Polyamines and abiotic stress tolerance in plants

REVIEW

review

Plant Signaling & Behavior 5:1, 26-33; January 2010; © 2010 Landes Bioscience

Polyamines and abiotic stress tolerance in plants Sarvajeet Singh Gill and Narendra Tuteja* Plant Molecular Biology Group; International Centre for Genetic Engineering and Biotechnology; Aruna Asaf Ali Marg, New Delhi, India

Key words: abiotic stress tolerance, putrescine, spermidine, spermine, polyamines Abbreviations: ACC, 1-aminocyclopropane-1-carboxylic acid; ADC, arginine decarboxylase; ADI, agmatine deiminase; APX, ascorbate peroxidise; CPA, N-carbamoylputrescine amidohydrolase; DAO, diamine oxidase; dcSAM, decarboxylated S-adenosylmethionine; GABA, γ-aminobutyric acid; GR, glutathione reductase; MDA, malondialdehyde; MGBG, methyglyoxal-bis(guanylhydrazone); ODC, ornithine decarboxylase; PAs, polyamines; POD, peroxidise; Put, putrescine; SAMDC, S-adenosyl methionine decarboxylase; SMM, S-methylmethionine; SOD, superoxide dismutase, spermidine; SPDS, spermidine synthase; Spm, spermine; SPMS, spermine synthase

Environmental stresses including climate change, especially global warming, are severely affecting plant growth and productivity worldwide. It has been estimated that two-thirds of the yield potential of major crops are routinely lost due to the unfavorable environmental factors. On the other hand, the world population is estimated to reach about 10 billion by 2050, which will witness serious food shortages. Therefore, crops with enhanced vigour and high tolerance to various environmental factors should be developed to feed the increasing world population. Maintaining crop yields under adverse environmental stresses is probably the major challenge facing modern agriculture where polyamines can play important role. Polyamines (PAs)(putrescine, spermidine and spermine) are group of phytohormone-like aliphatic amine natural compounds with aliphatic nitrogen structure and present in almost all living organisms including plants. Evidences showed that polyamines are involved in many physiological processes, such as cell growth and development and respond to stress tolerance to various environmental factors. ın many cases the relationship of plant stress tolerance was noted with the production of conjugated and bound polyamines as well as stimulation of polyamine oxidation. Therefore, genetic manipulation of crop plants with genes encoding enzymes of polyamine biosynthetic pathways may provide better stress tolerance to crop plants. Furthermore, the exogenous application of PAs is also another option for increasing the stress tolerance potential in plants. Here, we have described the synthesis and role of various polyamines in abiotic stress tolerance in plants.

Introduction Plants are sessile and sensitive organisms that encounter a variety of environmental stresses throughout the life cycle. Plant development and productivity are negatively affected by environmental stresses.1-4 During the last decade, cultivated land in several regions of the world has been affected by environmental stresses *Correspondence to: Narendra Tuteja; Email: [email protected] Submitted: 10/06/09; Accepted: 10/07/09 Previously published online: www.landesbioscience.com/journals/psb/article/10291

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like salt, cold, drought and UV, which hinders crop cultivation and yield.5,6 Every year countries lose a substantial amount of money from reductions in crop productivity caused by abiotic stresses.5 It is predicted that these environmental stresses will become more intense and frequent with climate change, especially global warming. On the other hand, the world population is estimated to reach near 10 billion by 2050, which will witness serious food shortages. Therefore, tolerant crops should be developed to feed the increasing world population. Maintaining crop yields under adverse environmental stresses is probably the major challenge facing modern agriculture. Plants possess efficient defense mechanisms to cope with a plethora of environmental stresses, which includes drought, UV, high-salinity, cold stresses and pathogen attack.7 Polyamines (PAs), widely present in living organisms, are now regarded as a new class of growth substances which includes spermidine (Spd, a triamine), spermine (Spm, a tetramine), and their obligate precursor putrescine (Put, a diamine) which play a pivotal role in the regulation of plant developmental and physiological processes.8 PAs may also function as stress messengers in plant responses to different stress signals.9,10 PAs are small ubiquitous polycations involved in many processes of plant growth and development and are well known for their anti-senescence and anti-stress effects due to their acid neutralizing and antioxidant properties, as well as for their membrane and cell wall stabilizing abilities.11 It has been suggested that PAs play important roles in modulating the defense response of plants to diverse environmental stresses,12 which includes metal toxicity,13 oxidative stress,14 drought,15 salinity16 and chilling stress.17,18 It has been reported that exogenous application of PAs is also effective approach for enhancing stress tolerance of crops for enhanced crop productivity. Exogenous application of Put has been successfully used to enhance salinity,19-21 cold,22,23 drought,24 heavy metals,25 osmotic stress,26 high-temperature,27 water logging28 and flooding tolerance of plants.29 Furthermore, it has been noted that genetic transformation with polyamine biosynthetic genes encoding arginine decarboxylase (ADC), ornithine decarboxylase (ODC), S-adenosylmethionine decarboxylase (SAMDC) or Spd synthase (SPDS) improved environmental stress tolerance in various plant species.10 It is interesting

