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The LINC Complex pp 3–15Cite as

EM Tomography of Meiotic LINC Complexes

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Part of the book series: Methods in Molecular Biology ((MIMB,volume 1840))

Abstract

Electron microscope (EM) tomography is a powerful technique that enables the three-dimensional analysis of subcellular structures at high resolution. We have applied this method to the quantitative analysis of LINC complex distribution and interaction with the cytoskeleton in meiotic cells from male mice. In this chapter, we describe methods to generate and analyze the tomograms.

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References

  1. Ungricht R, Kutay U (2017) Mechanisms and functions of nuclear envelope remodeling. Nat Rev Mol Cell Biol 18:229–245

    Article  CAS  PubMed  Google Scholar 

  2. Chang W, Worman HJ, Gundersen GG (2015) Accessorizing and anchoring the LINC complex for multifunctionality. J Cell Biol 208:11–22

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  3. Link J, Jahn D, Alsheimer M (2015) Structural and functional adaptations of the mammalian nuclear envelope to meet the meiotic requirements. Nucleus 6:93–101

    Article  PubMed  PubMed Central  Google Scholar 

  4. Link J, Benavente R, Alsheimer M (2016) Analysis of meiotic telomere behavior in the mouse. Methods Mol Biol 1411:195–208

    Article  CAS  PubMed  Google Scholar 

  5. Liebe B, Alsheimer M, Höög C et al (2004) Telomere attachment, meiotic chromosome condensation, pairing, and bouquet stage duration are modified in spermatocytes lacking axial elements. Mol Biol Cell 15:827–837

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  6. Schmitt J, Benavente R, Hodzic D et al (2007) Transmembrane protein Sun2 is involved in tethering mammalian meiotic telomeres to the nuclear envelope. Proc Natl Acad Sci U S A 104:7426–7431

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  7. Ding X, Xu R, Yu J et al (2007) SUN1 is required for telomere attachment to nuclear envelope and gametogenesis in mice. Dev Cell 12:863–872

    Article  CAS  PubMed  Google Scholar 

  8. Morimoto A, Shibuya H, Zhu X et al (2012) A conserved KASH domain protein associated with telomeres, SUN1 and dynactin during mammalian meiosis. J Cell Biol 198:165–172

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  9. Horn HF, Kim DI, Wright GD et al (2013) A mammalian KASH domain protein coupling meiotic chromosomes to the cytoskeleton. J Cell Biol 202:1023–1039

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  10. Spindler MC, Redolfi J, Helmprobst F et al (2018) Electron tomography of LINC complex association with microtubules at meiotic telomere attachment sites, submitted.

    Google Scholar 

  11. Reynolds ES (1963) The use of lead citrate at high pH as an electron-opaque stain in electron microscopy. J Cell Biol 17:208

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  12. Kremer JR, Mastronarde DN, McIntosh JR (1996) Computer visualization of three-dimensional image data using IMOD. J Struct Biol 116:71–76

    Article  CAS  PubMed  Google Scholar 

  13. Mastronarde DN (2005) Automated electron microscope tomography using robust prediction of specimen movements. J Struct Biol 152:36–51

    Article  PubMed  Google Scholar 

  14. Dhanyasi N, Segal D, Shimoni E et al (2015) Surface apposition and multiple cell contacts promote myoblast fusion in drosophila flight muscles. J Cell Biol 211:191–203

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  15. Weimer RM (2006) Preservation of C. elegans tissue via high-pressure freezing and freeze-substitution for ultrastructural analysis and immunocytochemistry. Methods Mol Biol 351:203–221

    PubMed  Google Scholar 

  16. Stigloher C, Zhan H, Zhen M et al (2011) The presynaptic dense projection of the Caenorhabditis elegans cholinergic neuromuscular junction localizes synaptic vesicles at the active zone through SYD-2/Liprin and UNC-10/RIM-dependent interactions. J Neurosci 31:4388–4396

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  17. Helmprobst F, Frank M, Stigloher C (2015) Presynaptic architecture of the larval zebrafish neuromuscular junction. J Comp Neurol 523:1984–1997

    Article  PubMed  Google Scholar 

  18. http://bio3d.colorado.edu/imod/. Accessed 8 June 2017.

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Acknowledgments

We are highly indebted to Markus Engstler (Biocenter, Würzburg) for equipment available in our laboratories. R.B. is supported by grant Be 1168/8-1 of the DFG.

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Authors and Affiliations

  1. Department of Cell and Developmental Biology, Biocenter, University of Würzburg, Würzburg, Germany

    Marie-Christin Spindler & Ricardo Benavente

  2. Imaging Core Facility, Biocenter, University of Würzburg, Würzburg, Germany

    Frederik Helmprobst & Christian Stigloher

Authors
  1. Marie-Christin Spindler
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  2. Frederik Helmprobst
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  3. Christian Stigloher
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  4. Ricardo Benavente
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Corresponding author

Correspondence to Ricardo Benavente .

Editor information

Editors and Affiliations

  1. Department of Pathology and Cell Biology, Columbia University, New York, New York, USA

    Gregg G. Gundersen

  2. Department of Medicine and Department of Pathology and Cell Biology, Columbia University, New York, New York, USA

    Howard J. Worman

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Spindler, MC., Helmprobst, F., Stigloher, C., Benavente, R. (2018). EM Tomography of Meiotic LINC Complexes. In: Gundersen, G., Worman, H. (eds) The LINC Complex. Methods in Molecular Biology, vol 1840. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-8691-0_1

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  • DOI: https://doi.org/10.1007/978-1-4939-8691-0_1

  • Published:

  • Publisher Name: Humana Press, New York, NY

  • Print ISBN: 978-1-4939-8690-3

  • Online ISBN: 978-1-4939-8691-0

  • eBook Packages: Springer Protocols

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