Skip to main content
SpringerLink
Log in
Book cover

Cyclic Nucleotide Signaling in Plants pp 107–119Cite as

Calcium Imaging of the Cyclic Nucleotide Response

  • Protocol
  • First Online:

Part of the book series: Methods in Molecular Biology ((MIMB,volume 1016))

Abstract

Calcium (Ca2+) is a key component of the signalling network by which plant cells respond to developmental and environmental signals. A change in guard cell cytosolic free Ca2+([Ca2+]cyt) is an early event in the response of stomata to both opening and closing stimuli, and cyclic nucleotide-mediated Ca2+ signalling has been implicated in the regulation of stomatal aperture. A range of techniques have been used to measure [Ca2+]cyt in plant cells. Here we describe a potential method for imaging cyclic nucleotide-induced changes in [Ca2+]cyt in guard cells using the cameleon ratiometric Ca2+ reporter protein.

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

Protocol
USD   49.95
Price excludes VAT (USA)
  • Available as PDF
  • Read on any device
  • Instant download
  • Own it forever
eBook
USD   84.99
Price excludes VAT (USA)
  • Available as EPUB and PDF
  • Read on any device
  • Instant download
  • Own it forever
Softcover Book
USD   139.00
Price excludes VAT (USA)
  • Compact, lightweight edition
  • Dispatched in 3 to 5 business days
  • Free shipping worldwide - see info
Hardcover Book
USD   109.99
Price excludes VAT (USA)
  • Durable hardcover edition
  • Dispatched in 3 to 5 business days
  • Free shipping worldwide - see info

Tax calculation will be finalised at checkout

Purchases are for personal use only

Learn about institutional subscriptions

Springer Nature is developing a new tool to find and evaluate Protocols. Learn more

References

  1. Dodd AN, Kudla J, Sanders D (2010) The language of calcium signaling. Annu Rev Plant Biol 61:593–620

    Article  PubMed  CAS  Google Scholar 

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

    Article  PubMed  CAS  Google Scholar 

  3. Knight H, Trewavas AJ, Knight MR (1997) Calcium signalling in Arabidopsis thaliana responding to drought and salinity. Plant J 12:1067–1078

    Article  PubMed  CAS  Google Scholar 

  4. Ranf S, Wunnenberg P, Lee J et al (2008) Loss of the vacuolar cation channel, AtTPC1, does not impair Ca2+ signals induced by abiotic and biotic stresses. Plant J 53:287–299

    Article  PubMed  CAS  Google Scholar 

  5. McAinsh MR, Clayton H, Mansfield TA et al (1996) Changes in stomatal behavior and guard cell cytosolic free calcium in response to oxidative stress. Plant Physiol 111:1031–1042

    PubMed  CAS  Google Scholar 

  6. Evans NH, McAinsh MR, Hetherington AM et al (2005) ROS perception in Arabidopsis thaliana, the ozone-induced calcium response. Plant J 41:615–626

    Article  PubMed  CAS  Google Scholar 

  7. Knight MR, Campbell AK, Smith SM et al (1991) Transgenic plant aequorin reports the effects of touch and cold-shock and elicitors on cytoplasmic calcium. Nature 352:524–526

    Article  PubMed  CAS  Google Scholar 

  8. Knight H, Trewavas AJ, Knight MR (1996) Cold calcium signaling in Arabidopsis involves two cellular pools and a change in calcium signature after acclimation. Plant Cell 8:489–503

    PubMed  CAS  Google Scholar 

  9. Clayton H, Knight MR, Knight H et al (1999) Dissection of the ozone-induced calcium signature. Plant J 17:575–579

    Article  PubMed  CAS  Google Scholar 

  10. Short EF, North KA, Roberts MR et al (2012) A stress-specific calcium signature regulating an ozone-responsive gene expression network in Arabidopsis. Plant J 71:948–961

    Article  PubMed  CAS  Google Scholar 

  11. Shacklock PS, Read ND, Trewavas AJ (1992) Cytosolic free calcium mediates red light-induced photomorphogenesis. Nature 358:753–755

    Article  CAS  Google Scholar 

  12. McAinsh MR, Brownlee C, Hetherington AM (1990) Abscisic acid-induced elevation of guard cell cytosolic Ca2+ precedes stomatal closure. Nature 343:186–188

    Article  CAS  Google Scholar 

  13. Allen GJ, Chu SP, Harrington CL et al (2001) A defined range of guard cell calcium oscillation parameters encodes stomatal movements. Nature 411:1053–1057

