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New Imaging Tools to Analyze Mitochondrial Morphology in Caenorhabditis elegans

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Mitochondria

Part of the book series: Methods in Molecular Biology ((MIMB,volume 1567))

Abstract

Mitochondria are highly dynamic organelles that constantly fuse and divide. This process is essential as several neurodegenerative diseases have been associated with defects in mitochondrial fusion or fission. Several tools have been developed over the years to visualize mitochondria in organisms such as Caenorhabditis elegans. Combining these tools with the powerful genetics of C. elegans has led to the discovery of new regulators of mitochondrial morphology. In this chapter, we present additional tools to further characterize mitochondrial morphology as well as regulators of mitochondrial morphology. Specifically, we introduce a photoactivatable mitoGFP (PAmitoGFP) that allows to investigate the connectivity of complex mitochondrial networks. In addition, we describe an immunostaining protocol that enables localization studies of these newly identified regulators of mitochondrial morphology.

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References

  1. Mishra P, Chan DC (2014) Mitochondrial dynamics and inheritance during cell division, development and disease. Nat Rev Mol Cell Biol 15(10):634–646. doi:10.1038/nrm3877

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  2. Ichishita R, Tanaka K, Sugiura Y, Sayano T, Mihara K, Oka T (2008) An RNAi screen for mitochondrial proteins required to maintain the morphology of the organelle in Caenorhabditis elegans. J Biochem 143(4):449–454. doi:10.1093/jb/mvm245

    Article  CAS  PubMed  Google Scholar 

  3. Kanazawa T, Zappaterra MD, Hasegawa A, Wright AP, Newman-Smith ED, Buttle KF, McDonald K, Mannella CA, van der Bliek AM (2008) The C. elegans Opa1 homologue EAT-3 is essential for resistance to free radicals. PLoS Genet 4(2):e1000022. doi:10.1371/journal.pgen.1000022

    Article  PubMed  PubMed Central  Google Scholar 

  4. Labrousse AM, Zappaterra MD, Rube DA, van der Bliek AM (1999) C. elegans dynamin-related protein DRP-1 controls severing of the mitochondrial outer membrane. Mol Cell 4(5):815–826

    Article  CAS  PubMed  Google Scholar 

  5. Rawson RL, Yam L, Weimer RM, Bend EG, Hartwieg E, Horvitz HR, Clark SG, Jorgensen EM (2014) Axons degenerate in the absence of mitochondria in C. elegans. Curr Biol : CB 24(7):760–765. doi:10.1016/j.cub.2014.02.025

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  6. Rolland SG (2014) How to analyze mitochondrial morphology in healthy cells and apoptotic cells in Caenorhabditis elegans. Methods Enzymol 544:75–98. doi:10.1016/B978-0-12-417158-9.00004-2

    Article  CAS  PubMed  Google Scholar 

  7. Kuznetsov AV, Hermann M, Saks V, Hengster P, Margreiter R (2009) The cell-type specificity of mitochondrial dynamics. Int J Biochem Cell Biol 41(10):1928–1939. doi:10.1016/j.biocel.2009.03.007

    Article  CAS  PubMed  Google Scholar 

  8. Stiernagle T (2006) Maintenance of C. elegans. WormBook : the online review of C elegans biology:1–11. doi:10.1895/wormbook.1.101.1

  9. Rolland SG, Motori E, Memar N, Hench J, Frank S, Winklhofer KF, Conradt B (2013) Impaired complex IV activity in response to loss of LRPPRC function can be compensated by mitochondrial hyperfusion. Proc Natl Acad Sci U S A 110(32):E2967–E2976. doi:10.1073/pnas.1303872110

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  10. Schermelleh L, Heintzmann R, Leonhardt H (2010) A guide to super-resolution fluorescence microscopy. J Cell Biol 190(2):165–175. doi:10.1083/jcb.201002018

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  11. Duerr JS (2006) Immunohistochemistry. WormBook : the online review of C elegans biology:1–61. doi:10.1895/wormbook.1.105.1

