Automatic Chloroplast Movement Analysis

Protocol
Part of the Methods in Molecular Biology book series (MIMB, volume 1398)

Abstract

In response to low or high intensities of light, the chloroplasts in the mesophyll cells of the leaf are able to increase or decrease their exposure to light by accumulating at the upper and lower sides or along the side walls of the cell respectively. This movement, regulated by the phototropin blue light photoreceptors phot1 and phot2, results in a decreased or increased transmission of light through the leaf. This way the plant is able to optimize harvesting of the incoming light or avoid damage caused by excess light. Here we describe a method that indirectly measures the movement of chloroplasts by taking advantage of the resulting change in leaf transmittance. By using a microplate reader, quantitative measurements of chloroplast accumulation or avoidance can be monitored over time, for multiple samples with relatively little hands-on time.

Key words

Phototropin Arabidopsis Chloroplast movements Microplate reader 

Notes

Acknowledgements

We would like to thank Jon Hughes for support and funding H.J. (Hu702/5-3), the members of his lab for valuable suggestions on the experimental setup, and Nora Schorn for proofreading the manuscript.

References

  1. 1.
    Kasahara M, Kagawa T, Oikawa K, Suetsugu N, Miyao M, Wada M (2002) Chloroplast avoidance movement reduces photodamage in plants. Nature 420(6917):829–832. doi: 10.1038/nature01213 CrossRefPubMedGoogle Scholar
  2. 2.
    Trojan A, Gabrys H (1996) Chloroplast distribution in Arabidopsis thaliana (L.) depends on light conditions during growth. Plant Physiol 111(2):419–425PubMedCentralPubMedGoogle Scholar
  3. 3.
    Kong SG, Wada M (2014) Recent advances in understanding the molecular mechanism of chloroplast photorelocation movement. Biochim Biophys Acta 1837(4):522–530. doi: 10.1016/j.bbabio.2013.12.004 CrossRefPubMedGoogle Scholar
  4. 4.
    Kagawa T, Sakai T, Suetsugu N, Oikawa K, Ishiguro S, Kato T, Tabata S, Okada K, Wada M (2001) Arabidopsis NPL1: a phototropin homolog controlling the chloroplast high-light avoidance response. Science 291(5511):2138–2141. doi: 10.1126/science.291.5511.2138 CrossRefPubMedGoogle Scholar
  5. 5.
    Sakai T, Kagawa T, Kasahara M, Swartz TE, Christie JM, Briggs WR, Wada M, Okada K (2001) Arabidopsis nph1 and npl1: blue light receptors that mediate both phototropism and chloroplast relocation. Proc Natl Acad Sci U S A 98(12):6969–6974. doi: 10.1073/pnas.101137598 PubMedCentralCrossRefPubMedGoogle Scholar
  6. 6.
    Wada M (2013) Chloroplast movement. Plant Sci 210:177–182. doi: 10.1016/j.plantsci.2013.05.016 CrossRefPubMedGoogle Scholar
  7. 7.
    Kagawa T, Wada M (2000) Blue light-induced chloroplast relocation in Arabidopsis thaliana as analyzed by microbeam irradiation. Plant Cell Physiol 41(1):84–93CrossRefPubMedGoogle Scholar
  8. 8.
    Inoue Y, Shibata K (1973) Light-induced chloroplast rearrangements and their action spectra as measured by absorption spectrophotometry. Planta 114(4):341–358. doi: 10.1007/BF00387947 CrossRefPubMedGoogle Scholar
  9. 9.
    Berg R, Koniger M, Schjeide BM, Dikmak G, Kohler S, Harris GC (2006) A simple low-cost microcontroller-based photometric instrument for monitoring chloroplast movement. Photosynth Res 87(3):303–311. doi: 10.1007/s11120-005-9012-1 CrossRefPubMedGoogle Scholar
  10. 10.
    DeBlasio SL, Mullen JL, Luesse DR, Hangarter RP (2003) Phytochrome modulation of blue light-induced chloroplast movements in Arabidopsis. Plant Physiol 133(4):1471–1479. doi: 10.1104/pp. 103.029116 PubMedCentralCrossRefPubMedGoogle Scholar
  11. 11.
    Kodama Y, Suetsugu N, Kong SG, Wada M (2010) Two interacting coiled-coil proteins, WEB1 and PMI2, maintain the chloroplast photorelocation movement velocity in Arabidopsis. Proc Natl Acad Sci U S A 107(45):19591–19596. doi: 10.1073/pnas.1007836107 PubMedCentralCrossRefPubMedGoogle Scholar
  12. 12.
    Kadota A, Sato Y, Wada M (2000) Intracellular chloroplast photorelocation in the moss Physcomitrella patens is mediated by phytochrome as well as by a blue-light receptor. Planta 210(6):932–937Google Scholar
  13. 13.
    Kawai H, Kanegae T, Christensen S, Kiyosue T, Sato Y, Imaizumi T, Kadota A, Wada M (2003) Responses of ferns to red light are mediated by an unconventional photoreceptor. Nature 421(6920):287–290. doi: 10.1038/nature01310 CrossRefPubMedGoogle Scholar

Copyright information

© Springer Science+Business Media New York 2016

Authors and Affiliations

  1. 1.Department of Plant PhysiologyJustus Liebig UniversityGiessenGermany
  2. 2.Institute of Biology/Applied Genetics, Dahlem Centre of Plant Sciences (DCPS)Freie Universität BerlinBerlin-DahlemGermany

Personalised recommendations