Plant Signaling & Behavior Volume 5 Issue 1

REVIEW

review

Figure 1. Schematic presentation of PA biosynthetic pathway for Put, Spd and Spm in plants and its relationships with ethylene biosynthesis. ADC, Arginine decarboxylase; CPA, N-Carbamoylputrescine amidohydrolase; ADI, Agmatine deiminase; ODC, Ornithine decarboxylase; DAO, Diamine oxidase; SPDS, Spermidine synthase; SPMS, Spermine synthase; SAM DC, SAM decarboxylase; dcSAM, decarboxylated S-adenosylmethionine; ACC, 1-aminocyclopropane-1-carboxylic acid.

to note that transgenic plants overexpressing ADC,30 SPDS31-33 or SAMDC34 can tolerate multiple stresses including salinity, drought, low and high temperature and parquet toxicity. Such multiple abiotic stress tolerance is of practical importance since plants often suffer from several concurrent forms of environmental stress during their life cycle. In this review, we tried to summarize the knowledge that has been gathered over the last couple of decades concerning the changes in polyamine metabolism (biosynthesis, catabolism and regulation) in plants under various environmental stress factors. Attempts has been made to discuss and understand the new advances concerning the mechanisms of polyamine action in plants in response to salinity, drought, freezing and heavy metal stress. Polyamines PAs are known as a group of natural compounds with aliphatic nitrogen structure, present in almost all living organisms, play important roles in many physiological processes, such as cell growth and development and respond to environmental stresses. Put, Spd and Spm are the most commonly found PAs in higher plants and could be present in free, soluble conjugated, and insoluble bound forms.35 Soluble conjugated PAs are covalently conjugated to small molecules such as phenolic compounds, while insoluble bound PAs are covalently bound to macromolecules such as nucleic acids and proteins.16 In addition, uncommon PAs, like homospermidine, 1,3-diaminopropane, cadaverine and canavalmine have been detected in a large number of biological systems, including plants, animals, algae and bacteria. At the physiological pH, PAs are found as cations. This polycationic

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nature of PAs is one of their important properties effectuating their biological activities.36 Large body of evidence suggested that plant transformation with genes of PAs biosynthetic enzymes or the exogenous application of PAs such as Put, Spd and Spm results in abiotic stress tolerance in various plants. Synthesis of Polyamines PAs are a group of phytohormone-like aliphatic amine compounds, major types of which are the, triamine Spd [NH2 (CH2)3NH(CH2)4NH2], tetramine Spm [NH2 (CH2)3NH(CH2)4NH(CH2)3NH2] and their diamine obligate precursor Put [NH2 (CH2)4NH2] that are ubiquitous in all plant cells. The biosynthetic pathways of main PAs such as Put, Spd and Spm are shown in Figure 1. These three main PAs differ in the number of positive charges exhibited in the physiological pH of the cell. The diamine Put synthesis proceeds through either ADC via agmatine (Agm) or ODC, while the triamine Spd is synthesized by SPDS through the addition to Put of an aminopropyl moiety donated by decarboxylated S-adenosylmethionine (dcSAM) formed by SAMDC. Conversion of Agm into Put requires two distinct enzymes: N-carbamoylputrescine amidohydrolase (CPA) and agmatine deiminase (ADI). Spd functions as a substrate for the synthesis of the higher polyamine Spm. Put is catabolized by diamine oxidases (DAOs), in a reaction that converts Put into Δ1-pyrroline and generates ammonia and H2O2 as byproducts (Fig. 1). DAOs are preferentially localized in plant cell walls, and hydrogen peroxide resulting from Put catabolism may be important in lignifications and cross-linking reactions under normal and stress conditions. Following the oxidation of Put, Δ1-pyrroline is catabolized into γ-aminobutyric acid (GABA)

