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Bimolecular Fluorescent Complementation (BiFC) by MAP Kinases and MAPK Phosphatases

  • Alois SchweighoferEmail author
  • Volodymyr Shubchynskyy
  • Vaiva Kazanaviciute
  • Armin Djamei
  • Irute Meskiene
Protocol
Part of the Methods in Molecular Biology book series (MIMB, volume 1171)

Abstract

The adaptation of plants to the environment is a key property for survival. Adaptation responses to environmental cues are generated in cells by signaling initiated from cell receptors. Signal transduction is based on protein phosphorylation that is employed in mitogen-activated protein kinase (MAPK) cascades to integrate signals from receptors to cellular responses. MAPK activity is determined by phosphorylation of amino acid residues within the kinase activation loop and their dephosphorylation by phosphatases is essential to control signal duration and intensity.

Monitoring protein–protein interactions (PPIs) of MAPKs with MAPK phosphatases in vivo provides valuable information about specificity and intracellular localization of the protein complex. Here, we report studying PPIs between Arabidopsis MAPKs and PP2C-type MAPK phosphatases using bimolecular fluorescent complementation (BiFC) in suspension cell protoplasts. The interactions of the MAPKs MPK3, MKP4 and MPK6 with the phosphatases AP2C1 and AP2C3 have been tested.

Key words

Arabidopsis BiFC Localisation PP2C MAPK phosphatase Protein interaction 

Notes

Acknowledgements

We thank Dierk Scheel and Thomas Kroj for sharing the protoplast isolation method for parsley and Andrej Belokurov for cultivation of suspension culture cells. This work has received funding from the Lithuanian-Swiss cooperation program to reduce economic and social disparities within the enlarged European Union under project agreement No CH-3-ŠMM-01/10 and from the Austrian Science Fund (FWF) I255, L687 and W1220-B09.

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Copyright information

© Springer Science+Business Media New York 2014

Authors and Affiliations

  • Alois Schweighofer
    • 1
    • 2
    Email author
  • Volodymyr Shubchynskyy
    • 1
  • Vaiva Kazanaviciute
    • 2
  • Armin Djamei
    • 3
  • Irute Meskiene
    • 1
    • 2
  1. 1.Max F. Perutz LaboratoriesUniversity and Medical University of ViennaViennaAustria
  2. 2.Institute of Biotechnology (IBT)University of VilniusVilniusLithuania
  3. 3.Gregor Mendel Institute of Molecular Plant Biology (GMI)ViennaAustria

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