  12. Baddeley D, Chagin VO, Schermelleh L, Martin S, Pombo A, Carlton PM, Gahl A, Domaing P, Birk U, Leonhardt H, Cremer C, Cardoso MC (2010) Measurement of replication structures at the nanometer scale using super-resolution light microscopy. Nucleic Acids Res 38(2):e8. doi:10.1093/nar/gkp901

    Article  CAS  PubMed  Google Scholar 

  13. Pourkarimi E, Greiss S, Gartner A (2012) Evidence that CED-9/Bcl2 and CED-4/Apaf-1 localization is not consistent with the current model for C. elegans apoptosis induction. Cell Death Differ 19(3):406–415. doi:10.1038/cdd.2011.104

    Article  CAS  PubMed  Google Scholar 

  14. Timmons L, Court DL, Fire A (2001) Ingestion of bacterially expressed dsRNAs can produce specific and potent genetic interference in Caenorhabditis elegans. Gene 263(1-2):103–112

    Article  CAS  PubMed  Google Scholar 

  15. Kamath RS, Ahringer J (2003) Genome-wide RNAi screening in Caenorhabditis elegans. Methods 30(4):313–321

    Article  CAS  PubMed  Google Scholar 

  16. Kim E, Sun L, Gabel CV, Fang-Yen C (2013) Long-term imaging of Caenorhabditis elegans using nanoparticle-mediated immobilization. PLoS One 8(1):e53419. doi:10.1371/journal.pone.0053419

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  17. Mondal S, Koushika SP (2014) Microfluidic devices for imaging trafficking events in vivo using genetic model organisms. Methods Mol Biol 1174:375–396. doi:10.1007/978-1-4939-0944-5_26

    Article  PubMed  Google Scholar 

  18. Labrador L, Barroso C, Lightfoot J, Muller-Reichert T, Flibotte S, Taylor J, Moerman DG, Villeneuve AM, Martinez-Perez E (2013) Chromosome movements promoted by the mitochondrial protein SPD-3 are required for homology search during Caenorhabditis elegans meiosis. PLoS Genet 9(5):e1003497. doi:10.1371/journal.pgen.1003497

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  19. Liu Y, Samuel BS, Breen PC, Ruvkun G (2014) Caenorhabditis elegans pathways that surveil and defend mitochondria. Nature 508(7496):406–410. doi:10.1038/nature13204

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  20. Berman SB, Chen YB, Qi B, McCaffery JM, Rucker EB 3rd, Goebbels S, Nave KA, Arnold BA, Jonas EA, Pineda FJ, Hardwick JM (2009) Bcl-x L increases mitochondrial fission, fusion, and biomass in neurons. J Cell Biol 184(5):707–719. doi:10.1083/jcb.200809060

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  21. Sarov M, Murray JI, Schanze K, Pozniakovski A, Niu W, Angermann K, Hasse S, Rupprecht M, Vinis E, Tinney M, Preston E, Zinke A, Enst S, Teichgraber T, Janette J, Reis K, Janosch S, Schloissnig S, Ejsmont RK, Slightam C, Xu X, Kim SK, Reinke V, Stewart AF, Snyder M, Waterston RH, Hyman AA (2012) A genome-scale resource for in vivo tag-based protein function exploration in C. elegans. Cell 150(4):855–866. doi:10.1016/j.cell.2012.08.001

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  22. Dunn KW, Kamocka MM, McDonald JH (2011) A practical guide to evaluating colocalization in biological microscopy. Am J Physiol Cell Physiol 300(4):C723–C742. doi:10.1152/ajpcell.00462.2010

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  23. Rolland SG, Lu Y, David CN, Conradt B (2009) The BCL-2-like protein CED-9 of C. elegans promotes FZO-1/Mfn1,2- and EAT-3/Opa1-dependent mitochondrial fusion. J Cell Biol 186(4):525–540. doi:10.1083/jcb.200905070

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  24. Head BP, Zulaika M, Ryazantsev S, van der Bliek AM (2011) A novel mitochondrial outer membrane protein, MOMA-1, that affects cristae morphology in Caenorhabditis elegans. Mol Biol Cell 22(6):831–841. doi:10.1091/mbc.E10-07-0600