    Article  PubMed  CAS  Google Scholar 

  14. Ehrhardt DW, Wais R, Long SR (1996) Calcium spiking in plant root hairs responding to Rhizobium nodulation signals. Cell 85:673–681

    Article  PubMed  CAS  Google Scholar 

  15. Capoen W, Sun J, Wysham D et al (2011) Nuclear membranes control symbiotic calcium signaling of legumes. Proc Natl Acad Sci U S A 108:14348–14353

    Article  PubMed  CAS  Google Scholar 

  16. Rall TW, Sutherland EW, Berthet J (1957) The relation of epinephrine and glucagon to liver phosphorylase. J Biol Chem 224:1987–1995

    Google Scholar 

  17. Botsford JL, Harman JH (1998) cAMP in prokaryotes. Microbiol Rev 56:100–132

    Google Scholar 

  18. Penson SP, Schuurink RC, Fath A et al (1996) cGMP is required for gibberellic acid-induced gene expression in barley aleurone. Plant Cell 8:2325–2333

    PubMed  CAS  Google Scholar 

  19. Durner J, Wendehenne D, Klessig D (1998) Defence gene induction in tobacco by nitric oxide, cyclic GMP and cyclic ADP-ribose. Proc Natl Acad Sci U S A 95:10328–19333

    Article  PubMed  CAS  Google Scholar 

  20. Donaldson L, Ludidi N, Knight MR et al (2004) Salt and osmotic stress cause rapid increases in Arabidopsis thaliana cGMP levels. FEBS Lett 569:317–320

    Article  PubMed  CAS  Google Scholar 

  21. Newton RP, Smith CJ (2004) Cyclic nucleotides. Phytochemistry 65:2423–2437

    Article  PubMed  CAS  Google Scholar 

  22. Dubovaskaya LV, Bakakina YS, Kolesneva EV et al (2011) cGMP-dependent ABA-induced stomatal closure in the ABA-insensitive Arabidopsis mutant abi1-1. New Phytol 191:57–69

    Article  Google Scholar 

  23. Maathuis FJM, Sanders D (2001) Sodium uptake in Arabidopsis roots is regulated by cyclic nucleotides. Plant Physiol 127:1617–1625

    Article  PubMed  CAS  Google Scholar 

  24. Essah PA, Davenport R, Tester M (2003) Sodium influx and accumulation in Arabidopsis. Plant Physiol 133:307–318

    Article  PubMed  CAS  Google Scholar 

  25. Maathuis FJM (2006) cGMP modulates gene transcription and cation transport in Arabidopsis roots. Plant J 45:700–711

    Article  PubMed  CAS  Google Scholar 

  26. Bowler C, Neuhaus G, Yamagata H et al (1994) Cyclic GMP and calcium mediate phytochrome phototransduction. Cell 77:73–81

    Article  PubMed  CAS  Google Scholar 

  27. Ma Y, Szostkiewicz I, Korte A, Moes D et al (2009) Regulators of PP2C phosphatase activity function as abscisic acid sensors. Science 324:1064–1068

    PubMed  CAS  Google Scholar 

  28. Kurosaki F, Kaburakim H, Nishi A (1994) Involvement of plasma membrane-located calmodulin in the response decay of cyclic nucleotide-gated cation channel in cultured carrot cells. FEBS Lett 340:193–196

    Article  PubMed  CAS  Google Scholar 

  29. Volotovski ID, Sokolovsky SG, Molchan OV et al (1998) Second messengers mediate increases in cytosolic calcium in tobacco protoplasts. Plant Physiol 117:1023–1030

    Article  PubMed  CAS  Google Scholar 

  30. Malho R, Camacho L, Moutinho A (2000) Signalling pathways in pollen tube growth and reorientation. Ann Bot 85:59–68

    Article  CAS  Google Scholar 

  31. Kaupp UB, Seifert R (2002) Cyclic nucleotide-gated ion channels. Physiol Rev 82:769–824

    PubMed  CAS  Google Scholar 

  32. Schuurink RC, Shartzer SF, Fath A et al (1998) Characterization of a calmodulin-binding transporter from the plasma membrane of barley aleurone. Proc Natl Acad Sci U S A 95:1944–1949

    Article  PubMed  CAS  Google Scholar 

  33. Kohler C, Merkle T, Neuhaus G (1999) Characterisation of a novel gene family of putative cyclic nucleotide- and calmodulin-regulated ion channels in Arabidopsis thaliana. Plant J 18:97–104

