Improvment of cold tolerance of chickpea through heavy polyamines catabolism and ethylene phytohormone

Document Type : Research Paper


1 Assistant Professor, Seed and Plant Certification and Registration Research Institute (SPCRI), Agricultural Research, Education and Extension Organization (AREEO), Karaj, Iran

2 Faculty member of Agronomy and Plant Breeding Department, College of Agriculture and Natural Resources, University of Tehran, Karaj,


The current study was undertaken to investigate if there is a relationship between metabolism of ethylene and heavy polyamines (PAs) under cold stress in cold-tolerant and cold-sensitive chickpea (Cicer arietinum L.) genotypes.
Materials and methods
In this research, content of ethylene, heavy polyamines (spermidine (Spd) and spermine (Spm)), activities of PAs degradation pathway enzymes (polyamine oxidase (PAO) and diamine oxidase (DAO)), hydrogen peroxide (H2O2) and relative expression of 1-aminocyclopropane-1-carboxylic acid synthase (ACS) and 1-aminocyclopropane 1-carboxylic acid oxidase (ACO) genes in cold-tolerant (Sel 96th11439) and cold-sensitive (ILC 533) chickpea (Cicer arietinum L.) genotypes during the first and sixth days of cold stress at 4 °C compared to control condition as a factorial experiment in a Completely Randomized Design were investigated.
During cold stress, both genotypes showed a significant increase in Spd and Spm content (66.66 and 96.23%). Ethylene production was declined in cold-sensitive genotype under cold stress compared to control conditions (up to 26.08%) while in the cold-tolerant genotype, the unique ethylene peak in early response (on the first day of stress) comared to control conditions (15.62%) was closely related to increased heavy polyamine accumulation. In the tolerant genotype, the increase in polyamine oxidase (PAO) and diamine oxidase (DAO) activity in early responses, (By 2.6- and 3.01-fold, respectively) was related to the increase in ethylene biosynthesis, as well as a concomitant increase in heavy polyamine (Spd & Spm) content by cold stress. In the tolerant genotype, the relative expression of ACS and ACO genes, after a significant increase on the first day of cold stress (5.2- and 4.03-fold, respectively), showed a significant decrease on the sixth day of the stress compared to the control plants, while a continuous decreasing trend (35-and 21.7-fold, respectively) was observed in the sensitive genotype compared to the control condition.
Findings of this research suggest that ethylene is intimately involved in improvement of cold stress tolerance through activation of a complex pathway of signalling by H2O2 that is polyamine catabolism-dependent.


Abts W, Van de Poel, B, Vandenbussche B, De Proft MP (2014) Ethylene is differentially regulated during sugar beet germination and affects early root growth in a dose-dependent manner. Planta 240, 679–686.
Alcázar R, Altabella T, Marco F, et al. (2010) Polyamines: molecules with regulatory functions in plant abiotic stress tolerance. Planta 231, 1237–1249.
Amini S, Maali-Amiri R, Kazemi-Shahandashti SS, et al. (2021) Effect of cold stress on polyamine metabolism and antioxidant responses in chickpea. J Plant Physiol 258, 153387.
Amini S, Maali-Amiri R, Mohammadi R, Kazemi-Shahandashti SS (2017) cDNA-AFLP analysis of transcripts induced in chickpea plants by TiO2 nanoparticles during cold stress. Plant Physiol Biochem 111, 39-49.
Azevedo Neto AD, Prisco JT, Enéas-Filho J, et al. (2004) Effects of salt stress on plant growth, stomatal response and solute accumulation of different maize genotypes. Braz J Plant Physiol 16, 31–38. Bitrián M, Zarza X, Altabella T, Tiburcio AF, Alcázar R (2012) Polyamines under abiotic stress: metabolic crossroads and hormonal cross talks in plants. Metabolites 2(3), 516-528.
Bulens I, Van de Poel B, Hertog MLATM, et al. (2014) Dynamic changes of the ethylene biosynthesis in ‘Jonagold’ apple. Physiol Plant 150, 161–173.
Campestre MP, Bordenave CD, Origone AC, et al. (2011) Polyamine catabolism is involved in response to salt stress in soybean hypocotyls. J Plant Physiol 168, 1234–1240.
Cervelli M, Di Caro O, Di Penta A, et al. (2004) A novel C-terminal sequence from barley polyamine oxidase is a vacuolar sorting signal. Plant J 40, 410–418.
Chen D, Ma X, Li C, et al. (2014) A wheat aminocyclopropane-1-carboxylate oxidase gene, TaACO1, negatively regulates salinity stress in Arabidopsis thaliana. Plant Cell Rep 33, 1815–1827.