Plant Signaling & Behavior

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(Fig. 1), which is ultimately converted into succinic acid, a component of the Krebs cycle.37 Polyamines and Abiotic Stress Tolerance in Plants PAs are involved in the regulation of many basic cellular processes, including DNA replication, transcription, translation, cell proliferation, modulation of enzyme activities, cellular cationanion balance and membrane stability. It has been illustrated that PAs also play pivotal roles in a wide range of growth and developmental processes such as cell division, dormancy breaking of tubers and germination of seeds, stimulation, support and development of flower buds, embryogenesis, fruit set and growth, fruit ripening, plant morphogenesis and response to biotic and abiotic stresses.12,18 Recently, a large body of study shows that plant PAs are involved in the acquisition of tolerance to such stresses as high and low temperatures, salinity, hyperosmosis, hypoxia and atmospheric pollutants.10,38 Zapata et al.39 studied the effect of salinity on plant growth, ethylene production and polyamine levels in Spinacia oleracea, Lactuca sativa, Cucumis melo, Capsicum annum, Brassica oleraceae, Beta vulgaris and Lycopersicon esculentum. They found that PA levels changed with salinity and in most cases Put decreased while Spd and/or Spm increased. The (Spd + Spm)/ Put ratio increased with salinity in all species increased salinity tolerance. It is shown that there is a general response by different plant species to salinity in relation to PA production, but not with regard to ethylene production. Liu et al.26 reported that PEG 6000 treatment significantly increased the free-Spd and free-Spm levels in leaves of Triticum aestivum drought-tolerant cv. Yumai No. 18. Whereas, Yangmai No. 9 cv. (drought-sensitive) showed a significant increase of free-Put level. Thus, Yumai No. 18 cv. gave a higher ratio of (free-Spd + free-Spm)/free-Put than Yangmai No. 9 cv. in leaves in response to osmotic stress. They suggested that free-Spd, free-Spm and PIS-bound Put facilitated the osmotic stress tolerance of wheat seedlings. Garcıa-Jimenez et al.38 reported that a moderate hyposaline shock caused an increase in the free fraction of Put, Spd and Spm, mainly due to a decrease in TGase activity, together with an apparent increase in the L-arginine dependent PAs synthesis (ODC and arginase decreased, and ADC slightly increased). Lefevre et al.35 studied the importance of ionic and osmotic components of salt stress on modification of free PA level in the seedling of two sensitive (IKP) and tolerant (Pokkali) cultivars of rice isoosmotic concentrations of NaCl or PEG and found that putrescine have differential role in non-photosynthetic organs versus photosynthetic because it accumulated to high amounts in the roots of Pokkali in comparison to IKP, whereas, an opposite trend was recorded in the shoots. They have noted that tyramine was also present at higher concentrations in the roots of Pokkali and its level clearly increased in response to ionic stresses while cadaverine level increased in water stress conditions only. Scaramagli et al.40 investigated the response of Put and Spd and Spm status in Solanum tuberosum cells grown in medium supplemented with 2,4-D and kinetin, and acclimated or not to low water potential. The di- and polyamine (Spd and Spm) status in cells gradually acclimated to increasing concentrations (up to