    Article  PubMed  CAS  Google Scholar 

  34. Arzai T, Kaplan B, Fromm H (2000) A high-affinity calmodulin-binding site in a tobacco plasma-membrane channel protein coincides with a characteristic element of cyclic nucleotide-binding domains. Plant Mol Biol 42:591–601

    Article  Google Scholar 

  35. Talke IN, Blaudez D, Maathuis FJM et al (2003) CNGCs: prime targets of plant cyclic nucleotide signalling? Trends Plant Sci 8:286–293

    Article  PubMed  CAS  Google Scholar 

  36. Lemtiri-Chlieh F, Berkowitz GA (2004) Cyclic adenosine monophosphate regulates calcium channels in the plasma membrane of Arabidopsis leaf guard and mesophyll cells. J Biol Chem 279:35306–35312

    Article  PubMed  CAS  Google Scholar 

  37. Clough SJ, Fengler KA, Yu IC et al (2000) The Arabidopsisdnd1 “defense, no death” gene encodes a mutated cyclic nucleotide-gated ion channel. Proc Natl Acad Sci U S A 97: 9323–9328

    Article  PubMed  CAS  Google Scholar 

  38. Balagué C, Lin BQ, Alcon C et al (2003) HLM1, an essential signaling component in the hypersensitive response, is a member of the cyclic nucleotide-gated channel ion channel family. Plant Cell 15:365–379

    Article  PubMed  Google Scholar 

  39. Yoshioka K, Moeder W, Kang HG et al (2006) The chimeric Arabidopsis CYCLIC NUCLEOTIDE-GATED ION CHANNEL11/12 activates multiple pathogen resistance responses. Plant Cell 18:747–763

    Article  PubMed  CAS  Google Scholar 

  40. Frietsch S, Wang YF, Sladek C et al (2007) A cyclic nucleotide-gated channel is essential for polarized tip growth of pollen. Proc Natl Acad Sci U S A 104:14531–14536

    Article  PubMed  CAS  Google Scholar 

  41. Chang F, Yan A, Zhao LN et al (2007) A putative calcium-permeable cyclic nucleotide-gated channel, CNGC18, regulates polarized pollen tube growth. J Integr Plant Biol 49:1261–1270

    Article  CAS  Google Scholar 

  42. Rudd JJ, Franklin-Tong V (2001) Unravelling response-specificity in Ca2+ signalling pathways in plant cells. New Phytol 151:7–33

    Article  CAS  Google Scholar 

  43. Swanson SS, Choi WG, Chanoca A et al (2011) In vivo imaging of Ca2+, pH, and reactive oxygen species using fluorescent probes in plants. Annu Rev Plant Biol 62:273–297

    Article  PubMed  CAS  Google Scholar 

  44. Miller AJ, Sanders D (1987) Depletion of cytosolic free calcium induced by photosynthesis. Nature 326:397–400

    Article  CAS  Google Scholar 

  45. Felle H (1988) Auxin causes oscillations of cytosolic free calcium and pH in Zea mays coleoptiles. Planta 174:495–499

    Article  CAS  Google Scholar 

  46. McAinsh MR, Pittman JK (2009) Shaping the calcium signature. New Phytol 181:275–294

    Article  PubMed  CAS  Google Scholar 

  47. Batistič O, Kudla J (2012) Analysis of calcium signaling pathways in plants. Biochim Biophys Acta 1820:1283–1293

    Article  PubMed  Google Scholar 

  48. Williamson RE, Ashley CC (1982) Free Ca2+and cytoplasmic streaming in the alga Chara. Nature 296:647–651

    Article  PubMed  CAS  Google Scholar 

  49. Kiegle E, Moore CA, Haseloff J et al (2000) Cell-type-specific calcium responses to drought, salt and cold in the Arabidopsis root. Plant J 23:267–278

    Article  PubMed  CAS  Google Scholar 

  50. Dodd AN, Jakobsen MK, Baker AJ et al (2006) Time of day modulates low-temperature Ca2+ signals in Arabidopsis. Plant J 48:962–973

    Article  PubMed  CAS  Google Scholar 

  51. Miyawaki A, Llopis J, Heim R et al (1997) Fluorescent indicators for Ca2+ based on green fluorescent proteins and calmodulin. Nature 388:882–887

    Article  PubMed  CAS  Google Scholar 

  52. Emmanouilidou E, Teschemacher AG, Pouli AE et al (1999) Imaging Ca2+ concentration changes at the secretory vesicle surface with a recombinant targeted cameleon. Curr Biol 9:915–918