Ciardi JA, Deikman J, Orzolek MD (1997) Increased ethylene synthesis enhances chilling tolerance in tomato. Physiol Plant 101, 333–340.
Dong H, Zhen Z, Peng J, et al. (2011) Loss of ACS7 confers abiotic stress tolerance by modulating ABA sensitivity and accumulation in Arabidopsis. J Exp Bot 62, 4875–4887.
Freitas VS, Souza Miranda R, Costa JH, et al. (2018) Ethylene triggers salt tolerance in maize genotypes by modulating polyamine catabolism enzymes associated with H2O2 production. Environ Exp Bot 145, 75-86.
Gong B, Li X, VandenLangenberg KM, et al. (2014) Overexpression of S-adenosyl-L methionine synthetase increased tomato tolerance to alkali stress through polyamine metabolism. Plant Biotechnol J 12, 694–708.
Gong B, Wang X, Wei M, et al. (2016) Overexpression of S-adenosylmethionine synthetase1 enhances tomato callus tolerance to alkali stress through polyamine and hydrogen peroxide cross-linked networks. Plant Cell Tiss Organ Cult 124, 377–439.
Groppa MD, Benavides MP (2008) Polyamines and abiotic stress: recent advances. Amino Acids 34, 35–45.
Grzesiak M, Filek M, Barbasz A, et al. (2013) Relationships between polyamines, ethylene, osmoprotectants and antioxidant enzymes activities in wheat seedlings after short-term PEG-and NaCl-induced stresses. Plant Growth Regul 69(2), 177-189.
Guo Z, Tan J, Zhuo C, et al. (2014) Abscisic acid, H2O2 and nitric oxide interactions mediated cold-induced S-adenosylmethionine synthetase in Medicago sativa subsp. falcata that confers cold tolerance through up-regulating polyamine oxidation. Plant Biotech J 12, 601–612.
Gupta K, Dey A, Gupta B (2013) Plant polyamines in abiotic stress responses. Acta Physiol Plant 35(7), 2015-2036.
Habben JE, Bao X, Bate NJ, et al. (2014) transgenic alteration of ethylene biosynthesis increases grain yield in maize under field drought‐stress conditions. Plant Biotech J 12(6), 685-693.
Heidarvand L, Amiri RM, Naghavi MR et al. (2011) Physiological and morphological characteristics of chickpea accessions under low temperature stress. Russ J Plant Physiol 58, 157-163.
Heidarvand L, Maali-Amiri R (2013) Physio-biochemical and proteome analysis of chickpea in early phases of cold stress. J Plant Physiol 170, 459-469.
Hugo JP, Jan MC (1987) High speed HPLC analysis of polyamines in plant tissues. Plant Physiol 83:232-234.
Hurry VM, Huner NPA (1991) Low growth temperature effects a differential inhibition of photosynthesis in spring and winter wheat. Plant Physiol 96,491–497.
Jha UC, Nayyar H, Jha R, et al. (2020) Chickpea Breeding for Abiotic Stress: Breeding Tools and ‘Omics’ Approaches for Enhancing Genetic Gain. In Accelerated Plant Breeding, Volume 3 (pp. 211-234). Springer, Cham.
Jiang C, Belfield EJ, Cao Y, et al. (2013) An Arabidopsis soil salinity-tolerance mutation confers ethylene-mediated enhancement of sodium/potassium homeostasis. Plant Cell 25, 3535–3552.
Jiménez-Bremont JF, Ruiz OA, Rodríguez-Kessler M (2007) Modulation of spermidine and spermine levels in maize seedlings subjected to long-term salt stress. Plant Physiol Bioch 45, 812–821.
Kazemi Shahandashti SS, Maali Amiri R, Zeinali H,  Ramezanpour SS (2013) Change in membrane fatty acid compositions and cold-induced responses in chickpea. Mol Biol Rep 40(2), 893-903.
Kazemi-Shahandashti SS, Maali-Amiri R (2018) Global insights of protein responses to cold stress in plants: Signaling, defence, and degradation. J Plant Physiol 226, 123-135.
Kazemi-Shahandashti SS, Maali-Amiri R, Zeinali H et al. (2014) Effect of short-term cold stress on oxidative damage and transcript accumulation of defense-related genes in chickpea seedlings. J Plant Physiol 171, 1106-1116.
Lasanajak Y, Minocha R, Minocha SC, et al. (2014) Enhanced flux of substrates into polyamine biosynthesis but not ethylene in tomato fruit engineered with yeast S-adenosylmethionine decarboxylase gene. Amino Acids 46(3), 729-742.
