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Nuclear export of cyclin B1 and its possible role in the DNA damage-induced G2 checkpoint
F Toyoshima1, T Moriguchi, A Wada
1Department of Biophysics, Graduate School of Science, Kyoto University, Kyoto, Japan.
Abstract:
M-phase-promoting factor (MPF), a complex of cdc2 and a B-type cyclin, is a key regulator of the G2/M cell cycle transition. Cyclin B1 accumulates in the cytoplasm through S and G2 phases and translocates to the nucleus during prophase. We show here that cytoplasmic localization of cyclin B1 during interphase is directed by its nuclear export signal (NES)-dependent transport mechanism. Treatment of HeLa cells with leptomycin B (LMB), a specific inhibitor of the NES-dependent transport, resulted in nuclear accumulation of cyclin B1 in G2 phase. Disruption of an NES which has been identified in cyclin B1 here abolished the nuclear export of this protein, and consequently the NES-disrupted cyclin B1 when expressed in cells accumulated in the nucleus. Moreover, we show that expression of the NES-disrupted cyclin B1 or LMB treatment of the cells is able to override the DNA damage-induced G2 checkpoint when combined with caffeine treatment. These results suggest a role of nuclear exclusion of cyclin B1 in the DNA damage-induced G2 checkpoint.
Insights
Cyclin B1 nuclear export, regulated by its NES, is crucial for the DNA damage G2 checkpoint. Disrupting this export overrides the checkpoint, suggesting nuclear exclusion
Area of Science:
- Cell Biology
- Molecular Biology
- Cell Cycle Regulation
Background:
- M-phase-promoting factor (MPF), composed of cdc2 and cyclin B, controls the G2/M cell cycle transition.
- Cyclin B1 normally accumulates in the cytoplasm during S and G2 phases before nuclear translocation in prophase.
Purpose of the Study:
- To investigate the mechanism of cytoplasmic localization of cyclin B1 during interphase.
- To determine the role of cyclin B1's nuclear export in the DNA damage-induced G2 checkpoint.
Main Methods:
- Treatment of HeLa cells with leptomycin B (LMB), a nuclear export inhibitor.
- Identification and disruption of a nuclear export signal (NES) within cyclin B1.
- Analysis of cyclin B1 localization and its effect on the G2 checkpoint, often in combination with caffeine treatment.
Main Results:
- Leptomycin B treatment caused nuclear accumulation of cyclin B1 in G2 phase HeLa cells.
- Disruption of cyclin B1's NES abolished nuclear export, leading to nuclear accumulation.
- NES-disrupted cyclin B1 expression or LMB treatment, combined with caffeine, overrode the DNA damage-induced G2 checkpoint.
Conclusions:
- Cyclin B1's cytoplasmic localization during interphase is actively maintained by an NES-dependent nuclear export mechanism.
- Nuclear exclusion of cyclin B1 plays a significant role in the DNA damage-induced G2 checkpoint.
- Targeting this nuclear export pathway offers potential strategies for manipulating cell cycle checkpoints.