    Article  PubMed  CAS  Google Scholar 

  53. Krebs M, Held K, Binder A et al (2012) FRET-based genetically encoded sensors allow high-resolution live cell imaging of Ca2+ dynamics. Plant J 69:181–192

    Article  PubMed  CAS  Google Scholar 

  54. Costa A, Drago I, Behera S et al (2010) H2O2 in plant peroxisomes: an in vivo analysis uncovers a Ca2+-dependent scavenging system. Plant J 62:760–772

    Article  PubMed  CAS  Google Scholar 

  55. Iwano M, Entani T, Shiba H et al (2009) Fine-tuning of the cytoplasmic Ca2+ concentration is essential for pollen tube growth. Plant Physiol 150:1322–1334

    Article  PubMed  CAS  Google Scholar 

  56. Allen GJ, Kwak JM, Chu SP et al (1999) Cameleon calcium indicator reports cytoplasmic calcium dynamics in Arabidopsis guard cells. Plant J 19:735–747

    Article  PubMed  CAS  Google Scholar 

  57. Kosuta S, Hazledine S, Sun J et al (2008) Differential and chaotic calcium signatures in the symbiosis signaling pathway of legumes. Proc Natl Acad Sci U S A 105:9823–9828

    Article  PubMed  CAS  Google Scholar 

  58. Chabaud M, Genre A, Sieberer BJ et al (2011) Arbuscular mycorrhizal hyphopodia and germinated spore exudates trigger Ca2+ spiking in the legume and nonlegume root epidermis. New Phytol 189:347–355

    Article  PubMed  CAS  Google Scholar 

  59. Allen GJ, Chu SP, Schumacher K et al (2000) Alteration of stimulus-specific guard cell calcium oscillations and stomatal closing in Arabidopsis det3 mutant. Science 289:2338–2342

    Article  PubMed  CAS  Google Scholar 

  60. Nagai T, Yamada S, Tominaga T et al (2004) Expanded dynamic range of fluorescent indicators for Ca2+ by circularly permuted yellow fluorescent proteins. Proc Natl Acad Sci U S A 101:10554–10559

    Article  PubMed  CAS  Google Scholar 

  61. Tanaka K, Swanson SJ, Gilroy S et al (2010) Extracellular nucleotides elicit cytosolic free calcium oscillations in Arabidopsis. Plant Physiol 154:705–719

    Article  PubMed  CAS  Google Scholar 

  62. Rincon-Zachary M, Teaster ND, Sparks JA et al (2010) Fluorescence resonance energy transfer-sensitized emission of yellow cameleon 3.60 reveals root zone-specific calcium signatures in Arabidopsis in response to aluminum and other trivalent cations. Plant Physiol 152:1442–1458

    Article  PubMed  CAS  Google Scholar 

  63. Monshausen GB, Messerli MA, Gilroy S (2008) Imaging of the Yellow Cameleon 3.6 indicator reveals that elevations in cytosolic Ca2+ follow oscillating increases in growth in root hairs of Arabidopsis. Plant Physiol 147:1690–1698

    Article  PubMed  CAS  Google Scholar 

  64. Monshausen GB, Bibikova TN, Weisenseel MH et al (2009) Ca2+ regulates reactive oxygen species production and pH during mechanosensing in Arabidopsis roots. Plant Cell 21:2341–2356

    Article  PubMed  CAS  Google Scholar 

  65. Jin XC, Wu WH (1999) Involvement of cyclic AMP in ABA- and Ca2+-mediated signal transduction of stomatal regulation in Vicia faba. Plant Cell Physiol 40:1127–1133

    Article  CAS  Google Scholar 

  66. Cousson A (2001) Pharmacological evidence for the implication of both cyclic GMP-dependent and -independent transduction pathways within auxin-induced stomatal opening in Commelina communis(L.). Plant Sci 161:249–258

    Article  PubMed  CAS  Google Scholar 

  67. Cousson A (2003) Pharmacological evidence for a positive influence of the cyclic GMP-independent transduction on the cyclic GMP-mediated Ca2+-dependent pathway within Arabidopsis stomatal opening in response to auxin. Plant Sci 164:759–767

    Article  CAS  Google Scholar 

  68. Pharmawati M, Billington T, Gehring CA (1998) Stomatal guard cell responses to kinetin and natriuretic peptides are cGMP-dependent. Cell Mol Life Sci 54:272–276

