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Functional role of Mdm2 phosphorylation by ATR in attenuation of p53 nuclear export

Oncogene volume 22, pages 8870–8880 (2003)Cite this article

Abstract

Mdm2 oncoprotein plays a major role in inhibiting the p53 tumor suppressor protein. Here, we investigate phosphorylation of Mdm2 at serine 407 (S407). S407 is phosphorylated in cells after treatment with camptothecin (CPT) or hydroxyurea, inhibitors of DNA replication. S407 phosphorylation after CPT treatment is induced upon cell cycle arrest during S phase and prevented if entry into S phase of cell cycle is blocked. We found that a major kinase responsible for S407 phosphorylation is ATR, a DNA damage checkpoint protein that induces cell cycle arrest and promotes DNA repair in response to impaired DNA replication; induction of S407 phosphorylation is enhanced after expression of wild-type ATR, while it is inhibited by a dominant-negative form of ATR. Further, S407 is specifically phosphorylated by ATR in vitro. Substitution of S407 with aspartate (S407D), but not with alanine (S407A), promotes nuclear localization of p53. Taken together, our data indicate that S407 phosphorylation of Mdm2 by ATR reduces Mdm2-dependent export of p53 from nuclei to cytoplasm.

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Acknowledgements

The Flag-tagged ATR expression vectors are kind gifts from Robert Abraham. We are indebted to Jiangdong Chen and Christian Gaiddon for providing us the Flag-tagged Mdm2 expression vector and the HA-tagged p53 expression vector, respectively. The His-ubiquitin expression plasmid is a kind gift from Dirk Bohmann. We also thank Satomi Yamazaki for technical assistance. We thank Issei Kitabayashi for critical reading of the manuscript. This work is supported by a Grant-in Aid for Scientific Research on Priority Area from the Ministry of Education, Culture, Sports, Science and Technology of Japan, the Program for Promotion of Fundamental Studies in Health Sciences of Organization for Pharmaceutical Safety and Research of Japan, Research Grant of the Princess Takamatsu Cancer Research Fund, and Takeda Science Foundation to YT.

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  1. Radiobiology Division, National Cancer Center Research Institute, Tsukiji 5-1-1, Chuo-ku, Tokyo, 104-0045, Japan

    Tomomi Shinozaki, Ayumi Nota, Yoichi Taya & Koji Okamoto

  2. CREST, Japan Science and Technology Corporation, Japan

    Tomomi Shinozaki, Ayumi Nota, Yoichi Taya & Koji Okamoto

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  1. Tomomi Shinozaki
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Correspondence to Koji Okamoto.

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Shinozaki, T., Nota, A., Taya, Y. et al. Functional role of Mdm2 phosphorylation by ATR in attenuation of p53 nuclear export. Oncogene 22, 8870–8880 (2003). https://doi.org/10.1038/sj.onc.1207176

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