Negative regulation of prolactin receptor stability and signaling mediated by SCF(beta-TrCP) E3 ubiquitin ligase

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Chemical Inactivation of the E3 Ubiquitin Ligase Cereblon by Pomalidomide-based Homo-PROTACs
10:44

Chemical Inactivation of the E3 Ubiquitin Ligase Cereblon by Pomalidomide-based Homo-PROTACs

Published on: May 15, 2019

Ying Li1, K G Kuresh Kumar, Weigang Tang

  • 1Department of Animal Biology, University of Pennsylvania, Philadelphia, Pennsylvania 19104, USA.

Molecular and Cellular Biology
|April 15, 2004
PubMed

Insights

Prolactin receptor (PRLR) degradation is regulated by beta-TrCP, an E3 ubiquitin ligase. Inhibiting beta-TrCP enhances prolactin signaling and cell growth, revealing a new control point for hormone action.

Area of Science:

  • Cell Biology
  • Endocrinology
  • Molecular Biology

Background:

  • Hormone receptor regulation is crucial for cellular responses.
  • Ubiquitin-dependent degradation controls hormone signaling duration and magnitude.
  • Prolactin (PRL) signaling, mediated by its receptor (PRLR), impacts cell differentiation, proliferation, and survival.

Purpose of the Study:

  • To investigate the role of ubiquitin-dependent degradation in regulating prolactin receptor (PRLR) stability and signaling.
  • To identify the E3 ubiquitin ligase involved in PRLR degradation.
  • To elucidate the mechanism by which PRLR is targeted for degradation.

Main Methods:

  • Co-immunoprecipitation assays to detect protein interactions.
  • Site-directed mutagenesis to create PRLR phosphorylation site mutants.
  • In vitro ubiquitination assays using purified SCF(beta-TrCP) ligase.
  • Western blotting to assess protein levels and phosphorylation.
  • siRNA-mediated knockdown of beta-TrCP.
  • Cellular proliferation assays.

Main Results:

  • PRL stimulation promotes interaction between PRLR and beta-TrCP2, an F-box protein subunit of the SCF(beta-TrCP) E3 ubiquitin ligase.
  • Phosphorylation of PRLR at serine 349 is required for beta-TrCP2 interaction and subsequent ubiquitination.
  • A mutant PRLR (S349A) resistant to ubiquitination exhibited increased stability.
  • Knockdown of beta-TrCP inhibited PRLR ubiquitination and degradation, enhancing PRL-induced Stat5 activation and transcription.
  • PRLR activation of Stat5 and PRL-stimulated cell growth were augmented in cells expressing the S349A mutant.

Conclusions:

  • PRLR is a novel substrate of the SCF(beta-TrCP) E3 ubiquitin ligase.
  • beta-TrCP acts as a negative regulator of PRL signaling by promoting PRLR degradation.
  • Targeting the beta-TrCP-PRLR interaction could offer therapeutic strategies for modulating PRL-dependent processes.

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