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20%, w/v) of polyethylene glycol (PEG 8000), was compared with that of unacclimated cells abruptly exposed (shocked) or not (controls) to 20% (w/v) PEG. They found that with respect to di- and polyamines, acquired tolerance to low water potential in potato cells leads principally to changes in Put biosynthesis and conjugation which may be involved in ensuring cell survival. In another study, Urano et al.41 studied the role of putrescine in salt tolerance in Arabidopsis. They found the induction of AtADC2 in response to salt stress causing the accumulation of free Put. Further, to analyze the roles of stress-inducible AtADC2 gene and endogenous Put in stress tolerance, they isolated a Ds insertion mutant of AtADC2 gene (adc2-1) and characterized its phenotypes under salt stress and found that in the adc2-1 mutant, free Put content was reduced to about 25% of that in the control plants and did not increase under salt stress and the adc2-1 mutant was more sensitive to salt stress than the control plants but stress sensitivity of adc2-1 was recovered by the addition of exogenous Put. They concluded that endogenous Put plays an important role in salt tolerance in Arabidopsis and AtADC2 is a key gene for the production of Put under not only salinity conditions, but also normal conditions. Duan et al.16 reported that salinity stress increased the superoxide and hydrogen peroxide production, particularly in Cucumis sativus cv. Jinchun No. 2 roots, while the salinity-induced increase in antioxidant enzyme activities and proline contents in the roots was higher in cv. Changchun mici than in cv. Jinchun No. 2. They have also noted a marked increase in ADO, ODC, SAMDC and DAO activities, as well as free Spd and Spm, soluble conjugated and insoluble bound Put, Spd and Spm contents in the roots Changchun mici than Jinchun No. 2 under salt stress. A substantial increase in Put, Spd and cadaverine contents at low temperature under normal light conditions in winter wheat MvEmese, while in the spring wheat Nadro, only the levels of Spd and Spm increased under these conditions.42 In another study, Garnica et al.43 reported that nitrate application induces changes in PA content and ethylene production in wheat plants grown with ammonium and found that nitrate’s effects on Put, Spd and Spm contents of ammonium-fed plants tended to follow the pattern associated with strict nitrate nutrition. Camacho-Cristobal et al.44 studied the behaviour of PA accumulation in leaves and roots of Nicotiana tabacum under short-term boron deficiency and found a noticeable decrease in plant growth. However, boron deficiency did not lead to a significant decrease in leaf or root ion concentrations when compared to control treatment; however, as expected, leaf boron concentration was lower in boron-deficient plants in comparison to the control. They have found that the levels of free Put and Spd in leaves were similar in both treatments but in roots, a short term boron deficiency caused an increase in free Put concentration. Moreover, boron deficient plants had higher conjugated polyamine concentration than boron-sufficient plants, which was especially evident for conjugated Put in leaves. Racz et al.45 reported that S-methylmethionine (SMM) treatment reduced cell membrane damage, and thus the electrolyte leakage under low-temperature stress in the leaves and roots of peas, maize, soy beans and eight winter wheat varieties with different levels of frost resistance. The interaction between SMM and PA


