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Yeast As a Chassis for Developing Functional Assays to Study Human P53
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Activation of p53 Facilitates the Target Search in DNA by Enhancing the Target Recognition Probability

Yuji Itoh1, Agato Murata1, Seiji Sakamoto2

  • 1Institute for Multidisciplinary Research for Advanced Materials, Tohoku University, Katahira 2-1-1, Aoba-ku, Sendai 980-8577, Japan; Department of Chemistry, Graduate School of Science, Tohoku University, Sendai 980-8578, Japan.

Journal of Molecular Biology
|June 14, 2016
PubMed

Insights

Tumor suppressor p53

Area of Science:

  • Molecular Biology
  • Genetics
  • Biophysics

Background:

  • The tumor suppressor p53 protein regulates gene expression by binding to specific DNA targets.
  • p53 utilizes a combination of 3D diffusion and 1D sliding to locate its genomic targets.
  • Understanding p53's target binding mechanism is crucial for comprehending its role in tumor suppression.

Purpose of the Study:

  • To investigate the regulatory mechanisms governing p53's target DNA binding.
  • To compare the target recognition probability (TRP) and binding dynamics of wild-type p53 with its activated and inactive mutants.

Main Methods:

  • Construction of pseudo-wild type (pseudo-WT), activated (S392E), and inactive (R248Q) p53 mutants.
  • Single-molecule fluorescence imaging to observe p53 binding to long DNA molecules.
  • Quantification of target recognition probability (TRP) and binding event fractions.

Main Results:

  • Pseudo-WT p53 exhibited a TRP of 7±2%, with 63±9% of binding events involving 1D sliding.
  • The activated mutant (S392E) showed a significantly increased TRP of 18±2% and an elevated 1D sliding fraction (87±2%).
  • The inactive mutant (R248Q) demonstrated a complete loss of target recognition (0% TRP).

Conclusions:

  • p53's target binding is modulated by its activation state, influencing both recognition probability and search strategy.
  • Increased TRP and reliance on 1D sliding characterize the activated p53 mutant.
  • Modulation of TRP upon activation may represent a general mechanism for transcription factor regulation.
Keywords:
mutationp53single moleculetarget bindingtranscription factor

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