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Quantitative Analysis of Photobodies

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Phytochromes

Part of the book series: Methods in Molecular Biology ((MIMB,volume 2026))

Abstract

Photobodies are membraneless subnuclear organelles that contain the red and far-red photoreceptors, phytochromes. Photobody biogenesis has been postulated to play important roles in early light signaling events. The size and number of photobodies are highly dynamic in response to the quality and quantity of light and correlated tightly with phytochrome-mediated seedling morphogenesis. Here, we provide a detailed protocol for characterization of the three-dimensional morphology of photobodies, including sample preparation, fluorescence microscopy, and image analysis. Although this method was developed initially for characterizing photobodies, it can be adopted to analyze other membraneless or membrane-bound subcellular organelles.

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References

  1. Chen M, Chory J (2011) Phytochrome signaling mechanisms and the control of plant development. Trends Cell Biol 21:664–671

    Article  CAS  Google Scholar 

  2. Yamaguchi R, Nakamura M, Mochizuki N, Kay SA, Nagatani A (1999) Light-dependent translocation of a phytochrome B-GFP fusion protein to the nucleus in transgenic Arabidopsis. J Cell Biol 145:437–445

    Article  CAS  Google Scholar 

  3. Kircher S, Gil P, Kozma-Bognár L, Fejes E, Speth V, Husselstein-Müller T, Bauer D, Ádám É, Schäfer E, Nagy F (2002) Nucleocytoplasmic partitioning of the plant photoreceptors phytochrome A, B, C, D, and E is regulated differentially by light and exhibits a diurnal rhythm. Plant Cell 14:1541–1555

    Article  CAS  Google Scholar 

  4. Kleiner O, Kircher S, Harter K, Batschauer A (1999) Nuclear localization of the Arabidopsis blue light receptor cryptochrome 2. Plant J 19:289–296

    Article  CAS  Google Scholar 

  5. Yu X, Sayegh R, Maymon M, Warpeha K, Klejnot J, Yang H, Huang J, Lee J, Kaufman L, Lin C (2009) Formation of nuclear bodies of Arabidopsis CRY2 in response to blue light is associated with its blue light–dependent degradation. Plant Cell 21:118–130

    Article  CAS  Google Scholar 

  6. Gu N-N, Zhang Y-C, Yang H-Q (2012) Substitution of a conserved glycine in the PHR domain of Arabidopsis CRYPTOCHROME 1 confers a constitutive light response. Mol Plant 5:85–97

    Article  CAS  Google Scholar 

  7. Lian H-L, He S-B, Zhang Y-C, Zhu D-M, Zhang J-Y, Jia K-P, Sun S-X, Li L, Yang H-Q (2011) Blue-light-dependent interaction of cryptochrome 1 with SPA1 defines a dynamic signaling mechanism. Genes Dev 25:1023–1028

    Article  CAS  Google Scholar 

  8. Liu B, Zuo Z, Liu H, Liu X, Lin C (2011) Arabidopsis cryptochrome 1 interacts with SPA1 to suppress COP1 activity in response to blue light. Genes Dev 25:1029–1034

    Article  CAS  Google Scholar 

  9. Más P, Devlin PF, Panda S, Kay SA (2000) Functional interaction of phytochrome B and cryptochrome 2. Nature 408:207–211

    Article  Google Scholar 

  10. Favory J-J, Stec A, Gruber H et al (2009) Interaction of COP1 and UVR8 regulates UV-B-induced photomorphogenesis and stress acclimation in Arabidopsis. EMBO J 28:591–601

    Article  CAS  Google Scholar 

  11. Van Buskirk EK, Decker PV, Chen M (2012) Photobodies in light signaling. Plant Physiol 158:52–60

    Article  Google Scholar 

  12. Kircher S, Kozma-Bognar L, Kim L, Adam E, Harter K, Schäfer E, Nagy F (1999) Light quality-dependent nuclear import of the plant photoreceptors phytochrome A and B. Plant Cell 11:1445–1456

    CAS  PubMed  PubMed Central  Google Scholar 

  13. Chen M, Schwab R, Chory J (2003) Characterization of the requirements for localization of phytochrome B to nuclear bodies. Proc Natl Acad Sci U S A 100:14493–14498

    Article  CAS  Google Scholar 

  14. Rockwell NC, Su Y-S, Lagarias JC (2006) Phytochrome structure and signaling mechanisms. Annu Rev Plant Biol 57:837–858

    Article  CAS  Google Scholar 

  15. Burgie ES, Vierstra RD (2014) Phytochromes: an atomic perspective on photoactivation and signaling. Plant Cell 26:4568–4583

    Article  CAS  Google Scholar 

  16. Jung J-H, Domijan M, Klose C et al (2016) Phytochromes function as thermosensors in Arabidopsis. Science 354:886–889

    Article  CAS  Google Scholar 

  17. Legris M, Klose C, Burgie ES, Rojas CCR, Neme M, Hiltbrunner A, Wigge PA, Schäfer E, Vierstra RD, Casal JJ (2016) Phytochrome B integrates light and temperature signals in Arabidopsis. Science 354:897–900