biosynthesis revealed that SMM increased the quantities of Agm and Put as well as that of Spd, while it had no effect on the quantity of Spm. Furthermore, by using a specific inhibitor, methylglyoxal-bis-guanyl hydrazone, it was noted that the PA metabolic pathway starting from methionine played no role in the synthesis of Spd or Spm, so there is a alternative pathway for the synthesis of SMM-induced PAs. Sood and Sweta46 studied the changes in endogenous PAs levels Polianthes tuberosa bulbs during the dormant periods and found that high free Put and low Spm and Spd levels were associated during initial stages of dormancy but high Spm and Spd levels were related with dormancy release. The conjugated Put level increased during the period with an increase in conjugated Spm and Spd levels. However, an inverse relationship between free and conjugated PAs was noticed only for Put. Ioannidis et al.47 reported that Put can increase light energy utilization through stimulation of photophosphorylation. Recently, Ioannidis and Kotzabasis48 reported that Put is an efficient stimulator of ATP synthesis in comparison to Spd and Spm in terms of maximal % stimulation but Spd and Spm are efficient stimulators of non-photochemical quenching. They found that Spd and Spm at high concentrations are efficient uncouplers of photophosphorylation. Furthermore, the higher the polycationic character of the amine being used, the higher was the effectiveness in PSII efficiency restoration, as well as stacking of low salt thylakoids. Spm with 50 µM increase F V as efficiently as 100 µM of Spd or 1,000 µM of Put or 1,000 µM of Mg2+. It is also demonstrated that the increase in F V derives mainly from the contribution of PSIIα centers. Their findings underline the importance of chloroplastic PAs in the functionality of the photosynthetic membrane.48 Sfakianaki et al.49 studied the response of low temperature on the structure and function of the photosynthetic apparatus in Phaseolus vulgaris and found that the photosynthetic apparatus is affected by the changes occurring in the pattern of LHCII-associated Put and Spm which adjust the size of LHCII. They reported that the decrease of Put/Spm ratio was mainly due to the reduction in the quantity of LHCII-associated Put that led to an increase of the LHCII. These alterations in the structure of the photosynthetic apparatus combined with the reduction in the photosynthetic electron transfer rate actually resulted in the inactivation of active reaction centers and the increase of dissipated energy which diminished the photosynthetic efficiency and the maximal photosynthetic rate. Furthermore, it was found that the photosynthetic mechanism recovered quite fast to the initial condition when plants were transferred to 26°C after low temperature stress. Wei et al.50 studied the effect of excess calcium nitrate (Ca(NO3)) on PA contents in leaves of grafted and nongrafted Solanum melongena seedlings, in which grafted plants were grafted on a salinity tolerant Solanum torvum. They noted that the contents of free, soluble conjugated and insoluble bound PAs of grafted seedlings were significantly higher than those of non-grafted seedlings, and activities of DAO and PAO of grafted seedlings were significantly lower than those of non-grafted seedlings. Furthermore, the activities of superoxide dismutase (SOD), peroxidise (POD), ascorbate peroxidise (APX) and glutathione reductase (GR) of grafted seedlings were significantly higher than those of non-grafted seedlings. They reported the synergy


between antioxidant enzymes and PAs in protective mechanism of grafted eggplant seedlings to excess Ca(NO3) stress. Rodrıguez-Kessler et al.51 analyzed the alterations occurred in PA metabolism of maize tumors formed during the interaction with the biotrophic pathogenic fungus Ustilago maydis and found a striking increase in maize polyamine biosynthesis, mainly free and conjugated Put occurred in the tumors induced by the fungus, and in the neighbor plant tissues. They correlated this increase with activation mainly of Adc, Samdc1, Zmsamdc2 and Zmsamdc3, but not of Zmodc, Zmspds1 and Zmspds2 genes, and an elevation in ADC activity, confirming a predominant role of this enzyme in the process. Evidences for a possible contribution of Spd and Spm degradation by PAO activity, probably related to cell wall stiffening or lignification during tumor growth, were also obtained. It is concluded that PAs, mainly Put, might play an active role in the pathosystem maize U. maydis. Transgenic Plants Engineered to Synthesize PAs for Enhanced Abiotic Stress Tolerance During the last few years several genes encoding biosynthetic enzymes of PA pathway such as ADC, ODC, SAMDC or SPDS improved environmental stress tolerance in various plant species10,52 (Table 1) and allowing the molecular approach to become a useful tool for gaining new insights on the regulation of PA metabolism. To understand the role of SPDS in the PA metabolism and in particular in affecting Spd endogenous levels, Franceschettia et al.53 reported that Nicotiana tabacum overexpressing Datura stramonium spermidine synthase showed all transgenic clones displayed a high level of overexpression of the exogenous cDNA with respect to the endogenous SPDS and no relationship was detected between the mRNA expression level of SAMDC, which did not change between the negative segregant control and the transgenic plants, and SPDS, suggesting the existence of an independent regulatory mechanism for transcription of the two genes. They found that ODC was the most active enzyme and its activity was equally distributed between the soluble and the particulate fraction, while ADC activity in the transgenic plants did not particularly change with respect to the controls. Furthermore, the transformed plants displayed an increased Spd to Put ratio in the majority of the clones assayed, while the total PA content remained almost unchanged in comparison to the controls. Their findings suggest a high capacity of the transformed plants to tightly regulate PA endogenous levels and provide evidence that SPDS is not a limiting step in the biosynthesis of PAs.53 Torrigiani et al.54 conducted an experiment to understand the regulation of PAs (spd and spm) biosynthesis. They introduced SAMDC cDNA of Datura stramonium into Nicotiana tabacum in antisense orientation and found that transgenics efficiently transcribed the antisense SAMDC gene, but SAMDC activity and PA titres did not change. Contrarily, SAMDC activity was remarkably lower in most transgenics than in controls, and the Put-to-Spd ratio was altered, mainly due to increased Put, even though Put oxidising activity (DAO) did not change relative to controls. Despite the reduction in SAMDC activity, the production of


