Advertisement

Plant Molecular Biology

, Volume 63, Issue 4, pp 491–503 | Cite as

Different phosphorylation mechanisms are involved in the activation of sucrose non-fermenting 1 related protein kinases 2 by osmotic stresses and abscisic acid

  • Marie Boudsocq
  • Marie-Jo Droillard
  • Hélène Barbier-Brygoo
  • Christiane LaurièreEmail author
Article

Abstract

In Arabidopsis cell suspension, hyperosmotic stresses (mannitol and NaCl) were previously shown to activate nine sucrose non-fermenting 1 related protein kinases 2 (SnRK2s) whereas only five of them were also activated by abscisic acid (ABA) treatment. Here, the possible activation by phosphorylation/dephosphorylation of each kinase was investigated by studying their phosphorylation state after osmotic stress, using the Pro-Q Diamond, a specific dye for phosphoproteins. All the activated kinases were phosphorylated after osmotic stress but the induced phosphorylation changes were clearly different depending on the kinase. In addition, the increase of the global phosphorylation level induced by ABA application was lower, suggesting that different mechanisms may be involved in SnRK2 activation by hyperosmolarity and ABA. On the other hand, SnRK2 kinases remain activated by hyperosmotic stress in ABA-deficient and ABA-insensitive mutants, indicating that SnRK2 osmotic activation is independent of ABA. Moreover, using a mutant form of SnRK2s, a specific serine in the activation loop was shown to be phosphorylated after stress treatments and essential for activity and/or activation. Finally, SnRK2 activity was sensitive to staurosporine, whereas SnRK2 activation by hyperosmolarity or ABA was not, indicating that SnRK2 activation by phosphorylation is mediated by an upstream staurosporine-insensitive kinase, in both signalling pathways. All together, these results indicate that different phosphorylation mechanisms and at least three signalling pathways are involved in the activation of SnRK2 proteins in response to osmotic stress and ABA.

Keywords

Abscisic acid Arabidopsis thaliana Osmotic stresses Phosphorylation Pro-Q® Diamond SnRK2 

Abbreviations

ABA

abscisic acid

MBP

myelin basic protein

SnRK2

sucrose non-fermenting 1 related protein kinase 2

Notes

Acknowledgements

We thank Dr. Pascale Bertrand and Dr. Yannick Saintigny for kind introduction to the Typhoon imaging system. We also thank Dr. Sylvain Merlot and Dr. Helen North for kindly providing abi1-1 and aba1-3 seeds, respectively.