Lin Y, Wang J, Zu Y, Tang Z (2012) Ethylene antagonizes the inhibition of germination in Arabidopsis induced by salinity by modulating the concentration of hydroge peroxide. Acta Physiol Plant 34, 1895–1904.
Livak KJ, Schmittgen TD (2001) Analysis of relative gene expression data using real-time quantitative PCR and the 2−ΔΔCT method. Methods 25, 402–408.
López-Gómez M, Hidalgo-Castellanos J, Iribarne C, Luch C (2014) Proline accumulation has prevalence over polyamines in nodules of Medicago sativa in symbiosis with Sinorhizobium meliloti during the initial response to salinity. Plant Soil 374, 149-159.
Loreto F, Velikova V (2001) Isoprene produced by leaves protects the photosynthetic apparatus against ozone damage, quenches ozone products, and reduces lipid peroxidation of cellular membranes. Plant Physiol 127, 1781-1787.
Lyzenga WJ, Booth JK, Stone SL (2012) The Arabidopsis RING‐type E3 ligase XBAT32 mediates the proteasomal degradation of the ethylene biosynthetic enzyme, 1‐aminocyclopropane‐1‐carboxylate synthase 7. Plant J 71(1), 23-34.
Merga B, Haji J (2019) Economic importance of chickpea: Production, value, and world trade. Cogent Food and Agricul 5:1615718.
Milhinhos A, Miguel CM (2013) Hormone interactions in xylem development: a matter of signals. Plant Cell Rep 32(6), 867-883.
Moeder W, Barry CS, Tauriainen AA, et al. (2002) Ethylene synthesis regulated by biphasic induction of 1-aminocyclopropane-1-carboxylic acid synthase and 1-aminocyclopropane-1-carboxylic acid oxidase genes is required for hydrogen peroxide accumulation and cell death in ozone-exposed tomato. Plant Physiol 130(4), 1918-1926.
Moschou, PN, Delis ID, Paschalidis KA, Roubelakis-Angelakis KA (2008a) transgenic tobacco plants overexpressing polyamine oxidase are not able to cope with oxidative burst generated by abiotic factors. Physiol Plant 133, 140–156.
Moschou PN, Paschalidis KA, Delis ID, et al. (2008b) Spermidine exodus and oxidation in the apoplast induced by abiotic stress is responsible for H2O2 signatures that direct tolerance responses in tobacco. Plant Cell 20, 1708–1724.
Moschou PN, Sanmartin M, Andriopoulou AH, et al. (2008c) Bridging the gap between plant and mammalian polyamine catabolism: a novel peroxisomal polyamine oxidas responsible for a full back-conversion pathway in Arabidopsis. Plant Physiol 147, 1845–1857.
Müller M, Munné-Bosch S (2015) Ethylene response factors: a key regulatory hub in hormone and stress signaling. Plant physiol 169(1), 32-41.
Nayyar H, Bains TS, Sanjeev K (2005) Chilling stressed chickpea seedlings: effect of cold acclimation, calcium and abscisic acid on cryoprotective solutes and oxidative damage. Environ Exp Bot 54, 275–285.
Peng H, Cheng H, Yu X et al. (2010) Molecular analysis of an actin gene, CarACT1, from chickpea (Cicer arietinum L.). Mol Biol Rep 37, 1081.
Pfaffl MW, Horgan GW, Dempfle L (2002) Relative expression software tool (REST) for group-wise comparison and statistical analysis of relative expression results in real-time PCR. Nucleic Acids Res 30, e36.
Rakei A, Maali-Amiri R, Zeinali H, Ranjbar M (2016) DNA methylation and physio-biochemical analysis of chickpea in response to cold stress. Protoplasma 253(1), 61-76.
Rodríguez-Kessler M, Alpuche-Solís AG, Ruiz OA, Jimenez-Bremont JF (2006). Effect of salt stress on the regulation of maize (Zea mays L.) genes involved in polyamine biosynthesis. Plant Growth Regul 48(2), 175-185.
Saeed A, Hovsepyan H, Darvishzadeh R et al. (2011) Genetic diversity of Iranian accessions, improved lines of chickpea (Cicer arietinum L.) and their wild relatives by using simple sequence repeats. Plant Mol Biol Rep 29, 848-858.
Sauter M, Moffatt B, Saechao MC, et al. (2013) Methionine salvage and S-adenosylmethionine: essential links between sulfur, ethylene and polyamine biosynthesis. Biochem J 451, 145–154.
Shi J, Habben JE, Archibald RL, et al. (2015) Overexpression of ARGOS genes modifies plant sensitivity to ethylene, leading to improved drought tolerance in both Arabidopsis and maize. Plant Physiol 169, 266–282.