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Table 1. Transgenic plants engineered to synthesize PAs (Put, Spd and Spm) for enhanced abiotic stress tolerance (modified and updated from plantstress.com) Gene ADC (Arginine decarboxylase) ADC1, ADC2 (mutants defective in putrescine biosynthesis)

SAMDC (S-adenosyl methionine decarboxylase)

Function & Gene product ADC is responsible for the biosynthesis of diamine Put from arginine

SAMDC is a key enzyme involved in the biosynthesis of the PAs, viz. Spd & Spm

MdSAMDC2

MdSPDS1 (Spermidine Synthase)

SPDS converts Put into spermidine

Source

Transgenic Plant

References

Avena sativa

Oryza sativa L.

Salt tolerance

66

Datura stramonium

Oryza sativa

Drought tolerance (PEG8000)

67

Arabidopsis mutants

Arabidopsis t haliana

Freezing tolerance and cold acclimation

17

Tritordeum

Oryza sativa

Salt tolerance

68

Human

Nicotiana tabacum var. xanthi

Salinity, drought and fungal wilts (caused by Verticillium dahliae and Fusarium oxysporum) stress tolerance

69

Saccharomyces cerevisiae

Lycopersicon esculentum

High temperature stress

70

Malus sylvestris

Malus sylvestris

Cold and Salt stress

71

Malus sylvestris

Pyrus communis L. ‘Ballad’

Salt tolerance

56

Malus sylvestris

Pyrus communis L. ‘Ballad’

Salt, osmotic and heavy metal stress tolerance

33

Cucurbita ficifolia

Arabidopsis t haliana L.

Chilling, freezing, salinity, hyperosmosis, drought and paraquat stress tolerance

72

ethylene, which shares with PAs the common precursor SAM, was not influenced by the foreign gene. Mohapatra et al.55 compared the activities of several enzymes and cellular metabolites involved in the ROS scavenging pathways in two isogenic cell lines of poplar (Populus nigra x maximowiczii) differing in their PA contents. Whereas, the control cell line was transformed with β-glucuronidase (GUS), the other, called HP (High Putrescine), was transformed with a mouse ornithine decarboxylase (mODC) gene. The expression of mODC resulted in several-fold increased production of Put as well its enhanced catabolism. The two cell lines followed a similar trend of growth over the seven-day culture cycle, but the HP cells had elevated levels of soluble proteins. Accumulation of H2O2 was higher in the HP cells than the control cells, and so were the activities of GR and MDHAR; the activity of APX was lower in the former. The contents of GSH and glutamate were significantly lower in the HP cells but proline was higher on some days of analysis. There was a small difference in mitochondrial activity between the two cell lines, and the HP cells showed increased membrane damage. In the HP cells, increased accumulation of Ca was concomitant with lower accumulation of K. We conclude that, while increased Put accumulation may have a protective role against ROS in plants, enhanced turnover of Put actually can make them vulnerable to increased oxidative damage. He et al.56 (2008) reported changes in enzymatic and non-enzymatic antioxidant capacity of the SPDS-overexpressing transgenic Pyrus communis in response to NaCl or mannitol stress. They found that before stress treatment the Spd contents and SPDS activity were higher than wild type. They reported that under salt or mannitol stress both the

