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Inhibition of p53-dependent transcription by BOX-I phospho-peptide mimetics that bind to p300

D Dornan1, T R Hupp

  • 1Department of Molecular & Cellular Pathology, University of Dundee, UK.

EMBO Reports
|March 22, 2001
PubMed

Insights

Phosphorylation of p53 at Thr18 or Ser20 stabilizes the p300-p53 complex, activating p53. This finding reveals potential therapeutic targets for modulating p53 activity.

Area of Science:

  • Molecular Biology
  • Cancer Research
  • Protein Biochemistry

Background:

  • The p53 tumor suppressor protein's activity is tightly regulated by its N-terminal domain.
  • This domain contains binding sites for MDM2 (negative regulator) and p300 (positive effector).
  • Two phosphorylation sites, Thr18 and Ser20, in the p53 N-terminal domain have been identified, but their effect on p300 interaction was unclear.

Purpose of the Study:

  • To investigate the impact of p53 phosphorylation at Thr18 or Ser20 on its interaction with p300.
  • To determine how these phosphorylations influence p53-dependent transcriptional activity.
  • To explore the potential for small molecules to selectively modulate p53 activity via MDM2 or p300 interactions.

Main Methods:

  • Biochemical assays to assess protein-protein interactions (p53-MDM2, p53-p300).
  • Use of engineered peptides (EGFP-BOX-I domain) with specific mutations (Aspartate substitution at Thr18 or Ser20) to mimic phosphorylation.
  • Reporter assays to measure p53-dependent transcriptional activity in various cellular contexts (cycling, irradiated, co-transfected cells).

Main Results:

  • Phosphorylation at Thr18 or Ser20 inhibits MDM2 binding to p53.
  • Crucially, these phosphorylations significantly stabilize the binding of p300 to p53.
  • Mutant peptides mimicking phosphorylation inhibited p53 transcription, while wild-type peptides stimulated it by inhibiting MDM2 binding.

Conclusions:

  • Phosphorylation of p53 at Thr18 or Ser20 acts as an activation mechanism by stabilizing the p300-p53 complex.
  • This study identifies a novel regulatory pathway for p53 activation.
  • The findings suggest the development of small molecules that can selectively target MDM2- or p300-dependent p53 activities for therapeutic purposes.

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