixth one (Leyser 2002).

8 Conclusion and Future Perspectives

Plants often experience a variety of changing environmental conditions like light conditions, temperature variations, water and nutrient availability, CO2 levels, etc. These changes often lead to stress which hampers the plant growth and development. Plants recognize and respond differently to different stresses for their survival. Plants can sense the external stimulus and lead to induction of defense mechanism. How plants cope with these demands is the subject of intensive research and in this review we tried to throw light on different signaling molecules during abiotic stress.

Calcium signaling is involved in the regulation of plant cell cycle progression in response to abiotic stress. Our knowledge regarding Ca2+ signaling processes in plants have increased tremendously because of the reverse and forward genetic approaches. Most of this research is focused on Ca2+ signal decoders. The versatility of Ca2+ ion signaling is amazing. Major contributor to this phenomenon is its unequal distribution which leads to rapid Ca2+ oscillations resulting in major concentration changes. Other contributors being highly evolved group of calcium sensors which sense these Ca2+ signatures and transduce them to downstream targets for phosphorylation and transcriptional responses. These sensors and transducers together form intricate signaling networks. Ca2+-regulated transcription factors also have an important role in this information processing. CAMTAs are the ones which are most emerging in this field. So, the important parameters which give specificity to this whole mechanism of sensing and decoding of Ca2+ signatures are brought out as the specific Ca2+-binding affinity, the specific cellular concentration and subcellular localization, and the specific interaction affinities of Ca2+ decoders. Now the important question is how calcium sensor proteins read the codes in calcium signatures and decode them downstream in such a precise manner. In order to answer this there is a need to analyze the structures of these sensors, their structural dynamics, the parameters which lead to association and dissociation of these Ca2+ sensors based on fluorescence resonance energy transfer, and the mathematical modeling approaches. In plants, studies can also be utilized to detect the dynamics of protein interactions and protein complex formation. Additionally, an important but not fully exploited area is the reverse genetics approach for exploring the functioning of Ca2+ sensor proteins. Gene knockouts prepared either via crosses or RNAi approaches can be proved beneficial to untangle the functional redundancy issues within and between Ca2+ sensor families. The mutant complementation analyses are useful to study specific pathways which may get influenced by disrupting the gene encoding Ca2+ sensor. MAPK cascades transduce the environmental and developmental signals into adaptivet and programmed responses. Physiological and developmental processes including stress and hormonal responses, innate immunity, etc. are regulated by MAPK cascades. More research is needed to identify substrates of MAPKs and crosstalk with other signaling molecules.

Chemicals such as NO, ABA, SA, JA, BRs, ethylene, and auxins are reported to respond to various environmental stresses and are reviewed in this chapter. Phytohormones SA, JA, and ET play a central role in plant metabolism. SA is known to play a central role in defense against biotrophic pathogens and systemic acquired resistance. JA plays a central role in defense responses to necrotrophic pathogens and herbivores, whereas ET plays a great role in fruit ripening and senescence and modulates defense responses. Intensive research is needed to know the role of different phytohormones in signaling. The overall progress of research on chemical-regulated stress-responsive genes and their products reflects their central role in plant growth and development under stress conditions.