O-GlcNAcylation contributes to the vascular effects of ET-1 via activation of the RhoA/Rho-kinase pathway

Cardiovasc Res. 2011 Feb 15;89(3):614-22. doi: 10.1093/cvr/cvq338. Epub 2010 Oct 26.

Abstract

Aims: Glycosylation with β-N-acetylglucosamine (O-GlcNAcylation) is one of the most complex post-translational modifications. The cycling of O-GlcNAc is controlled by two enzymes: UDP-NAc transferase (OGT) and O-GlcNAcase (OGA). We recently reported that endothelin-1 (ET-1) augments vascular levels of O-GlcNAcylated proteins. Here we tested the hypothesis that O-GlcNAcylation contributes to the vascular effects of ET-1 via activation of the RhoA/Rho-kinase pathway.

Methods and results: Incubation of vascular smooth muscle cells (VSMCs) with ET-1 (0.1 μM) produces a time-dependent increase in O-GlcNAc levels. ET-1-induced O-GlcNAcylation is not observed when VSMCs are previously transfected with OGT siRNA, treated with ST045849 (OGT inhibitor) or atrasentan (ET(A) antagonist). ET-1 as well as PugNAc (OGA inhibitor) augmented contractions to phenylephrine in endothelium-denuded rat aortas, an effect that was abolished by the Rho kinase inhibitor Y-27632. Incubation of VSMCs with ET-1 increased expression of the phosphorylated forms of myosin phosphatase target subunit 1 (MYPT-1), protein kinase C-potentiated protein phosphatase 1 inhibitor protein (protein kinase C-potentiated phosphatase inhibitor-17), and myosin light chain (MLC) and RhoA expression and activity, and this effect was abolished by both OGT siRNA transfection or OGT inhibition and atrasentan. ET-1 also augmented expression of PDZ-Rho GEF (guanine nucleotide exchange factor) and p115-Rho GEF in VSMCs and this was prevented by OGT siRNA, ST045849, and atrasentan.

Conclusion: We suggest that ET-1 augments O-GlcNAcylation and this modification contributes to increased vascular contractile responses via activation of the RhoA/Rho-kinase pathway.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Acetylglucosamine / metabolism*
  • Acylation / physiology
  • Animals
  • Aorta, Thoracic / cytology
  • Cardiovascular Diseases / metabolism
  • Cells, Cultured
  • Endothelin-1 / metabolism*
  • Endothelin-1 / pharmacology
  • Male
  • Muscle, Smooth, Vascular / cytology
  • Muscle, Smooth, Vascular / metabolism*
  • Phosphorylation / drug effects
  • Phosphorylation / physiology
  • Rats
  • Rats, Wistar
  • Signal Transduction / drug effects
  • Signal Transduction / physiology
  • Vasoconstriction / drug effects
  • Vasoconstriction / physiology*
  • rho-Associated Kinases / metabolism*
  • rhoA GTP-Binding Protein / metabolism*

Substances

  • Endothelin-1
  • rho-Associated Kinases
  • rhoA GTP-Binding Protein
  • Acetylglucosamine