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Updated: Sep 26, 2026
Yeast As a Chassis for Developing Functional Assays to Study Human P53
Published on: August 4, 2019
Loss of p53 induces M-phase retardation following G2 DNA damage checkpoint abrogation
Yuzuru Minemoto1, Sanae Uchida, Motoaki Ohtsubo
1Department of Life Science, Graduate School of Natural Science and Technology, General Education Hall, Kanazawa University, Kakuma-machi, Kanazawa 920-1192, Ishikawa, Japan.
Abstract:
Most cell lines that lack functional p53 protein are arrested in the G2 phase of the cell cycle due to DNA damage. When the G2 checkpoint is abrogated, these cells are forced into mitotic catastrophe. A549 lung adenocarcinoma cells, in which p53 was eliminated with the HPV16 E6 gene, exhibited efficient arrest in the G2 phase when treated with adriamycin. Administration of caffeine to G2-arrested cells induced a drastic change in cell phenotype, the nature of which depended on the status of p53. Flow cytometric and microscopic observations revealed that cells that either contained or lacked p53 resumed their cell cycles and entered mitosis upon caffeine treatment. However, transit to the M phase was slower in p53-negative cells than in p53-positive cells. Consistent with these observations, CDK1 activity was maintained at high levels, along with stable cyclin B1, in p53-negative cells. The addition of butyrolactone I, which is an inhibitor of CDK1 and CDK2, to the p53-negative cells reduced the floating round cell population and induced the disappearance of cyclin B1. These results suggest a relationship between the p53 pathway and the ubiquitin-mediated degradation of mitotic cyclins and possible cross-talk between the G2-DNA damage checkpoint and the mitotic checkpoint.
Insights
Cancer cells lacking p53 protein arrest in G2 phase. Caffeine treatment reinitiates cell cycle progression, but p53-negative cells show slower entry into mitosis, revealing cell cycle checkpoint interactions.
Area of Science:
- Cell Biology
- Molecular Biology
- Cancer Research
Background:
- Most cancer cells lacking functional p53 protein arrest in the G2 phase due to DNA damage.
- Abrogation of the G2 checkpoint in these cells can lead to mitotic catastrophe.
Purpose of the Study:
- To investigate the effect of caffeine on G2-arrested A549 lung adenocarcinoma cells with and without functional p53.
- To explore the relationship between the p53 pathway, cell cycle checkpoints, and mitotic cyclin degradation.
Main Methods:
- Utilized A549 lung adenocarcinoma cells with p53 eliminated via the HPV16 E6 gene.
- Treated G2-arrested cells with adriamycin and caffeine.
- Performed flow cytometry and microscopic observations.
- Assessed CDK1 activity and cyclin B1 levels.
- Used butyrolactone I, a CDK1/CDK2 inhibitor.
Main Results:
- Caffeine treatment induced cell cycle resumption and entry into mitosis in both p53-positive and p53-negative cells.
- p53-negative cells exhibited slower transit to M phase compared to p53-positive cells.
- High CDK1 activity and stable cyclin B1 were observed in p53-negative cells.
- Butyrolactone I treatment reduced floating cells and cyclin B1 levels in p53-negative cells.
Conclusions:
- Suggests a link between the p53 pathway and the ubiquitin-mediated degradation of mitotic cyclins.
- Indicates potential cross-talk between the G2-DNA damage checkpoint and the mitotic checkpoint.
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