Skip to main content
SpringerLink
Log in

Calcium triggers protein kinases-induced signal transduction for augmenting the thermotolerance of developing wheat (Triticum aestivum) grain under the heat stress

  • Original Article
  • Published:
Journal of Plant Biochemistry and Biotechnology Aims and scope Submit manuscript

Abstract

Calcium acts as a signaling molecule and plays very important role in plants-tolerance to the abiotic stresses including the heat stress. We studied the effect of exogenous Ca2+ (10 mM) prior to the heat stress (42 °C, 2 h) on different molecular and biochemical parameters associated with thermotolerance in wheat (Triticum aestivum). We could observe 64 and 52 unique protein spots in HD2967 (thermotolerant) and HD2329 (thermosusceptible) cultivars under normal and heat stress condition. Similarly, 13 and 9 (HD2967) and 19 and 9 proteins (HD2329) were observed to be up-regulated and down-regulated in response to HS. MALDI-TOF-TOF/MS characterization identified the differentially expressed protein (DEP) spots as calcium dependent protein kinase (CDPK), oxygen evolving enhancer protein, HSP17, HSP70, Rubisco activase etc. Transcript profiling of identified stress-associated genes (SAGs) showed very high expression of CDPK, HSFA4a, HSP17, SOD and APX in response to Ca2+ + HS in HD2967 compared to HD2329 cultivars of wheat. Similarly accumulation of signaling molecules (H2O2 and CDPK) as well as osmolyte was observed maximum in response to Ca2+ + HS in HD2967 compared to HD2329 cultivars. Very high activities of guaiacol-peroxidase and ascorbate peroxidase were observed in Ca-treated HD2967 compared to HD2329 when exposed to HS. It is the abundance of these chaperones and antioxidant enzymes in thermotolerant cultivar which limit the accumulation of H2O2 in response to Ca2+ and HS. Exogenous Ca2+ application showed negative correlation with lipid peroxidation and positive correlation with total antioxidant capacity of the cell system under the elevated temperature. Alteration in Ca2+ efflux triggers the activities of kinases and peroxidases more in tolerant compared to susceptible cultivars which regulate the accumulation of ROS inside the cell and attenuate the lipid peroxidation process. Ultimately, it enhances the total antioxidant capacity and thermotolerance of the plants under the heat stress.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Log in via an institution

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Fig. 1
Fig. 2
Fig. 3
Fig. 4

Similar content being viewed by others

Abbreviations

APX:

Ascorbate peroxidase

HS:

Heat stress

HSFs:

Heat shock transcription factors

HSP:

Heat shock protein

MDA:

Malondialdehyde

qRT-PCR:

Quantitative real-time PCR

SOD:

Superoxide dismutase

References

  • Amirjani M (2012) Estimation of wheat responses to high heat stress. Am-Eurasian J Sustain Agric 6:222–233

    Google Scholar 

  • Ara N, Nakkanong K, Lv W, Yang J, Hu Z, Zhang M (2013) Antioxidant enzymatic activities and gene expression associated with heat tolerance in the stems and roots of two cucurbit species (“Cucurbita maxima” and “Cucurbita moschata”) and their interspecific inbred line “maxchata”. Int J Mol Sci 14:24008–24028

    Article  PubMed Central  PubMed  Google Scholar 

  • Bates LS, Waldren RP, Teare ID (1973) Rapid determination of free proline for water-stress studies. Plant Soil 39:205–207

    Article  CAS  Google Scholar 

  • Benzie IFF, Strain JJ (1999) Ferric reducing/antioxidant power assay: direct measure of the total antioxidant activity of biological fluids and modified version for simultaneous measurement of total antioxidant power and ascorbic acid concentration. Meth Enzymol 299:15–27

    Article  CAS  PubMed  Google Scholar 

  • Blokhina O, Eija V, Kurt VF (2003) Antioxidants, oxidative damage and oxygen deprivation stress: a review. Ann Bot 91:179–194

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  • Bowne JB, Erwin TA, Juttner J, Schnurbusch T, Langridge P, Bacic A, Roessner U (2012) Drought responses of leaf tissues from wheat cultivars of differing drought tolerance at the metabolite level. Mol Plant 5(2):418–429

    Article  CAS  PubMed  Google Scholar 

  • Bradford M (1976) A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein dye binding. Anal Biochem 72:248–254

