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  • Original Paper
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F9 embryonal carcinoma cells fail to stop at G1/S boundary of the cell cycle after γ-irradiation due to p21WAF1/CIP1 degradation

Oncogene volume 19, pages 3858–3865 (2000)Cite this article

Abstract

We studied the ability of F9 teratocarcinoma cells to arrest in G1/S and G2/M checkpoints after γ-irradiation. Wild-type p53 protein was rapidly accumulated in F9 cells after γ-irradiation, however, this was followed not by a G1/S arrest but by a short and reversible delay of the cell cycle in G2/M. In order to elucidate the reasons of the lack of G1/S arrest in F9 cells, we investigated the expression of p53 downstream target Cdk inhibitor p21WAF1/CIP1. In spite of p53-dependent activation of p21WAF1/CIP1 gene promoter and p21WAF1/CIP1 mRNA accumulation upon irradiation, the p21WAF1/CIP1 protein was not detected by either immunoblot or immunofluorescence techniques. However, the cells treated with a specific proteasome inhibitor lactacystin revealed the p21WAF1/CIP1 protein both in non-irradiated and irradiated cells. Therefore we suggest thatp21WAF1/CIP1 protein is degraded by a proteasome-dependent mechanism in F9 cells and the lack of G1/S arrest after γ-irradiation is due to this degradation. We also examined the expression and activity of cell cycle regulatory proteins: G1- and G2-cyclins and cyclin-dependent kinases. In the absence of functionalp21WAF1/CIP1 inhibitor, the activity of G1 cyclin/Cdk complexes was insufficiently inhibited to cause a G1 arrest, whereas a decrease of cdc2 and cyclin B1-associated kinase activities was enough to contribute to a reversible G2 arrest following γ-irradiation. Afterγ-irradiation, the majority of F9 cells undergo apoptosis implying that wt-p53 likely triggers pro-apoptotic gene expression in DNA damaged cells. Elimination of defected cells might ensure maintenance of genome integrity in the remaining cell population.

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Acknowledgements

We are grateful to Dr Yu M Rosanov for assistance in flow cytometry experiments, we thank Dr W El-Deiry who kindly provided WWP-luc plasmid and also Dr A Zantema for anti-cdk2 antibodies. This work was supported by Russian Foundation of Basic Research grants 00-04-49389 and INTAS 99-1036 (VAP), 00-04-49024 (TVK) and NCI-NIH CDA-NIS (TVK), and by the grant for young scientists of St Petersburg's mayor ASP no. 298414 (ABM).

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  1. Institute of Cytology, Laboratory of Molecular Basis of Cell Differentiation, Russian Academy of Sciences, 194064 St Petersburg, Tikhoretsky, 4, Russia

    Anna B Malashicheva, Tatiana V Kislyakova, Nikolay D Aksenov, Konstantin A Osipov & Valery A Pospelov

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  1. Anna B Malashicheva
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  2. Tatiana V Kislyakova
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Malashicheva, A., Kislyakova, T., Aksenov, N. et al. F9 embryonal carcinoma cells fail to stop at G1/S boundary of the cell cycle after γ-irradiation due to p21WAF1/CIP1 degradation. Oncogene 19, 3858–3865 (2000). https://doi.org/10.1038/sj.onc.1203736

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