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In Vitro Analysis of E3 Ubiquitin Ligase Function
06:06

In Vitro Analysis of E3 Ubiquitin Ligase Function

Published on: May 14, 2021

HectD1 E3 ligase modifies adenomatous polyposis coli (APC) with polyubiquitin to promote the APC-axin interaction

Hoanh Tran1, Daisy Bustos, Ronald Yeh

  • 1Department of Cancer Targets, Genentech Inc., South San Francisco, California 94080, USA. tran.hoanh@gene.com

The Journal of Biological Chemistry
|January 2, 2013
PubMed

Insights

HectD1 E3 ubiquitin ligase promotes the adenomatous polyposis coli (APC) protein interaction with Axin. This interaction is crucial for the negative regulation of Wnt signaling, impacting beta-catenin stability.

Area of Science:

  • Molecular Biology
  • Cell Biology
  • Biochemistry

Background:

  • The adenomatous polyposis coli (APC) protein is a key negative regulator of the Wnt signaling pathway.
  • APC forms a destruction complex with Axin, CK1α, and GSK3β to target β-catenin for degradation.
  • The role of Lys-63-linked ubiquitylation on APC within the destruction complex is not fully understood.

Purpose of the Study:

  • To identify the E3 ubiquitin ligase responsible for Lys-63-linked ubiquitylation of APC.
  • To elucidate the functional consequence of HectD1-mediated APC ubiquitylation on Wnt signaling.

Main Methods:

  • Identification of HectD1 as an E3 ubiquitin ligase for APC.
  • Knockdown of HectD1 expression.
  • Assessment of APC ubiquitylation and APC-Axin interaction.
  • Analysis of Wnt signaling pathway activation, including β-catenin stabilization.

Main Results:

  • HectD1 was identified as an E3 ubiquitin ligase that modifies APC with Lys-63 polyubiquitin.
  • HectD1 knockdown reduced APC ubiquitylation and disrupted the APC-Axin interaction.
  • Loss of HectD1 function led to increased Wnt3a-induced β-catenin stabilization and signaling.

Conclusions:

  • HectD1 promotes the APC-Axin interaction through Lys-63-linked ubiquitylation.
  • HectD1 acts as a negative regulator of Wnt signaling by facilitating the APC-Axin interaction.

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