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Enhanced tolerance

wild type and transgenic exhibited accumulation of Spd with the latter accumulating more. The transgenic line contained higher antioxidant enzyme activities, less MDA (malondialdehyde) and hydrogen peroxide (H2O2) than the wild type, implying it suffered from less injury. These results suggested that increase of Spd content in the transgenic line could, at least in part, lead to enhancing enzymatic and non-enzymatic antioxidant capacity. Exogenous Application of Polyamines for Enhanced Abiotic Stress Tolerance Large number of evidence suggested that exogenous application of PAs (di- and tri- and tetra-amines) were shown to stabilize plant cell membranes, protecting them from damage under stress conditions10,56 and endogenous PAs are also suggested to participate in sustaining membrane integrity.58 Yiu et al.29 reported that exogenous Put reduces the flooding-induced oxidative damage in Allium fistulosum by increasing the antioxidant capacity. They found that exogenous application of Put resulted in reduced superoxide radical (O2•-) and H2O2 contents and thereby, less oxidative stress in plant cells. The antioxidant system, as an important component of the water logging-stress-protective mechanism including α,α-diphenyl-β-picrylhydrazyl (DPPH)-radical scavenging activity, O2•- scavenging, metal chelating activities and reducing power, was upgraded by Put, which is therefore able to moderate the radical scavenging system and to lesser oxidative stress. Their findings suggest that Put confers flooding tolerance to Allium fistulosum, probably through inducing the activities of various anti-oxidative systems. Ali59 reported that exogenous


application of Put reduced the net accumulation of Na+ and Clions in different organs of Atropa belladonna subjected to salinity stress. Put alleviated the adverse effect of NaCl during germination and early seedling growth and increased the alkaloids as well as endogenous Put of A. belladonna. Lutts et al.60 reported that Put increased the growth and the leaf tissue viability of salttreated plants in all cvs. of Oryza sativa. They suggested that this positive effect was associated with an increase in ethylene biosynthesis through an increase in ACC content and a suppression of NaCl-induced inhibition of ACC conversion to ethylene and suggested the involvement of Put in salinity tolerance in rice. Ndayiragije and Lutts61 studied the possible role of exogenous application of PAs on Oryza sativa and noted that addition of PAs in nutritive solution reduced plant growth in the absence of NaCl and did not afford protection in the presence of NaCl. PA-treated plants exhibited a higher K+/Na+ ratio in the shoots, suggesting an improved discrimination among monovalent cations at the root level, especially at the sites of xylem loading. Put induced a decrease in the shoot water content in the presence of NaCl, while Spd and Spm had no effects on the plant water status. In contrast to Spd, Spm was efficiently translocated to the shoots. Both PAs (Spd and Spm) induced a decrease in cell membrane stability as suggested by a strong increase in MDA content of PA-treated plants exposed to NaCl. Zhang et al.62 reported the protective effect of exogenous PAs application on Changchun mici (chilling-resistant) and Beijing jietou (chilling-sensitive) cv. of cucumber against chilling injury. They noted a remarkable increase in free Spd, Spm and Put in the leaves of cv. Changchun mici upon chilling treatment after 1 day of treatment but the induction of Put declined thereafter, whereas, Spd and Spm levels increased steadily. Whereas, in the leaves of cv. Beijing jietou, Put content was increased only at 1 day after chilling while Spd content decreased significantly upon chilling treatment. It was noted that chilling reduced the soluble protein content, activities of antioxidant enzymes (SOD, POD, CAT and APX) only in Beijing jietou. However, these changes could be renovated by exogenous application of Put and Spd. It was also found that pretreatment with Put and Spd diminished the increased electrolyte leakage and MDA content caused by chilling in the leaves of both cultivars. Pretreatment of methyglyoxal-bis-(guanylhydrazone) (MGBG), the PAs biosynthetic inhibitor cancelled the effects of PAs in most of the treatments. Furthermore, Changchun mici was found to contain higher endogenous free PAs contents compared to Beijing jietou. They concluded that PAs play important roles in the tolerance of cucumber against chilling stress, which is most likely achieved by acting as oxidative machinery against chilling injury. Verma and Mishra 20 also reported that PAs (Put) reversed the salinity induced reductions in seedling growth and biomass accumulation and increased O2•-, H2O2 levels, MDA content and electrolyte leakage in leaf tissues of Brassica juncea. Put also increased the activity of antioxidant enzymes and carotenoids in leaf tissues of salt stressed Brassica juncea seedlings. These finding suggests that Put might be activating antioxidant enzymes and elevating antioxidants there by controlling free radical generation, hence preventing membrane peroxidation and denaturation