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  25. Brys K, Castelein N, Matthijssens F, Vanfleteren JR, Braeckman BP (2010) Disruption of insulin signalling preserves bioenergetic competence of mitochondria in ageing Caenorhabditis elegans. BMC Biol 8:91. doi:10.1186/1741-7007-8-91

    Article  PubMed  PubMed Central  Google Scholar 

  26. Senoo-Matsuda N, Hartman PS, Akatsuka A, Yoshimura S, Ishii N (2003) A complex II defect affects mitochondrial structure, leading to ced-3- and ced-4-dependent apoptosis and aging. J Biol Chem 278(24):22031–22036. doi:10.1074/jbc.M211377200

    Article  CAS  PubMed  Google Scholar 

  27. Mun JY, Lee TH, Kim JH, Yoo BH, Bahk YY, Koo HS, Han SS (2010) Caenorhabditis elegans mitofilin homologs control the morphology of mitochondrial cristae and influence reproduction and physiology. J Cell Physiol 224(3):748–756. doi:10.1002/jcp.22177

    Article  CAS  PubMed  Google Scholar 

  28. Hadwiger G, Dour S, Arur S, Fox P, Nonet ML (2010) A monoclonal antibody toolkit for C. elegans. PLoS One 5(4):e10161. doi:10.1371/journal.pone.0010161

    Article  PubMed  PubMed Central  Google Scholar 

  29. Yokoyama K, Fukumoto K, Murakami T, Harada S, Hosono R, Wadhwa R, Mitsui Y, Ohkuma S (2002) Extended longevity of Caenorhabditis elegans by knocking in extra copies of hsp70F, a homolog of mot-2 (mortalin)/mthsp70/Grp75. FEBS Lett 516(1-3):53–57

    Article  CAS  PubMed  Google Scholar 

  30. Wirth M, Karaca S, Wenzel D, Ho L, Tishkoff D, Lombard DB, Verdin E, Urlaub H, Jedrusik-Bode M, Fischle W (2013) Mitochondrial SIRT4-type proteins in Caenorhabditis elegans and mammals interact with pyruvate carboxylase and other acetylated biotin-dependent carboxylases. Mitochondrion 13(6):705–720. doi:10.1016/j.mito.2013.02.002

    Article  CAS  PubMed  Google Scholar 

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Acknowledgments

We thank Barbara Conradt, Nadin Memar, and Anne Kathrin Müller-Rischart for their comments on the manuscript. This work was supported by funding from the National Institutes of Health (GM076651), the Center for Integrated Protein Science Munich (EXC114) and the Deutsche Forschungsgemeinschaft (C0204/6-1) to Barbara Conradt.

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Authors and Affiliations

  1. LMU Biocenter, Department Biology II, Ludwig-Maximilians-University Munich, Grosshadernerstr. 2, Planegg-Martinsried, Munich, 82152, Germany

    Saroj G. Regmi & Stéphane G. Rolland

  2. Department of Genetics, Geisel School of Medicine at Dartmouth, 7400 Remsen, Hanover, NH, 03755, USA

    Saroj G. Regmi

  3. National Institute of Child Health and Human Development, NIH, 18 Center Drive, Bethesda, 20892, MD, USA

    Saroj G. Regmi

Authors
  1. Saroj G. Regmi
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  2. Stéphane G. Rolland
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Corresponding author

Correspondence to Stéphane G. Rolland .

Editor information

Editors and Affiliations

  1. Biomedical Center-Physiological Chemistry, LMU Munich, Martinsried, Germany

    Dejana Mokranjac

  2. Gene Center, LMU Munich and Institute for Diabetes and Obesity, Helmholtz Zentrum München, Munich, Germany

    Fabiana Perocchi

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Regmi, S.G., Rolland, S.G. (2017). New Imaging Tools to Analyze Mitochondrial Morphology in Caenorhabditis elegans . In: Mokranjac, D., Perocchi, F. (eds) Mitochondria. Methods in Molecular Biology, vol 1567. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-6824-4_16

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  • DOI: https://doi.org/10.1007/978-1-4939-6824-4_16

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  • Publisher Name: Humana Press, New York, NY

  • Print ISBN: 978-1-4939-6822-0

  • Online ISBN: 978-1-4939-6824-4

  • eBook Packages: Springer Protocols

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