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Imaging Analysis of Cell Cycle-Dependent Degradation of Cdt1 in Mammalian Cells

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Cell Cycle Control

Part of the book series: Methods in Molecular Biology ((MIMB,volume 1170))

Abstract

Numerous cell cycle-regulating proteins are controlled by protein degradation. Recent work shows that ubiquitination-dependent proteolysis plays an important role in once-per-cell cycle control of DNA replication. Cdt1 is a licensing factor essential for assembling the pre-replicative complex on replication origins. Cdt1 is present in G1 phase, but after S phase ubiquitin-mediated proteolysis maintains Cdt1 at low levels. This is important to prevent the re-replication of chromosomal DNA. The cell cycle-dependent degradation of Cdt1 can be monitored by dual staining of the cell nuclei with antibodies against Cdt1- and S/G2-phase marker proteins, such as cyclin A or geminin.

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Acknowledgements

This work was financially supported by JSPS KAKENHI and MEXT KAKENHI, Grant in Aid from the Ministry of Education, Culture, Sports, Science, and Technology of Japan.

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Authors and Affiliations

  1. Graduate School of Life Science, University of Hyogo, Kamigori, Ako-gun, Hyogo, 678-1297, Japan

    Yasushi Shiomi, Naohiro Suenaga, Miyuki Tanaka, Akiyo Hayashi & Hideo Nishitani

Authors
  1. Yasushi Shiomi
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  2. Naohiro Suenaga
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  3. Miyuki Tanaka
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  4. Akiyo Hayashi
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  5. Hideo Nishitani
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Corresponding author

Correspondence to Hideo Nishitani .

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Editors and Affiliations

  1. Department of Biochemistry and Molecular Biology, Drexel University College of Medicine, Philadelphia, Pennsylvania, USA

    Eishi Noguchi

  2. Dept of Biochemistry & Molecular Biology, Drexel University College of Medicine, Philadelphia, USA

    Mariana C. Gadaleta

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Shiomi, Y., Suenaga, N., Tanaka, M., Hayashi, A., Nishitani, H. (2014). Imaging Analysis of Cell Cycle-Dependent Degradation of Cdt1 in Mammalian Cells. In: Noguchi, E., Gadaleta, M. (eds) Cell Cycle Control. Methods in Molecular Biology, vol 1170. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-0888-2_18

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  • DOI: https://doi.org/10.1007/978-1-4939-0888-2_18

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  • Publisher Name: Humana Press, New York, NY

  • Print ISBN: 978-1-4939-0887-5

  • Online ISBN: 978-1-4939-0888-2

  • eBook Packages: Springer Protocols

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