    Article  CAS  PubMed  Google Scholar 

  • Chen WP, Li PH (2001) Chilling induced Ca2+ over load enhances production of active oxygen species in maize (Zea mays L.) cultured cells: the effect of abscisic acid treatment. Plant Cell Environ 24:791–800

    Article  CAS  Google Scholar 

  • Dubrovina AS, Kiselev KV, Khristenko VS (2013) Expression of calcium-dependent protein kinase (CDPK) genes under abiotic stress conditions in wild-growing grapevine Vitis amurensis. J Plant Physiol 15:1491–1500

    Article  Google Scholar 

  • Evers J, Johnson MC, Marini MA (1994) Peroxidative activities of hemoglobin and hemoglobin derivatives. In: Evers J, Vandegriff KD, Winslow RM (eds) Methods of enzymology, vol 231. Academic, San Diego, pp 547–561

    Google Scholar 

  • Ge P, Ma C, Wang S, Gao L, Li X, Guo G, Ma W, Yan Y (2012) Comparative proteomic analysis of grain development in two spring wheat varieties under drought stress. Anal Bioanal Chem 402(3):1297–1313

    Article  CAS  PubMed  Google Scholar 

  • Heath RL, Packer L (1968) Photoperoxidation in isolated chloroplasts. Arch Biochem Biophys 125:189–198

    Article  CAS  PubMed  Google Scholar 

  • Hong Z, Lakkineni K, Zhang Z, Verma DP (2000) Removal of feedback inhibition of delta(1)-pyrroline-5-carboxylate synthetase results in increased proline accumulation and protection of plants from osmotic stress. Plant Physiol 122:1129–1136

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  • Hu M, Shi Z, Zhang Z, Zhang Y, Li H (2012) Effects of exogenous glucose on seed germination and antioxidant capacity in wheat seedlings under salt stress. Plant Growth Regul 68(2):177–188

    Article  CAS  Google Scholar 

  • Kamal MAH, Kim K-H, Shin K-H, Choi J-S, Baik B-K, Tsujimoto H, Heo HY, Park CS, Woo SH (2010) Abiotic stress responsive proteins of wheat grain determined using proteomics technique. Aust J Crop Sci 4(3):196–208

    CAS  Google Scholar 

  • Kovacs Z, Simon-Sarkadi L, Vashegyi I, Kocsy G (2012) Different accumulation of free amino acids during short-and long-term osmotic stress in wheat. Sci World J. doi:10.1100/2012/216521

    Google Scholar 

  • Kudla J, Batistic O, Hashimoto K (2010) Calcium signals: the lead currency of plant information processing. Plant Cell 22:541–563

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  • Kumar RR, Goswami S, Kumar N, Pandey SK, Pandey VC, Sharma SK, Pathak H, Rai RD (2011) Expression of novel ascorbate peroxidase isoenzymes of wheat (Triticum aestivum L) in response to heat stress. Int J Plant Physiol Biochem 3(11):188–194

    CAS  Google Scholar 

  • Kumar RR, Goswami S, Sharma SK, Singh K, Gadpayle KA, Kumar N, Rai GK, Singh M, Rai RD (2012) Protection against heat stress in wheat involves change in cell membrane stability, antioxidant enzymes, osmolyte, H2O2 and transcript of heat shock protein. Int J Plant Physiol Biochem 4:83–91

    CAS  Google Scholar 

  • Kumar RR, Goswami S, Sharma SK, Singh K, Gadpayle KA, Singh SD, Pathak H, Rai RD (2013) Differential expression of heat shock protein and alteration in osmolyte accumulation under heat stress in wheat. J Plant Biochem Biotechnol 22(1):16–26

    Article  CAS  Google Scholar 

  • Kumar RR, Singh GP, Goswami S, Pathak H, Rai RD (2014) Proteome analysis of wheat (Triticum aestivum) for the identification of differentially expressed heat-responsive proteins. Aust J Crop Sci 8(6):973–986

    CAS  Google Scholar 

  • Laemmli UK (1970) Cleavage of structural proteins during the assembly of the head of bacteriophage. Nature 227:680–685

    Article  CAS  PubMed  Google Scholar 

  • Liu HT, Gao F, Li GL, Han JL, Liu DL, Sun DY, Zhou RG (2008) The calmodulin-binding protein kinase 3 is part of heat-shock signal transduction in Arabidopsis thaliana. Plant J 55:760–773