    Article  CAS  Google Scholar 

  18. Klose C, Venezia F, Hussong A, Kircher S, Schäfer E, Fleck C (2015) Systematic analysis of how phytochrome B dimerization determines its specificity. Nat Plants 1:15090

    Article  CAS  Google Scholar 

  19. Chen M, Galvão RM, Li M, Burger B, Bugea J, Bolado J, Chory J (2010) Arabidopsis HEMERA/pTAC12 initiates photomorphogenesis by phytochromes. Cell 141:1230–1240

    Article  CAS  Google Scholar 

  20. Galvão RM, Li M, Kothadia SM, Haskel JD, Decker PV, Van Buskirk EK, Chen M (2012) Photoactivated phytochromes interact with HEMERA and promote its accumulation to establish photomorphogenesis in Arabidopsis. Genes Dev 26:1851–1863

    Article  Google Scholar 

  21. Huang H, Yoo CY, Bindbeutel R, Goldsworthy J, Tielking A, Alvarez S, Naldrett MJ, Evans BS, Chen M, Nusinow DA (2016) PCH1 integrates circadian and light-signaling pathways to control photoperiod-responsive growth in Arabidopsis. elife 5:e13292

    Article  Google Scholar 

  22. Van Buskirk EK, Reddy AK, Nagatani A, Chen M (2014) Photobody localization of phytochrome B is tightly correlated with prolonged and light-dependent inhibition of hypocotyl elongation in the dark. Plant Physiol 165:595–607

    Article  Google Scholar 

  23. Qiu Y, Li M, Pasoreck EK et al (2015) HEMERA couples the proteolysis and transcriptional activity of PHYTOCHROME INTERACTING FACTORs in Arabidopsis photomorphogenesis. Plant Cell 27:1409–1427

    Article  CAS  Google Scholar 

  24. Qiu Y, Pasoreck EK, Reddy AK, Nagatani A, Ma W, Chory J, Chen M (2017) Mechanism of early light signaling by the carboxy-terminal output module of Arabidopsis phytochrome B. Nat Commun 8:1905

    Article  Google Scholar 

  25. Bauer D, Viczián A, Kircher S et al (2004) CONSTITUTIVE PHOTOMORPHOGENESIS 1 and multiple photoreceptors control degradation of PHYTOCHROME INTERACTING FACTOR 3, a transcription factor required for light signaling in Arabidopsis. Plant Cell 16:1433–1445

    Article  CAS  Google Scholar 

  26. Al-Sady B, Ni W, Kircher S, Schäfer E, Quail PH (2006) Photoactivated phytochrome induces rapid PIF3 phosphorylation prior to proteasome-mediated degradation. Mol Cell 23:439–446

    Article  CAS  Google Scholar 

  27. Gendreau E, Traas J, Desnos T, Grandjean O, Caboche M, Höfte H (1997) Cellular basis of hypocotyl growth in Arabidopsis thaliana. Plant Physiol 114:295–305

    Article  CAS  Google Scholar 

  28. Wang Z-Y, Nakano T, Gendron J et al (2002) Nuclear-localized BZR1 mediates brassinosteroid-induced growth and feedback suppression of brassinosteroid biosynthesis. Dev Cell 2:505–513

    Article  CAS  Google Scholar 

  29. Nybo K (2012) GFP imaging in fixed cells. BioTechniques 52:359–360

    CAS  PubMed  Google Scholar 

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Acknowledgments

This work was supported by National Institute of Health grant R01GM087388 to M.C.

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Authors and Affiliations

  1. Department of Botany and Plant Sciences, Institute for Integrative Genome Biology, University of California, Riverside, CA, USA

    Chan Yul Yoo, Desiree Williams & Meng Chen

Authors
  1. Chan Yul Yoo
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  2. Desiree Williams
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  3. Meng Chen
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Corresponding author

Correspondence to Meng Chen .

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Editors and Affiliations

  1. Faculty of Biology, Institute of Biology II and Signalling Research Centres BIOSS and CIBSS, University of Freiburg, Freiburg, Germany

    Andreas Hiltbrunner

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Yoo, C.Y., Williams, D., Chen, M. (2019). Quantitative Analysis of Photobodies. In: Hiltbrunner, A. (eds) Phytochromes. Methods in Molecular Biology, vol 2026. Humana, New York, NY. https://doi.org/10.1007/978-1-4939-9612-4_10

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  • DOI: https://doi.org/10.1007/978-1-4939-9612-4_10

  • Published:

  • Publisher Name: Humana, New York, NY

  • Print ISBN: 978-1-4939-9611-7

  • Online ISBN: 978-1-4939-9612-4

  • eBook Packages: Springer Protocols

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