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Functional and physical interactions of the ARF tumor suppressor with p53 and Mdm2

T Kamijo1, J D Weber, G Zambetti

  • 1Howard Hughes Medical Institute, St. Jude Children's Research Hospital, 332 North Lauderdale, Memphis, TN 38105, USA.

Proceedings of the National Academy of Sciences of the United States of America
|July 8, 1998
PubMed

Insights

The ARF protein directly interacts with p53, stabilizing it and promoting cell cycle arrest. However, supra-physiologic ARF levels can impair p53 function, suggesting a complex regulatory role in cancer suppression.

Area of Science:

  • Molecular Biology
  • Cancer Research
  • Cell Cycle Regulation

Background:

  • The INK4a-ARF locus is crucial for cell growth control, encoding p16(INK4a) and p19(ARF).
  • Disruptions in the INK4a-ARF locus are frequent in human cancers, second only to p53 alterations.
  • p19(ARF) and p53 are key tumor suppressors involved in cell cycle regulation.

Purpose of the Study:

  • To investigate the direct interaction between mouse p19(ARF) and p53.
  • To elucidate the mechanisms by which p19(ARF) influences p53 function and cell proliferation.
  • To understand the role of the ARF N-terminal domain in p53 binding and cell cycle arrest.

Main Methods:

  • Utilized baculovirus vectors in insect cells and retroviral vectors in NIH 3T3 fibroblasts.
  • Performed co-immunoprecipitation assays to detect protein interactions.
  • Analyzed p53 half-life, p53-dependent transcriptional activity, and cell cycle arrest in response to p19(ARF) overexpression.

Main Results:

  • Demonstrated direct interaction between mouse p19(ARF) and p53, as well as with mdm2.
  • Showed that p19(ARF) binds to p53-DNA complexes and requires functional p53 for inducing mdm2 and p21(Cip1).
  • Identified the ARF N-terminal domain (amino acids 1-62) as essential for p53 binding and cell cycle arrest, and observed that supra-physiologic p19(ARF) levels can paradoxically inhibit p53-mediated responses.

Conclusions:

  • p19(ARF) directly interacts with p53, stabilizing it and promoting cell cycle arrest through transcriptional induction of mdm2 and p21(Cip1).
  • The N-terminal domain of ARF is critical for its interaction with p53 and subsequent cell cycle inhibition.
  • ARF modulates p53 function through both stabilization and an additional, yet to be fully defined, mechanism that can be impaired at supra-physiologic concentrations.

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