Stearns JC, Glick BR (2003) Transgenic plants with altered ethylene biosynthesis or perception. Biotechnol Adv 21, 193–210.
Stepanova AN, Alonso JM (2009) Ethylene signaling and response: where different regulatory modules meet. Curr Opin Plant Biol 12,548-555.
Sudhakar C, Veeranagamallaiah G, Nareshkumar A, et al. (2014) Polyamine metabolism influences antioxidant defense mechanism in foxtail millet (Setaria italica L.) cultivars with different salinity tolerance. Plant Cell Rep 34, 141–156.
Tavladoraki P, Cona A, Federico R, et al. (2012) Polyamine catabolism: target for antiproliferative therapies in animals and stress tolerance strategies in plants. Amino Acids 42, 411–426.
Tavladoraki P, Rossi MN, Saccuti G, et al. (2006) Heterologous expression and biochemical characterization of a polyamine oxidase from Arabidopsis involved in polyamine back conversion. Plant Physiol 141, 1519–1532.
Thudi M, Chitikineni A, Liu X, et al. (2016) Recent breeding programs enhanced genetic diversity in both desi and kabuli varieties of chickpea (Cicer arietinum L.). Sci Rep 6(1), 1-10.
Van de Poel B, Smet D, Van Der Straeten D (2015) Ethylene and hormonal cross talk in vegetative growth and development. Plant Physiol 169(1), 61-72.
Van de Poel B, Van Der Straeten D (2014) 1-aminocyclopropane-1-carboxylic acid (ACC) in plants: more than just the precursor of ethylene! Front Plant Sci 5, 640.
Wang B, Zhang J, Xia X, Zhang WH (2011) Ameliorative effect of brassinosteroid and ethylene on germination of cucumber seeds in the presence of sodium chloride. Plant Growth Regul 65, 407–413.
Wang H, Liang X, Huang J, et al. (2010) Involvement of ethylene and hydrogen peroxide in induction of alternative respiratory pathway in salt-treated Arabidopsis calluses. Plant Cell Physiol 51, 1754–1765.
Wang X, Shi G, Xu Q, Hu J (2007) exogenous polyamines enhance copper tolerance of Nymphoides peltatum. J Plant Physiol 164, 1062–1070.
Wi SJ, Jang SJ, Park KY (2010) Inhibition of biphasic ethylene production enhances tolerance to abiotic stress by reducing the accumulation of reactive oxygen species in Nicotiana tabacum. Mol Cell 30, 37–49.
Wi SJ, Kim SJ, Kim WT, Park KY (2014) Constitutive S-adenosylmethionine decarboxylase gene expression increases drought tolerance through inhibition of reactive oxygen species accumulation in Arabidopsis. Planta 239, 979–988.
Wu L, Zhang Z, Zhang H, et al (2008) Transcriptional modulation of ethylene response factor protein JERF3 in the oxidative stress response enhances tolerance of tobacco seedlings to salt, drought, and freezing. Plant Physiol 148, 1953–1963.
Yang L, Zu YG, Tang ZH (2013) Ethylene improves Arabidopsis salt tolerance mainly via retaining K+ in shoots and roots rather than decreasing tissue Na+ content. Environ Exp Bot 86, 60–69.
Yang SF, Hoffman NE (1984) Ethylene biosynthesis and its regulation in higher plants. Ann Rev Plant Physiol 35(1), 155-189.
Yin ZP, Li S, Ren J, Song XS (2014) Role of spermidine and spermine in alleviation of drought-induced oxidative stress and photosynthetic inhibition in Chinese dwarf cherry (Cerasus humilis) seedlings. Plant Growth Regul 74, 209–218.
Zarza X, Atanasov KE, Marco F, et al. (2017) Polyamine oxidase 5 loss-of-function mutations in Arabidopsis thaliana trigger metabolic and transcriptional reprogramming and promote salt stress tolerance. Plant Cell Environ 40, 527–542.
Zhai, Y., Wang, Y., Li, Y., et al. (2013) Isolation and molecular characterization of GmERF7, a soybean ethylene-response factor that increases salt stress tolerance in tobacco. Gene 513, 174–183.
Zhang Z, Huang R (2010) Enhanced tolerance to freezing in tobacco and tomato overexpressing transcription factor TERF2/LeERF2 is modulated by ethylene biosynthesis. Plant Mol Biol 73, 241–249.
Zhao M, Liu W, Xia X, et al. (2014) Cold acclimation-induced freezing tolerance of Medicago truncatula seedlings is negatively regulated by ethylene. Physiol Plant 152, 115–129.