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A ribonucleotide reductase gene involved in a p53-dependent cell-cycle checkpoint for DNA damage

Abstract

The p53 gene is frequently inactivated in human cancers. Here we have isolated a p53-inducible gene, p53R2, by using differential display to examine messenger RNAs in a cancer-derived human cell line carrying a highly regulated wild-type p53 expression system. p53R2 contains a p53-binding sequence in intron 1 and encodes a 351-amino-acid peptide with striking similarity to the ribonucleotide reductase small subunit (R2), which is important in DNA synthesis during cell division. Expression of p53R2, but not R2, was induced by ultraviolet and γ-irradiation and adriamycin treatment in a wild-type p53-dependent manner. Induction of p53R2 in p53-deficient cells caused G2/M arrest and prevented cells from death in response to adriamycin. Inhibition of endogenous p53R2 expression in cells that have an intact p53-dependent DNA damage checkpoint reduced ribonucleotide reductase activity, DNA repair and cell survival after exposure to various genotoxins. Our results indicate that p53R2 encodes a ribonucleotide reductase that is directly involved in the p53 checkpoint for repair of damaged DNA. The discovery of p53R2 clarifies a relationship between a ribonucleotide reductase activity involved in repair of damaged DNA and tumour suppression by p53.

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Figure 1: Isolation and characterization of the p53R2 gene.
Figure 2: Comparison of the amino-acid sequences of p53R2, R2, RNR2 and RNR4.
Figure 3: The p53-responsive site in the p53R2 gene.
Figure 4: -dependent induction of p53R2 transcription in human cells after DNA damage by 14 Gy γ-radiation (a) or 0.2 µg ml-1 adriamycin (b).
Figure 5: Ectopic expression of p53R2 in p53-deficient cancer cells restores G2/M arrest and cell survival after DNA damage.
Figure 6: Cell-cycle arrest, p53R2 expression and DNA repair activity through p53R2 ribonucleotide reductase activity after DNA damage.
Figure 7: Inhibition of either p53 or p53R2 protein expression reduces DNA repair activity through ribonucleotide reductase after various genotoxic stresses.
Figure 8: Inhibition of either p53 or p53R2 protein expression reduces cell survival after genotoxic stress.
Figure 9: Hypothetical models for DNA damage repair and replication.

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Acknowledgements

This work was supported in part by the ‘Research for the Future’ Program of the Japan Society for the Promotion of Science.

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Correspondence to Yusuke Nakamura.

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Tanaka, H., Arakawa, H., Yamaguchi, T. et al. A ribonucleotide reductase gene involved in a p53-dependent cell-cycle checkpoint for DNA damage. Nature 404, 42–49 (2000). https://doi.org/10.1038/35003506

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