    Article  PubMed  CAS  Google Scholar 

  69. Pharmawati M, Maryani MM, Nikolakopoulos T et al (2001) Cyclic GMP modulates stomatal opening induced by natriuretic peptides and immunoreactive analogues. Plant Physiol Biochem 39:385–394

    Article  CAS  Google Scholar 

  70. Kim TH, Bohmer M, Hu HH et al (2010) Guard cell signal transduction network: advances in understanding abscisic acid, CO2, and Ca2+signaling. Annu Rev Plant Biol 61:561–591

    Article  PubMed  CAS  Google Scholar 

  71. Murashige T, Skoog F (1962) A revised medium for rapid growth and bio assays with tobacco tissue cultures. Physiol Plant 15:473–497

    Article  CAS  Google Scholar 

  72. Ng CKY, Carr K, McAinsh MR et al (2001) Drought-induced guard cell signal transduction involves sphingosine-1-phosphate. Nature 410:596–599

    Article  PubMed  CAS  Google Scholar 

  73. Mori C, Murata Y, Yang Y et al (2006) CDPKs CPK6 and CPK3 function in ABA regulation of guard cell S-type anion- and Ca2+-permeable channels and stomatal closure. PLoS Biol 4:10

    Article  Google Scholar 

  74. Yang Y, Costa A, Leonhardt N et al (2008) Isolation of a strong Arabidopsis guard cell promoter and its potential as a research tool. Plant Methods 4:6

    Article  PubMed  Google Scholar 

  75. Trejo CL, Clephan AL, Davies WJ (1995) How do stomata read abscisic acid-signals. Plant Physiol 109:803–811

    PubMed  CAS  Google Scholar 

  76. Schroeder JI, Hagiwara S (1990) Repetitive increases in cytosolic Ca2+ of guard cells by abscisic acid activation of nonselective Ca2+ permeable channels. Proc Natl Acad Sci U S A 87:9305–9309

    Article  PubMed  CAS  Google Scholar 

  77. Lemtiri-Chlieh F, MacRobbie EAC, Webb AAR et al (2003) Inositol hexakisphosphate mobilizes an endomembrane store of calcium in guard cells. Proc Natl Acad Sci U S A 100:10091–10095

    Article  PubMed  CAS  Google Scholar 

  78. Young JJ, Mehta S, Israelsson M et al (2006) CO2 signaling in guard cells: calcium sensitivity response modulation, a Ca2+-independent phase, and CO2 insensitivity of the gca2 mutant. Proc Natl Acad Sci U S A 103:7506–7511

    Article  PubMed  CAS  Google Scholar 

Download references

Acknowledgments

The authors thank the European Community, The Royal Society, the Natural Environment Research Council (UK), the Biotechnology and Biological Sciences Research Council (UK), INTAS, and Belarusian Republican Foundation for Fundamental Research for funding.

Author information

Authors and Affiliations

  1. Lancaster Environment Centre, Lancaster University, Lancaster, UK

    Martin R. McAinsh

  2. Biomedical and Life Sciences Division, Lancaster University, Lancaster, UK

    Stephen K. Roberts

  3. Institute of Biophysics and Cell Engineering, National Academy of Sciences of Belarus, Minsk, Belarus

    Lyudmila V. Dubovskaya

Authors
  1. Martin R. McAinsh
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Stephen K. Roberts
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Lyudmila V. Dubovskaya
    View author publications

    You can also search for this author in PubMed Google Scholar

Editor information

Editors and Affiliations

  1. , Division of Chemical and Life Sciences a, 4700 King Abdullah University of Science, N/A, Thuwal, 23955-6900, Saudi Arabia

    Chris Gehring

Rights and permissions

Reprints and permissions

Copyright information

© 2013 Springer Science+Business Media New York

About this protocol

Cite this protocol

McAinsh, M.R., Roberts, S.K., Dubovskaya, L.V. (2013). Calcium Imaging of the Cyclic Nucleotide Response. In: Gehring, C. (eds) Cyclic Nucleotide Signaling in Plants. Methods in Molecular Biology, vol 1016. Humana Press, Totowa, NJ. https://doi.org/10.1007/978-1-62703-441-8_8

Download citation

  • DOI: https://doi.org/10.1007/978-1-62703-441-8_8

  • Published:

  • Publisher Name: Humana Press, Totowa, NJ

  • Print ISBN: 978-1-62703-440-1

  • Online ISBN: 978-1-62703-441-8

  • eBook Packages: Springer Protocols

Publish with us

Policies and ethics

Search

Navigation