    Article  CAS  PubMed  Google Scholar 

  • 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

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  • Mattioli R, Marchese D, D’Angeli S, Altamura MM, Costantino P, Trovato M (2008) Modulation of intracellular proline levels affects flowering time and inflorescence architecture in Arabidopsis. Plant Mol Biol 66:277–288

    Article  CAS  PubMed  Google Scholar 

  • Miller G, Honig A, Stein H, Suzuki N, Mittler R, Zilberstein A (2009) Unraveling delta 1-pyrroline-5-carboxylate proline cycle in plants by uncoupled expression of proline oxidation enzymes. J Biol Chem 284:26482–26492

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  • Nagi KM, Devaraj VR (2011) Antioxidant and other biochemical defense responses of Macrotyloma uniflorum (Lam) Verdc. (Horse gram) induced by high temperature and salt stress. Braz J Plant Physiol 23:187–195

    Google Scholar 

  • Nakano Y, Asada K (1981) Hydrogen peroxide is scavenged by ascorbate-specific peroxidase in spinach chloroplasts. Plant Cell Physiol 22:867–880

    CAS  Google Scholar 

  • Neill SJ, Desikan R, Clarke A, Hurst RD, Hencock JT (2002) Hydrogen peroxide and nitric oxide as signaling molecules in plants. J Exp Bot 53:1237–1247

    Article  CAS  PubMed  Google Scholar 

  • Orozco-Cardenas ML, Narvaez-Vasquez J, Ryan CA (2001) Hydrogen peroxide acts as a second messenger for the induction of defense genes in tomato plants in response to wounding, systemin and methyl jasmonate. Plant Cell 13:179–191

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  • Passardi F, Penel C, Dunand C (2004) Performing the paradoxical: how plant peroxidases modify the cell wall. Trends Plant Sci 9:534–540

    Article  CAS  PubMed  Google Scholar 

  • Pfaffl MW (2001) A new mathematical model for relative quantification in real-time RT-PCR. Nucleic Acids Res 29(9):45

    Article  Google Scholar 

  • Polle H, Rennenberg (1993) Significance of antioxidants in plant adaptation to environmental stress. In: Mansfield T, Fowden L, Stoddard F (eds) Plant adaptation to environmental stress. Chapman and Hall, London, pp 263–273

    Google Scholar 

  • Saijo Y, Hata S, Kyozuka J, Shimamoto K, Izui K (2000) Over-expression of a single Ca2+-dependent protein kinase confers both cold and salt/drought tolerance on rice plants. Plant J 23:319–327

    Article  CAS  PubMed  Google Scholar 

  • Sathiyaraj G, Lee OR, Parvin S, Khorolragchaa A, Kim Y-J, Yang DC (2011) Transcript profiling of antioxidant genes during biotic and abiotic stresses in Panax ginseng. Mol Biol Rep 38(4):2761–2769

    Article  CAS  PubMed  Google Scholar 

  • Sharma P, Dubey RS (2005) Drought induces oxidative stress and enhances the activities of antioxidant enzymes in growing rice seedlings. Plant Growth Regul 46:209–221

    Article  CAS  Google Scholar 

  • Siddiqui MH, Al-Whaibi MH, Basalah MO (2011) Interactive effect of calcium and gibberellin on nickel tolerance in relation to antioxidant systems in Triticum aestivum L. Protoplasma 248:503–511

    Article  CAS  PubMed  Google Scholar 

  • Sircelj H, Tausz M, Grill D, Batie F (2005) Biochemical responses in leaves of two apple tree cultivars subjected to progressing drought. J Plant Physiol 162:1308–1318

    Article  CAS  PubMed  Google Scholar 

  • Szekely G, Abraham E, Cseplo A, Rigo G, Zsigmond L, Csiszar J, Ayaydin F, Strizhov N, Jasik J, Schmelzer E, Koncz C, Szabados L (2008) Duplicated P5CS genes of Arabidopsis play distinct roles in stress regulation and developmental control of proline biosynthesis. Plant J 53:11–28

    Article  CAS  PubMed  Google Scholar 

  • Tan W, Meng QW, Brestic M, Olsovska K, Yang XH (2011) Photosynthesis is improved by exogenous calcium in heat-stressed tobacco plants. J Plant Physiol 168:2063–2071

