Journal
CELL CYCLE
Volume 17, Issue 21-22, Pages 2496-2516Publisher
TAYLOR & FRANCIS INC
DOI: 10.1080/15384101.2018.1547001
Keywords
Live-cell imaging; G1/S transition; single-cell dynamics
Categories
Funding
- NIH Office of the Director [DP2-HD091800]
- National Cancer Institute [T32CA009156]
- National Institute of General Medical Sciences [GM102413, T32GM007040, GM083024]
- W. M. Keck Foundation [Medical Research Grant]
- EUNICE KENNEDY SHRIVER NATIONAL INSTITUTE OF CHILD HEALTH & HUMAN DEVELOPMENT [DP2HD091800] Funding Source: NIH RePORTER
- NATIONAL CANCER INSTITUTE [T32CA009156] Funding Source: NIH RePORTER
- NATIONAL INSTITUTE OF GENERAL MEDICAL SCIENCES [T32GM007040, R01GM102413, R01GM083024] Funding Source: NIH RePORTER
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Cell cycle phase transitions are tightly orchestrated to ensure efficient cell cycle progression and genome stability. Interrogating these transitions is important for understanding both normal and pathological cell proliferation. By quantifying the dynamics of the popular FUCCI reporters relative to the transitions into and out of S phase, we found that their dynamics are substantially and variably offset from true S phase boundaries. To enhance detection of phase transitions, we generated a new reporter whose oscillations are directly coupled to DNA replication and combined it with the FUCCI APC/C reporter to create PIP-FUCCI. The PIP degron fusion protein precisely marks the G1/S and S/G2 transitions; shows a rapid decrease in signal in response to large doses of DNA damage only during G1; and distinguishes cell type-specific and DNA damage source-dependent arrest phenotypes. We provide guidance to investigators in selecting appropriate fluorescent cell cycle reporters and new analysis strategies for delineating cell cycle transitions.
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