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Purification of Ubiquitinated p53 Proteins from Mammalian Cells
10:55

Purification of Ubiquitinated p53 Proteins from Mammalian Cells

Published on: March 21, 2022

Regulation of the MDM2-p53 pathway by ribosomal protein L11 involves a post-ubiquitination mechanism

Mu-Shui Dai1, Dingding Shi, Yetao Jin

  • 1Department of Biochemistry and Molecular Biology, School of Medicine, Oregon Health and Science University, Portland, Oregon 97239.

The Journal of Biological Chemistry
|June 29, 2006
PubMed

Insights

Ribosomal protein L11 uniquely inhibits MDM2 turnover by preventing proteasomal degradation of ubiquitinated MDM2. This post-ubiquitination mechanism differentiates L11 from L5 and L23, impacting MDM2-p53 feedback loop regulation.

Area of Science:

  • Molecular Biology
  • Cellular Biology
  • Biochemistry

Background:

  • The MDM2-p53 feedback loop is crucial for regulating p53 stability during cellular stress.
  • Ribosomal proteins L5, L11, and L23 can inhibit this loop, particularly under ribosomal stress, by affecting MDM2-mediated p53 ubiquitination and degradation.

Purpose of the Study:

  • To investigate the differential effects of ribosomal proteins L5, L11, and L23 on MDM2 regulation.
  • To elucidate the specific mechanism by which L11 influences MDM2 levels and activity.

Main Methods:

  • Western blotting to detect ubiquitinated and native MDM2.
  • In vitro assays to assess proteasome-mediated degradation.
  • Cellular half-life measurements of MDM2.

Main Results:

  • Ribosomal protein L11, unlike L5 and L23, causes significant accumulation of ubiquitinated and native MDM2.
  • This effect relies on MDM2's ubiquitin ligase activity and L11's central binding domain (residues 51-108).
  • L11 inhibits the in vitro 26 S proteasome degradation of ubiquitinated MDM2 and prolongs MDM2's half-life in cells.

Conclusions:

  • L11 differentially regulates ubiquitinated p53 and MDM2 levels compared to L5 and L23.
  • L11 inhibits MDM2 turnover and activity via a post-ubiquitination mechanism, distinct from previously understood pathways.

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