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Updated: Apr 19, 2026
Studying Proteolysis of Cyclin B at the Single Cell Level in Whole Cell Populations
Published on: September 17, 2012
Nuclear translocation of Cyclin B1 marks the restriction point for terminal cell cycle exit in G2 phase
Erik Müllers1, Helena Silva Cascales, Himjyot Jaiswal
1a Department of Cell and Molecular Biology; Karolinska Institutet ; Stockholm , Sweden.
Abstract:
Upon DNA damage, cell cycle progression is temporally blocked to avoid propagation of mutations. While transformed cells largely maintain the competence to recover from a cell cycle arrest, untransformed cells past the G1/S transition lose mitotic inducers, and thus the ability to resume cell division. This permanent cell cycle exit depends on p21, p53, and APC/C(Cdh1). However, when and how permanent cell cycle exit occurs remains unclear. Here, we have investigated the cell cycle response to DNA damage in single cells that express Cyclin B1 fused to eYFP at the endogenous locus. We find that upon DNA damage Cyclin B1-eYFP continues to accumulate up to a threshold level, which is reached only in G2 phase. Above this threshold, a p21 and p53-dependent nuclear translocation required for APC/C(Cdh1)-mediated Cyclin B1-eYFP degradation is initiated. Thus, cell cycle exit is decoupled from activation of the DNA damage response in a manner that correlates to Cyclin B1 levels, suggesting that G2 activities directly feed into the decision for cell cycle exit. Once Cyclin B1-eYFP nuclear translocation occurs, checkpoint inhibition can no longer promote mitotic entry or re-expression of mitotic inducers, suggesting that nuclear translocation of Cyclin B1 marks the restriction point for permanent cell cycle exit in G2 phase.
Insights
DNA damage halts cell cycle progression. Untransformed cells permanently exit the cell cycle in G2 phase when Cyclin B1 reaches a threshold, marking a restriction point for cell division.
Area of Science:
- Cell Biology
- Molecular Biology
- Genetics
Background:
- DNA damage triggers cell cycle arrest to prevent mutations.
- Untransformed cells permanently exit the cell cycle after G1/S, unlike transformed cells.
- The precise timing and mechanisms of permanent cell cycle exit remain unclear.
Purpose of the Study:
- To investigate the cell cycle response to DNA damage in single cells.
- To determine the role of Cyclin B1 levels in permanent cell cycle exit.
- To identify the restriction point for irreversible cell cycle arrest.
Main Methods:
- Utilized single-cell analysis of Cyclin B1 fused to eYFP at the endogenous locus.
- Monitored Cyclin B1-eYFP accumulation and localization upon DNA damage.
- Investigated the involvement of p21, p53, and APC/C(Cdh1) in the process.
Main Results:
- Cyclin B1-eYFP accumulates to a threshold level specifically in G2 phase following DNA damage.
- Above this threshold, p21 and p53 mediate nuclear translocation of Cyclin B1-eYFP.
- Nuclear translocation triggers degradation of Cyclin B1-eYFP and initiates permanent cell cycle exit.
- Cell cycle exit is decoupled from DNA damage response activation and linked to Cyclin B1 levels.
Conclusions:
- Cyclin B1 nuclear translocation in G2 phase represents a critical restriction point for permanent cell cycle exit.
- G2 phase activities, particularly Cyclin B1 levels, directly influence the decision for irreversible cell cycle arrest.
- Checkpoint inhibition is ineffective after Cyclin B1 nuclear translocation, preventing mitotic re-entry.
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