References

  1. Abbasi F, Onodera H, Toki S, Tanaka H, Komatsu S (2004) OsCDPK13, a calcium-dependent protein kinase gene from rice, is induced by cold and gibberellin in rice leaf sheath. Plant Mol Biol 55:541–552PubMedCrossRefGoogle Scholar
  2. Blom N, Sicheritz-Ponten P, Gupta R, Gammeltoft S, Brunak S (2004) Prediction of post-translational glycosylation and phosphorylation of proteins from the amino acid sequence. Proteomics 4:1633–1649PubMedCrossRefGoogle Scholar
  3. Boudsocq M, Barbier-Brygoo H, Laurière C (2004) Identification of nine sucrose nonfermenting 1-related protein kinases 2 activated by hyperosmotic and saline stresses in Arabidopsis thaliana. J Biol Chem 279:41758–41766PubMedCrossRefGoogle Scholar
  4. Boudsocq M, Laurière C (2005) Osmotic signaling in Plants. Multiple pathways mediated by emerging kinase families. Plant Physiol 138:1185–1194PubMedCrossRefGoogle Scholar
  5. Bradford MM (1976) A rapid and sensitive method for the quantification of μg quantities of proteins utilising the principle of protein-dye binding. Anal Biochem 72:248–254PubMedCrossRefGoogle Scholar
  6. Cardinale F, Meskiene I, Ouaked F, Hirt H (2002) Convergence and divergence of stress-induced mitogen-activated protein kinase signaling pathways at the level of two distinct mitogen-activated protein kinase kinases. Plant Cell 14:703–711PubMedGoogle Scholar
  7. Droillard MJ, Boudsocq M, Barbier-Brygoo H, Laurière C (2002) Different protein kinase families are activated by osmotic stresses in Arabidopsis thaliana cell suspensions. Involvement of the MAP kinases AtMPK3 and AtMPK6. FEBS Lett 527:43–50PubMedCrossRefGoogle Scholar
  8. Furdui CM, Lew ED, Schlessinger J, Anderson KS (2006) Autophosphorylation of FGFR1 kinase is mediated by a sequential and precisely ordered reaction. Mol Cell 21:711–717PubMedCrossRefGoogle Scholar
  9. Furihata T, Maruyama K, Fujita Y, Umezawa T, Yoshida R, Shinozaki K, Yamaguchi-Shinozaki K (2006) Abscisic acid-dependent multisite phosphorylation regulates the activity of a transcription activator AREB1. Proc Natl Acad Sci USA 103:1988–1993PubMedCrossRefGoogle Scholar
  10. Guo Y, Halfter U, Ishitani M, Zhu JK (2001) Molecular characterization of functional domains in the protein kinase SOS2 that is required for plant salt tolerance. Plant Cell 13:1383–1399PubMedCrossRefGoogle Scholar
  11. Halfter U, Ishitani M, Zhu JK (2000) The Arabidopsis SOS2 protein kinase physically interacts with and is activated by the calcium-binding protein SOS3. Proc Natl Acad Sci USA 97:3735–3740PubMedCrossRefGoogle Scholar
  12. Harper JF, Breton G, Harmon A (2004) Decoding Ca2+ signals through plant protein kinases. Annu Rev Plant Biol 55:263–288PubMedCrossRefGoogle Scholar
  13. Hoyos ME, Zhang SQ (2000) Calcium-independent activation of salicylic acid-induced protein kinase and a 40-kilodalton protein kinase by hyperosmotic stress. Plant Physiol 122:1355–1363PubMedCrossRefGoogle Scholar
  14. Hrabak EM, Chan CWM, Gribskov M, Harper JF, Choi JH, Halford N, Kudla J, Luan S, Nimmo HG, Sussman MR, Thomas M, Walker Simmons K, Zhu JK, Harmon AC (2003) The Arabidopsis CDPK-SnRK superfamily of protein kinases. Plant Physiol 132:666–680PubMedCrossRefGoogle Scholar
  15. Johnson LN, Noble MEM, Owen DJ (1996) Active and inactive protein kinases: structural basis for regulation. Cell 85:149–158PubMedCrossRefGoogle Scholar
  16. Kanhonou R, Serrano R, Palau RR (2001) A catalytic subunit of the sugar beet protein kinase CK2 is induced by salt stress and increases NaCl tolerance in Saccharomyces cerevisiae. Plant Mol Biol 47:571–579PubMedCrossRefGoogle Scholar
  17. Kelner A, Pekala I, Kaczanowski S, Muszynska G, Hardie DG, Dobrowolska G (2004) Biochemical characterization of the tobacco 42-kD protein kinase activated by osmotic stress. Plant Physiol 136:3255–3265PubMedCrossRefGoogle Scholar
  18. Kiegerl S, Cardinale F, Siligan C, Gross A, Baudouin E, Liwosz A, Eklof S, Till S, Bogre L, Hirt H, Meskiene I (2000) SIMKK, a mitogen-activated protein kinase (MAPK) kinase, is a specific activator of the salt stress-induced MAPK, SIMK. Plant Cell 12:2247–2258PubMedCrossRefGoogle Scholar
  19. Kobayashi Y, Murata M, Minami H, Yamamoto S, Kagaya Y, Hobo T, Yamamoto A, Hattori T (2005) Abscisic acid-activated SnRK2 protein kinases function in the gene-regulation pathway of ABA signal transduction by phosphorylating ABA response element-binding factors. Plant J 44:939–949PubMedCrossRefGoogle Scholar
  20. Kobayashi Y, Yamamoto S, Minami H, Kagaya Y, Hattori T (2004) Differential activation of the rice sucrose nonfermenting1-related protein kinase2 family by hyperosmotic stress and abscisic acid. Plant Cell 16:1163–1177PubMedCrossRefGoogle Scholar
  21. Leung J, Merlot S, Giraudat J (1997) The Arabidopsis Abscisic acid-insensitive 2 (ABI2) and ABI1 genes encode homologous protein phosphatases 2C involved in abscisic acid signal transduction. Plant Cell 9:759–771PubMedCrossRefGoogle Scholar
