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Author Notes:

Gowen Cook, jean_cook@med.unc.edu

J.P. Matson designed and performed the experiments and analyzed the results. A.M. House tracked cells from live cell imaging and processed data. G.D. Grant tracked cells and ran the microscope. H. Wu performed some immunoblots. J. Perez created the RPE1 Cdc6-mVenus, PCNA-mTurq2, DHB-mCherry cell line. J.G. Cook designed experiments and supervised the project. J.P. Matson and J.G. Cook wrote the manuscript with input from the other authors.

We thank Jeffrey Jones for managerial assistance and the Cook laboratory for helpful discussion. We thank Dr. Sam Wolff, Seraphina Wong, Aimee Littlejohn, and Walli Driggers for support. We thank Dr. Kasia Kedziora for help with imaging analysis. The DHB-mCherry plasmid was a gift from Dr. Sabrina Spencer. The Cdc6-mVenus plasmid was a gift from Dr. Michael Brandeis. The RPE1_hTert p53 CRIPSR null line was a gift from Dr. Prasad Jallepalli.

The authors declare no competing financial interests.

Subjects:

Research Funding:

This work was supported by a fellowship from the National Science Foundation (DGE-1144081) to J.P. Matson, a University of North Carolina Dissertation Completion Fellowship to J.P. Matson, and grants from the National Institutes of Health/National Institute of General Medical Sciences to J.G. Cook (GM083024, GM102413, and R25GM089569; R25GM089569 also supported J. Perez). The University of North Carolina Flow Cytometry Core Facility is supported in part by P30 CA016086.

Keywords:

  • Adaptor Proteins, Signal Transducing
  • Cell Cycle
  • Cell Cycle Proteins
  • Cell Division
  • Cell Nucleus
  • Cell Proliferation
  • Chromatin
  • DNA Replication
  • Flow Cytometry
  • G1 Phase
  • Genomic Instability
  • Humans
  • Nuclear Proteins
  • Replication Origin
  • S Phase

Intrinsic checkpoint deficiency during cell cycle re-entry from quiescence

Tools:

Journal Title:

Journal of Cell Biology

Volume:

Volume 218, Number 7

Publisher:

, Pages 2169-2184

Type of Work:

Article | Final Publisher PDF

Abstract:

To maintain tissue homeostasis, cells transition between cell cycle quiescence and proliferation. An essential G1 process is minichromosome maintenance complex (MCM) loading at DNA replication origins to prepare for S phase, known as origin licensing. A p53-dependent origin licensing checkpoint normally ensures sufficient MCM loading before S phase entry. We used quantitative flow cytometry and live cell imaging to compare MCM loading during the long first G1 upon cell cycle entry and the shorter G1 phases in the second and subsequent cycles. We discovered that despite the longer G1 phase, the first G1 after cell cycle re-entry is significantly underlicensed. Consequently, the first S phase cells are hypersensitive to replication stress. This underlicensing results from a combination of slow MCM loading with a severely compromised origin licensing checkpoint. The hypersensitivity to replication stress increases over repeated rounds of quiescence. Thus, underlicensing after cell cycle re-entry from quiescence distinguishes a higher-risk first cell cycle that likely promotes genome instability.

Copyright information:

© 2019 Matson et al.

This is an Open Access work distributed under the terms of the Creative Commons Attribution-NonCommercial-ShareAlike 4.0 International License (https://creativecommons.org/licenses/by-nc-sa/4.0/rdf).
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