of biomolecules resulting into improved seedling growth under salinity. Whereas, Liu et al.26 reported that exogenous application of Spd resulted in an increase of free-Spd + free-Spm in leaves of, in concert with an alleviation of PEG-induced injury to cv. angmai No. 9 (drought sensitive) in comparison to Yumai No. 18 (drought tolerant) Triticum aestivum seedlings. Huang and Bie 63 reported that 0.1 mM cinnamic acid (CA) treatment increased the Put level, but decreased Spd and Spm levels, thereby reducing the (Spd + Spm)/Put ratio in the leaves of Vigna unguiculata. The exogenous application of 1 mM Spd markedly reversed these CA-induced effects for PA and partially restored the PAs ratio and RuBPC activity in leaves. Methylglyoxal-bis (guanylhydrazone) (MGBG), which is an inhibitor of SAMDC, results in the inability of activated cells to synthesize Spd and exacerbates the negative effects induced by CA. Furthermore, the exogenous application of 1 mM D-arginine (D-Arg), which is an inhibitor of Put biosynthesis, decreased the levels of Put, but increased the PAs ratio and RuBPC activity in leaves. These results suggest that 0.1 mM CA inhibits RuBPC activity by decreasing the levels of endogenous free and perchloric acid soluble (PS) conjugated Spm, as well as the PAs ratio.63 Lazzarato et al.64 reported that the Spd and arsenic increased resistance responses in the well-known pathosystem NN tobacco/tobacco mosaic virus. They found that exogenous application of Spd, arsenic and DL -β-aminobutyric acid modulate the tobacco resistance to TMV, and also affect the local and systemic glucosylsalicylic acid levels and ADC gene expression in tobacco leaves. Shoeb et al.65 found that modification of cellular PA titers and their Put:Spd ratio by the addition of exogenous PAs (Put, Spd) or their biosynthesis inhibitor, difluoromethylarginine led to the induction/promotion of plant regeneration in poorly responding genotypes like HS, Bindli, DV-85, ACB-72, IR-64 and IR-72. They found a close relationship between cellular PA levels and their Put:Spd ratio with in vitro morphogenetic capacity in indica rice and suggest that the cellular PAs and Put:Spd ratios are important determinants (biomarkers) of plant regeneration ability in indica rice, and the improvement/induction of plant regeneration in morphogenetically poor and recalcitrant species could be achieved by modulating PA metabolism. Conclusion and Future Prospects Various abiotic stresses including the global warming are negatively affecting the plant productivity worldwide. On the other hand the demand for food is expected to grow as a result of population growth and rising incomes. According to FAO, the World needs 70% more food by the year 2050. Therefore, it is necessary to obtain stress-tolerant varieties to cope with this upcoming problem of food security. The involvement of PAs in regulation of various cellular processes including growth, development and stress tolerance in plants might have general implications. However, in plants the role of PAs metabolism for the abiotic stress tolerance is just beginning to be understood. A lot of effort is still required to uncover in detail the molecular mechanism of protective role of Spd, Spm and Put in abiotic


31

stress tolerance. The knowledge gained so far about plant PAs has built a strong case for further studies towards careful analysis of the genes involved in abiotic stress tolerance. High throughput analysis including microarray, transcriptomics, metabolomics, reverse genetics approaches will also be helpful to understand the involvement of PAs biosynthetic pathways in abiotic stress tolerance. Further insights are also expected from three-dimensional structural studies of individual components of PAs biosynthetic pathways. Isolation and analysis of interacting partners of the enzymes of PAs biosynthetic pathways will also be of great help in our better understanding of the mechanism of stress tolerance. The pyramiding of the genes encoding the enzymes of PAs References 1. 2.

3.

4.

5.

6. 7.

8.

9.

10.

11.

12.

13.

14.

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biosynthetic pathways will also be helpful for further enhancing the tolerance potentials of crop plants for various stress factors. Furthermore, the use of PAs as external application can also be exploited for increasing tolerance to salinity, cold, drought, heavy metal, osmotic stress, high-temperature, water logging and flooding tolerance in various crop plants. Acknowledgements

Work on plant abiotic stress tolerance in NT’s laboratory is partially supported by Department of Science and Technology (DST), Government of India and Department of Biotechnology (DBT), Government of India.

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