  22. Li J, Assmann SM (1996) An abscisic acid-activated and calcium-independent protein kinase from guard cells of fava bean. Plant Cell 8:2359–2368PubMedCrossRefGoogle Scholar
  23. Li J, Wang XQ, Watson MB, Assmann SM (2000) Regulation of abscisic acid-induced stomatal closure and anion channels by guard cell AAPK kinase. Science 287:300–303PubMedCrossRefGoogle Scholar
  24. Martin ML, Busconi L (2001) A rice membrane-bound calcium-dependent protein kinase is activated in response to low temperature. Plant Physiol 125:1442–1449PubMedCrossRefGoogle Scholar
  25. Meggio F, Donella Deana A, Ruzzene M, Brunati AM, Cesaro L, Guerra B, Meyer T, Mett H, Fabbro D, Furet P, Dobrowolska G, Pinna LA (1995) Different susceptibility of protein kinases to staurosporine inhibition. Kinetic studies and molecular bases for the resistance of protein kinase CK2. Eur. J Biochem 234:317–322Google Scholar
  26. Mikolajczyk M, Awotunde OS, Muszynska G, Klessig DF, Dobrowolska G (2000) Osmotic stress induces rapid activation of a salicylic acid-induced protein kinase and a homolog of protein kinase ASK1 in tobacco cells. Plant Cell 12:165–178PubMedCrossRefGoogle Scholar
  27. Murashige T, Skoog F (1962) A revised medium fot rapid growth and bioassays with tobacco tissue cultures. Physiol Plant 15:473–497CrossRefGoogle Scholar
  28. Mustilli AC, Merlot S, Vavasseur A, Fenzi F, Giraudat J (2002) Arabidopsis OST1 protein kinase mediates the regulation of stomatal aperture by abscisic acid and acts upstream of reactive oxygen species production. Plant Cell 14:3089–3099PubMedCrossRefGoogle Scholar
  29. Riera M, Figueras M, Lopez C, Goday A, Pagès M (2004) Protein kinase CK2 modulates developmental functions of the abscisic acid responsive protein Rab17 from maize. Proc Natl Acad Sci USA 101:9879–9884PubMedCrossRefGoogle Scholar
  30. Rock CD, Zeevaart JAD (1991) The aba mutant of Arabidopsis thaliana is impaired in epoxy-carotenoid biosynthesis. Proc Natl Acad Sci USA 88:7496–7499PubMedCrossRefGoogle Scholar
  31. Romeis T, Ludwig AA, Martin R, Jones JDG (2001) Calcium-dependent protein kinases play an essential role in a plant defence response. EMBO J 20:5556–5567PubMedCrossRefGoogle Scholar
  32. Romeis T, Piedras P, Jones JDG (2000) Resistance gene-dependent activation of a calcium-dependent protein kinase in the plant defense response. Plant Cell 12:803–815PubMedCrossRefGoogle Scholar
  33. Sicheri F, Kuriyan J (1997) Structures of Scr-family tyrosine kinases. Curr Opin Struct Biol 7:777–785PubMedCrossRefGoogle Scholar
  34. Smits VAJ, Medema RH (2001) Checking out the G2/M transition. Biochim Biophys Acta 1519: 1–12PubMedGoogle Scholar
  35. Steinberg TH, Agnew BJ, Gee KR, Leung WY, Goodman T, Schulenberg B, Hendrickson J, Beechem JM, Haugland RP, Patton WF (2003) Global quantitative phosphoprotein analysis using multiplexed proteomics technology. Proteomics 3:1128–1144PubMedCrossRefGoogle Scholar
  36. Teige M, Scheikl E, Eulgem T, Doczi F, Ichimura K, Shinozaki K, Dangl JL, Hirt H (2004) The MKK2 pathway mediates cold and salt stress signaling in Arabidopsis. Mol Cell 15:141–152PubMedCrossRefGoogle Scholar
  37. Umezawa T, Yoshida R, Maruyama K, Yamaguchi-Shinozaki K, Shinozaki K (2004) SRK2C, a SNF1-related protein kinase 2, improves drought tolerance by controlling stress-responsive gene expression in Arabidopsis thaliana. Proc Natl Acad Sci USA 101:17306–17311PubMedCrossRefGoogle Scholar
  38. Yoshida R, Hobo T, Ichimura K, Mizoguchi T, Takahashi F, Aronso J, Ecker JR, Shinozaki K (2002) ABA-activated SnRK2 protein kinase is required for dehydration stress signaling in Arabidopsis. Plant Cell Physiol 43:1473–1483PubMedCrossRefGoogle Scholar
  39. Yoshida R, Umezawa T, Mizoguchi T, Takahashi S, Takahashi F, Shinozaki K (2006) The regulatory domain of SRK2E/OST1/SnRK2.6 interacts with ABI1 and integrates abscisic acid (ABA) and osmotic stress signals controlling stomatal closure in Arabidopsis. J Biol Chem 281:5310–5318PubMedCrossRefGoogle Scholar
  40. Zhang S, Du H, Klessig DF (1998) Activation of the tobacco SIP kinase by both a cell wall-derived carbohydrate elicitor and purified proteinaceous elicitins from Phytophthora spp. Plant Cell 10:435–449PubMedCrossRefGoogle Scholar

Copyright information

© Springer Science+Business Media B.V. 2006

Authors and Affiliations

  • Marie Boudsocq
    • 1
    • 2
    • 3
  • Marie-Jo Droillard
    • 1
  • Hélène Barbier-Brygoo
    • 1
  • Christiane Laurière
    • 1
    Email author
  1. 1.Institut des Sciences du Végétal, UPR 2355, CNRSGif sur Yvette CedexFrance
  2. 2.Department of GeneticsHarvard Medical SchoolBostonUSA
  3. 3.Department of Molecular BiologyMassachusetts General HospitalBostonUSA

Personalised recommendations