    Article  CAS  PubMed  Google Scholar 

  • Taulavuori E, Hellstrom E, Taulavuori K, Laine K (2001) Comparison of two methods used to analyze lipid peroxidation from Vaccinium myrtillus (L) during snow removal, reacclimation and cool acclimation. J Exp Bot 52:2375–2380

    Article  CAS  PubMed  Google Scholar 

  • Upadhyaya H, Panda SK, Dutta BK (2011) CaCl2 improves post drought recovery potential in Camellia sinensis (L) O Kuntze. Plant Cell Rep 30:495–503

    Article  CAS  PubMed  Google Scholar 

  • Vacca RA, de Pinto MC, Valenti D, Passarella S, Maria E, De Gara L (2004) Production of reactive oxygen species, alteration of cytosolic ascorbate peroxidase, and impairment of mitochondrial metabolism are early events in heat shock induced programmed cell death in tobacco Bright-Yellow 2 cells. Plant Physiol 134:1100–1112

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  • Varga B, Janda T, Laszlo E, Veisz O (2012) Influence of abiotic stresses on the antioxidant enzyme activity of cereals. Acta Physiol Plant 34(3):849–858

    Article  CAS  Google Scholar 

  • Veremeichik GN, Shkryl YN, Pinkus SA, Bulgakov VP (2014) Expression profiles of calcium-dependent protein kinase genes (CDPK1-14) in Agrobacterium rhizogenes pRiA4-transformed calli of Rubia cordifolia under temperature-and salt-induced stresses. J Plant Physiol 171(7):467–474

    Article  CAS  PubMed  Google Scholar 

  • Vranova E, Inze D, Breusegem FV (2002) Signal transduction during oxidative stress. J Exp Bot 53:1227–1236

    Article  CAS  PubMed  Google Scholar 

  • Wahid A, Gelani S, Ashraf M, Foolad MR (2007) Heat tolerance in plants: an overview. Environ Exp Bot 61:199–223

    Article  Google Scholar 

  • Wang C-T, Shao J-M (2013) Characterization of the ZmCK1 gene encoding a calcium-dependent protein kinase responsive to multiple abiotic stresses in maize. Plant Mol Biol Rep 31(1):222–230

    Article  CAS  Google Scholar 

  • Wang Y, Yu QY, Tang XX, Wang LL (2009) Calcium pretreatment increases thermotolerance of Laminaria japonica sporophytes. Prog Nat Sci 19:435–442

    Article  Google Scholar 

  • Zehra A, Gul B, Ansari R, Khan MA (2012) Role of calcium in alleviating effect of salinity on germination of Phragmites karka seeds. South Afr J Bot 78:122–128

    Article  CAS  Google Scholar 

  • Zhang W, Zhou R-G, Gao Y-J, Zheng S-Z, Xu P, Zhang S-Q, Sun D-Y (2009) Molecular and genetic evidence for the key role of AtCaM3 in heat-shock signal transduction in Arabidopsis. Plant Physiol 149:1773–1784

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  • Zhou B, Guo Z (2009) Calcium is involved in the abscisic acid-induced ascorbate peroxidase, superoxide dismutase and chilling resistance in Stylosanthes guianensis. Biol Plant 53:63–68

    Article  CAS  Google Scholar 

Download references

Acknowledgment

The Author’s sincerely thanks Indian Agricultural Research Institute (IARI) and Indian Council of Agriculture Research (ICAR) for providing the financial support under the National Initiative for Climate Resilient Agriculture (NICRA) project.

Conflict of interest

The authors declare that they have no competing interests.

