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Glycogen synthase kinase-3beta is activated by matrix metalloproteinase-2 mediated proteolysis in cardiomyoblasts

Arulmozhi D Kandasamy1, Richard Schulz

  • 1Department of Pediatrics, Cardiovascular Research Centre, University of Alberta, 4-62 Heritage Medical Research Centre, Edmonton, AB, Canada T6G 2S2.

Cardiovascular Research
|June 5, 2009
PubMed
Abstract

Insights

Matrix metalloproteinase-2 (MMP-2) cleaves glycogen synthase kinase-3beta (GSK-3beta), enhancing its activity. This MMP-2 mediated GSK-3beta activation may contribute to cardiac oxidative stress injury.

Area of Science:

  • Cardiovascular Biology
  • Enzymology
  • Proteolysis

Background:

  • Matrix metalloproteinase-2 (MMP-2) degrades cardiac proteins during oxidative stress.
  • Glycogen synthase kinase-3beta (GSK-3beta) is dysregulated and proteolytically cleaved under oxidative stress conditions.

Purpose of the Study:

  • To investigate if GSK-3beta is a substrate for MMP-2 following oxidative stress.
  • To determine the impact of MMP-2 cleavage on GSK-3beta activity.

Main Methods:

  • Incubation of purified MMP-2 and GSK-3beta, followed by mass spectrometry (LC-MS/MS) to identify cleavage products.
  • Measurement of GSK-3beta kinase activity using a peptide substrate and [gamma-(32)P]-ATP.
  • Induction of oxidative stress in H9c2 cardiomyoblasts using H(2)O(2) to assess MMP-2 and GSK-3beta levels and activities.

Main Results:

  • MMP-2 directly cleaved GSK-3beta in a time- and concentration-dependent manner, producing an N-terminal fragment.
  • GSK-3beta kinase activity significantly increased upon incubation with MMP-2, an effect blocked by MMP inhibitors.
  • Oxidative stress in H9c2 cells increased MMP-2 activity and levels, decreased GSK-3beta levels, and elevated GSK-3beta kinase activity, with MMP inhibitors mitigating these changes.

Conclusions:

  • GSK-3beta is a novel substrate for MMP-2, with cleavage enhancing its kinase activity.
  • MMP-2 likely cleaves the N-terminal inhibitory phosphorylation site (serine-9) of GSK-3beta.
  • MMP-2-induced augmentation of GSK-3beta activity may contribute to cardiac injury in conditions of heightened oxidative stress.

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