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Live Cell Imaging of the Cytoskeleton and Cell Wall Enzymes in Plant Cells

  • Arun SampathkumarEmail author
  • Raymond Wightman
Protocol
Part of the Methods in Molecular Biology book series (MIMB, volume 1242)

Abstract

The use of live imaging techniques to visualize the dynamic changes and interactions within plant cells has given us detailed information on the function and organization of the cytoskeleton and cell wall associated proteins. This information has grown with the constant improvement in imaging hardware and molecular tools. In this chapter, we describe the procedure for the preparation and live visualization of fluorescent protein fusions associated with the cytoskeleton and the cell wall in Arabidopsis.

Key words

Live cell imaging Microtubules Actin filaments Cell wall Point scanning confocal microscopy Spinning disc microscopy Hypocotyls Xylem vessels 

Notes

Acknowledgements

We would like to thank Dr. Staffan Persson at the Max Planck Institute of Molecular Plant Physiology for his comments on the chapter.

References

  1. 1.
    Hepler PK et al (1993) Cytoskeletal dynamics in living plant cells. Cell Biol Int 17(2):127–142CrossRefGoogle Scholar
  2. 2.
    Kost B, Spielhofer P, Chua NH (1998) A GFP-mouse talin fusion protein labels plant actin filaments in vivo and visualizes the actin cytoskeleton in growing pollen tubes. Plant J 16(3):393–401PubMedCrossRefGoogle Scholar
  3. 3.
    Marc J et al (1998) A GFP-MAP4 reporter gene for visualizing cortical microtubule rearrangements in living epidermal cells. Plant Cell 10(11):1927–1940PubMedCentralPubMedGoogle Scholar
  4. 4.
    Ketelaar T, Anthony RG, Hussey PJ (2004) Green fluorescent protein-mTalin causes defects in actin organization and cell expansion in Arabidopsis and inhibits actin depolymerizing factor’s actin depolymerizing activity in vitro. Plant Physiol 136(4):3990–3998PubMedCentralPubMedCrossRefGoogle Scholar
  5. 5.
    Sheahan MB et al (2004) A green fluorescent protein fusion to actin-binding domain 2 of Arabidopsis fimbrin highlights new features of a dynamic actin cytoskeleton in live plant cells. Plant Physiol 136(4):3968–3978PubMedCentralPubMedCrossRefGoogle Scholar
  6. 6.
    Ueda K, Matsuyama T, Hashimoto T (1999) Visualization of microtubules in living cells of transgenic Arabidopsis thaliana. Protoplasma 206(1–3):201–206CrossRefGoogle Scholar
  7. 7.
    Ledbetter MC, Porter KR (1963) A “microtubule” in plant cell fine structure. J Cell Biol 19(1):239–250PubMedCentralPubMedCrossRefGoogle Scholar
  8. 8.
    Mueller SC, Brown RM (1980) Evidence for an intramembrane component associated with a cellulose microfibril-synthesizing complex in higher plants. J Cell Biol 84(2):315–326PubMedCrossRefGoogle Scholar
  9. 9.
    Delmer DP, Amor Y (1995) Cellulose biosynthesis. Plant Cell 7(7):987–1000PubMedCentralPubMedCrossRefGoogle Scholar
  10. 10.
    Paredez AR, Somerville CR, Ehrhardt DW (2006) Visualization of cellulose synthase demonstrates functional association with microtubules. Science 312(5779):1491–1495PubMedCrossRefGoogle Scholar
  11. 11.
    Bischoff V et al (2011) Phytochrome regulation of cellulose synthesis in Arabidopsis. Curr Biol 21(21):1822–1827PubMedCrossRefGoogle Scholar
  12. 12.
    Desprez T et al (2007) Organization of cellulose synthase complexes involved in primary cell wall synthesis in Arabidopsis thaliana. Proc Natl Acad Sci U S A 104(39):15572–15577PubMedCentralPubMedCrossRefGoogle Scholar
  13. 13.
    Staiger CJ et al (2009) Actin filament dynamics are dominated by rapid growth and severing activity in the Arabidopsis cortical array. J Cell Biol 184(2):269–280PubMedCentralPubMedCrossRefGoogle Scholar
  14. 14.
    Shaw SL, Kamyar R, Ehrhardt DW (2003) Sustained microtubule treadmilling in Arabidopsis cortical arrays. Science 300(5626):1715–1718PubMedCrossRefGoogle Scholar
  15. 15.
    Sampathkumar A et al (2011) Live cell imaging reveals structural associations between the actin and microtubule cytoskeleton in Arabidopsis. Plant Cell 23(6):2302–2313PubMedCentralPubMedCrossRefGoogle Scholar
  16. 16.
    Sampathkumar A et al (2013) Patterning and lifetime of plasma membrane-localized cellulose synthase is dependent on actin organization in Arabidopsis interphase cells. Plant Physiol 162(2):675–688PubMedCentralPubMedCrossRefGoogle Scholar
  17. 17.
    Wightman R, Turner SR (2008) The roles of the cytoskeleton during cellulose deposition at the secondary cell wall. Plant J 54(5):794–805PubMedCrossRefGoogle Scholar
  18. 18.
    Wightman R, Marshall R, Turner SR (2009) A cellulose synthase-containing compartment moves rapidly beneath sites of secondary wall synthesis. Plant Cell Physiol 50(3):584–594PubMedCrossRefGoogle Scholar
  19. 19.
    Young B et al (2010) pH-sensitivity of YFP provides an intracellular indicator of programmed cell death. Plant Methods 6:27PubMedCentralPubMedCrossRefGoogle Scholar

Copyright information

© Springer Science+Business Media New York 2015

Authors and Affiliations

  1. 1.Division of Biology and Biological Engineering 156-29California Institute of TechnologyPasadenaUSA
  2. 2.Sainsbury LaboratoryUniversity of CambridgeCambridgeUK

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