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G2 Premature Chromosome Condensation/Chromosome Aberration Assay: Drug-Induced Premature Chromosome Condensation (PCC) Protocols and Cytogenetic Approaches in Mitotic Chromosome and Interphase Chromatin for Radiation Biology

  • Eisuke Gotoh
Protocol
Part of the Methods in Molecular Biology book series (MIMB, volume 1984)

Abstract

Chromosome analysis is a fundamental technique for a wide range of cytogenetic studies. Chromosome aberrations are easily introduced by many kinds of clastogenic agents such as ionizing irradiation, UV, or alkylating agents, and damaged chromosomes may be prone to cancer. Chromosomes are conventionally prepared from mitotic cells arrested by the colcemid block method. However, obtaining of mitotic chromosomes is sometimes hampered under several circumstances, for example after high-dose (over several Gys of γ-rays) ionizing irradiation exposure accident. As a result, cytogenetic analysis will be often difficult or even impossible in such cases. Premature chromosome condensation (PCC) is an alternative technique that has proved to be a unique and useful way in chromosome analysis. Previously, PCC has been achieved following cell fusion mediated either by fusogenic viruses (for example Sendai virus) or by polyethylene glycol (PEG) (cell-fusion PCC), but the cell-fusion PCC has several drawbacks. The novel drug-induced PCC use of specific inhibitors for serine/threonine protein phosphatase was introduced about 20 years ago. This method is much simple and easy even than the conventional mitotic chromosome preparation using colcemid block protocol and the obtained PCC index (equivalent to mitotic index for metaphase chromosome) is much higher. Furthermore, this method allows the interphase chromatin to be condensed and visualized like mitotic chromosomes, and thus has been opening the way for chromosome analysis not only in metaphase chromosomes but also in interphase chromatin. The drug-induced PCC has therefore proven the usefulness in cytogenetics and other many cell biology fields. Since the first version of drug-induced PCC protocol has been published in 2009 (Gotoh, Methods in molecular biology. Humana Press, New York, 2009), many newer applications of drug-induced PCC in radiation biology and chromosome science fields in a wide range of species from animal to plant have been reported (Gotoh et al., Biomed Res 16:63–68, 1995; Lamadrid Boada et al., Mutat Res 757:45–51, 2013; Ravi et al., Biochimie 95:124–33, 2013; Ono et al., J Cell Biol 200:429–41, 2013; Vagnarelli, Exp Cell Res 318:1435–41, 2012; Roukos et al., Nat Protoc 9:2476–92, 2014; Miura and Blakely, Cytometry A 79:1016–22, 2013; Zabka et al., J Plant Physiol 174:62–70, 2015; Samaniego et al., Planta 215:195–204, 2002; Rybaczek et al., Folia Histochem Cytobiol 40:51–9, 2002; Gotoh and Durante J Cell Physiol 209:297–304, 2006). Therefore as a new edition, I will write in this chapter the drug-induced PCC technique with newer findings, in particular focused drug-induced PCC protocols in radiation biology with referring updated articles published recently.

Key words

Cytogenetic study Chromosome analysis Radiation biology Biological dosimeter Chromosome damages and repair Repair kinetics Premature chromosome condensation (PCC) Prematurely condensed chromosomes (PCCs) Protein phosphatase inhibitors Calyculin A Okadaic acid Mitotic index PCC index 

Notes

Acknowledgments

I wish to express my thanks to Prof. Paul Wilson of Brookhaven National Laboratory and Prof. Takamitsu Kato of Colorado State University for their recommendation to give me a chance to contribute to this chapter.

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© Springer Science+Business Media, LLC, part of Springer Nature 2019

Authors and Affiliations

  • Eisuke Gotoh
    • 1
  1. 1.Department of RadiologyJikei University School of MedicineMinato-kuJapan

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