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Quantification of Mitotic Chromosome Alignment

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The Mitotic Spindle

Part of the book series: Methods in Molecular Biology ((MIMB,volume 1413))

Abstract

The alignment of chromosomes during metaphase is a hallmark of mitosis. For this reason, chromosome alignment has served as an informative functional assay for evaluating mitotic fidelity. The common approach of quantifying the number of mitotic cells with unaligned chromosomes within a population has led to the identification of many proteins required for this conserved process. However, more sensitive assays are now required to dissect the complex molecular control of chromosome alignment. In this chapter, we describe a microscopy-based method for objectively quantifying the distribution of fluorescently labeled chromosomes within the mitotic spindle that can be used to evaluate the extent of chromosome alignment within individual mitotic cells.

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References

  1. Mitchison TJ, Salmon ED (2001) Mitosis: a history of division. Nat Cell Biol 3(1):E17–E21

    Article  CAS  PubMed  Google Scholar 

  2. Amaro AC, Samora CP, Holtackers R, Wang E, Kingston IJ, Alonso M, Lampson M, McAinsh AD, Meraldi P (2010) Molecular control of kinetochore-microtubule dynamics and chromosome oscillations. Nat Cell Biol 12(4):319–329. doi:10.1038/ncb2033

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  3. Cai S, O'Connell CB, Khodjakov A, Walczak CE (2009) Chromosome congression in the absence of kinetochore fibres. Nat Cell Biol 11(7):832–838. doi:10.1038/ncb1890

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  4. Foltz DR, Jansen LE, Black BE, Bailey AO, Yates JR 3rd, Cleveland DW (2006) The human CENP-A centromeric nucleosome-associated complex. Nat Cell Biol 8(5):458–469. doi:10.1038/ncb1397

    Article  CAS  PubMed  Google Scholar 

  5. Goshima G, Vale RD (2003) The roles of microtubule-based motor proteins in mitosis: comprehensive RNAi analysis in the Drosophila S2 cell line. J Cell Biol 162(6):1003–1016

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  6. Lampson MA, Kapoor TM (2005) The human mitotic checkpoint protein BubR1 regulates chromosome-spindle attachments. Nat Cell Biol 7(1):93–98. doi:10.1038/ncb1208

    Article  CAS  PubMed  Google Scholar 

  7. McCleland ML, Gardner RD, Kallio MJ, Daum JR, Gorbsky GJ, Burke DJ, Stukenberg PT (2003) The highly conserved Ndc80 complex is required for kinetochore assembly, chromosome congression, and spindle checkpoint activity. Genes Dev 17(1):101–114. doi:10.1101/gad.1040903

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  8. Putkey FR, Cramer T, Morphew MK, Silk AD, Johnson RS, McIntosh JR, Cleveland DW (2002) Unstable kinetochore-microtubule capture and chromosomal instability following deletion of CENP-E. Dev Cell 3(3):351–365

    Article  CAS  PubMed  Google Scholar 

  9. Zhu C, Zhao J, Bibikova M, Leverson JD, Bossy-Wetzel E, Fan JB, Abraham RT, Jiang W (2005) Functional analysis of human microtubule-based motor proteins, the kinesins and dyneins, in mitosis/cytokinesis using RNA interference. Mol Biol Cell 16(7):3187–3199

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  10. Bissonette S, Stumpff J (2014) Quantifying mitotic chromosome dynamics and positioning. J Cell Physiol 229(10):1301–1305. doi:10.1002/jcp.24634

    Article  CAS  PubMed  Google Scholar 

  11. Kim H, Fonseca C, Stumpff J (2014) A unique kinesin-8 surface loop provides specificity for chromosome alignment. Mol Biol Cell 25(21):3319–3329. doi:10.1091/mbc.E14-06-1132

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  12. Stumpff J, Wagenbach M, Franck A, Asbury CL, Wordeman L (2012) Kif18A and chromokinesins confine centromere movements via microtubule growth suppression and spatial control of kinetochore tension. Dev Cell 22(5):1017–1029. doi:10.1016/j.devcel.2012.02.013

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  13. Genschik P, Criqui MC, Parmentier Y, Derevier A, Fleck J (1998) Cell cycle-dependent proteolysis in plants. Identification Of the destruction box pathway and metaphase arrest produced by the proteasome inhibitor mg132. Plant Cell 10(12):2063–2076

    CAS  PubMed  PubMed Central  Google Scholar 

  14. Lee DH, Goldberg AL (1996) Selective inhibitors of the proteasome-dependent and vacuolar pathways of protein degradation in Saccharomyces cerevisiae. J Biol Chem 271(44):27280–27284

    Article  CAS  PubMed  Google Scholar 

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Author information

Authors and Affiliations

  1. Department of Molecular Physiology and Biophysics, University of Vermont College of Medicine, Given Building, RM E217F, Burlington, VT, 05405, USA

    Cindy Fonseca & Jason Stumpff

Authors
  1. Cindy Fonseca
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  2. Jason Stumpff
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Corresponding author

Correspondence to Jason Stumpff .

Editor information

Editors and Affiliations

  1. Massachusetts Institute of Technolo, Cambridge, Massachusetts, USA

    Paul Chang

  2. Dept of Cell & Developmental Biolog, Vanderbilt University, Nashville, Tennessee, USA

    Ryoma Ohi

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© 2016 Springer Science+Business Media New York

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Fonseca, C., Stumpff, J. (2016). Quantification of Mitotic Chromosome Alignment. In: Chang, P., Ohi, R. (eds) The Mitotic Spindle. Methods in Molecular Biology, vol 1413. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-3542-0_16

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  • DOI: https://doi.org/10.1007/978-1-4939-3542-0_16

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  • Publisher Name: Humana Press, New York, NY

  • Print ISBN: 978-1-4939-3540-6

  • Online ISBN: 978-1-4939-3542-0

  • eBook Packages: Springer Protocols

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