Author information

Authors and Affiliations

  1. Division of Biochemistry, Indian Agricultural Research Institute, New Delhi, 110012, India

    Suneha Goswami, Ranjeet R. Kumar, Sushil K. Sharma, Khushboo Singh, Richa Gupta & Raj D. Rai

  2. Division of Genetics, Indian Agricultural Research Institute, New Delhi, India, 110012

    Yugal K. Kala, Gaurav Dhavan & Gyanendra P. Singh

  3. Sher-e-Kashmir University of Agricultural Sciences & Technology (SKUAST), Jammu, India

    Gyanendra K. Rai

  4. CESCRA, Indian Agricultural Research Institute, New Delhi, India, 110012

    Himanshu Pathak

Authors
  1. Suneha Goswami
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Ranjeet R. Kumar
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Sushil K. Sharma
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Yugal K. Kala
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Khushboo Singh
    View author publications

    You can also search for this author in PubMed Google Scholar

  6. Richa Gupta
    View author publications

    You can also search for this author in PubMed Google Scholar

  7. Gaurav Dhavan
    View author publications

    You can also search for this author in PubMed Google Scholar

  8. Gyanendra K. Rai
    View author publications

    You can also search for this author in PubMed Google Scholar

  9. Gyanendra P. Singh
    View author publications

    You can also search for this author in PubMed Google Scholar

  10. Himanshu Pathak
    View author publications

    You can also search for this author in PubMed Google Scholar

  11. Raj D. Rai
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Ranjeet R. Kumar.

Additional information

Author contributions

SG RRK- conceived and designed the experiments. SG RRK SKS KS- performed the transcript profiling. RG GD GKR GPS- performed biochemical estimations. SG RRK HP RDR- wrote the paper and edited the manuscript.

Equal contribution made by Suneha Goswami and Ranjeet R. Kumar.

Electronic supplementary material

Below is the link to the electronic supplementary material.

Table S1

List of primers used for the quantitative real-time PCR (qRT-PCR) of signaling molecules and stress-associated genes (SAGs) in HD2967 (thermotolerant) and HD2329 (thermosusceptible) cultivars of wheat in response to Ca2+ and HS (DOCX 16 kb)

Table S2

Unique protein spots identified in two dimensional electrophoresis (2-DE) gels of control (22 °C) and heat shock treated (42 °C, 2 h) samples of HD2967 (thermotolerant) cultivar of wheat; ImageMaster Platinum 7.0 (IMP7) software (GE Healthcare Life Sciences) was used for the gel characterization (DOCX 16 kb)

Table S3

Unique protein spots identified in two dimensional electrophoresis (2-DE) gels of control (22 °C) and heat shock treated (42 °C, 2 h) samples of HD2329 (thermosusceptible) cultivar of wheat; ImageMaster Platinum 7.0 (IMP7) software (GE Healthcare Life Sciences) was used for the gel characterization (DOCX 16 kb)

Fig. S1

Peptide mass fingerprinting of differentially expressed protein (DEP) spot (match ID 69) identified from the 2-DE gel using matrix-assisted laser desorption ionization - time of flight mass spectrometer (MALDI-TOF-TOF/MS), m/z data was subjected to MASCOT search (Matrix science) for the identification of protein. (DOCX 285 kb)

Fig. S2

Effects of exogenous Ca2+ and heat stress on proline accumulation and free amino acid content in thermotolerant (HD2967) and thermosusceptible (HD2329) cultivars of wheat, (a) Proline accumulation, (b) free amino acid content; T0 - 22 °C, T1 - HS of 42 °C for 2 h, T2 - Ca10mM, and T3 - Ca10mM + HS of 42 °C for 2 h; Means for varieties, treatments and interaction are significant at p = 0.05, vertical bars indicate s.e. (n = 3). (DOCX 76 kb)

Fig. S3

Effects of exogenous Ca2+ and heat stress on peroxidases activities in HD2967 (thermotolerant) and HD2329 (thermosusceptible) cultivars of wheat, (a) guaiacol peroxidase (GPX), and (b) ascorbate peroxidase (APX) activity; T0 - 22 °C, T1 - HS of 42 °C for 2 h, T2 - Ca10mM, and T3 - Ca10mM + HS of 42 °C for 2 h, Means for varieties, treatments and interaction are significant at p = 0.05, vertical bars indicate s.e. (n = 3). (DOCX 82 kb)

Rights and permissions

Reprints and permissions

About this article

Check for updates. Verify currency and authenticity via CrossMark

Cite this article

Goswami, S., Kumar, R.R., Sharma, S.K. et al. Calcium triggers protein kinases-induced signal transduction for augmenting the thermotolerance of developing wheat (Triticum aestivum) grain under the heat stress. J. Plant Biochem. Biotechnol. 24, 441–452 (2015). https://doi.org/10.1007/s13562-014-0295-1

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s13562-014-0295-1

Keywords

Search

Navigation