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

Victor V. Lima, Fernanda R. Giachini, Fernando S. Carneiro, Maria Helena C. Carvalho, Zuleica B. Fortes, R. Clinton Webb, Rita C. Tostes

Research output: Contribution to journalArticle

28 Citations (Scopus)

Abstract

AimsGlycosylation 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 resultsIncubation 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 (ETA 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.ConclusionWe suggest that ET-1 augments O-GlcNAcylation and this modification contributes to increased vascular contractile responses via activation of the RhoA/Rho-kinase pathway.

Original languageEnglish (US)
Pages (from-to)614-622
Number of pages9
JournalCardiovascular Research
Volume89
Issue number3
DOIs
StatePublished - Feb 15 2011

Fingerprint

rho-Associated Kinases
Endothelin-1
Blood Vessels
Vascular Smooth Muscle
Smooth Muscle Myocytes
Rho Guanine Nucleotide Exchange Factors
Small Interfering RNA
Protein Kinase C
Myosin-Light-Chain Phosphatase
Myosin Light Chains
Uridine Diphosphate
Acetylglucosamine
Phenylephrine
Post Translational Protein Processing
Transferases
Phosphoric Monoester Hydrolases
Endothelium
Transfection
Aorta
Proteins

Keywords

  • Endothelin-1
  • RhoA/Rho-kinase pathway
  • Vascular reactivity
  • β-N-acetylglucosamine

ASJC Scopus subject areas

  • Physiology
  • Cardiology and Cardiovascular Medicine
  • Physiology (medical)

Cite this

Lima, V. V., Giachini, F. R., Carneiro, F. S., Carvalho, M. H. C., Fortes, Z. B., Webb, R. C., & Tostes, R. C. (2011). O-GlcNAcylation contributes to the vascular effects of ET-1 via activation of the RhoA/Rho-kinase pathway. Cardiovascular Research, 89(3), 614-622. https://doi.org/10.1093/cvr/cvq338

O-GlcNAcylation contributes to the vascular effects of ET-1 via activation of the RhoA/Rho-kinase pathway. / Lima, Victor V.; Giachini, Fernanda R.; Carneiro, Fernando S.; Carvalho, Maria Helena C.; Fortes, Zuleica B.; Webb, R. Clinton; Tostes, Rita C.

In: Cardiovascular Research, Vol. 89, No. 3, 15.02.2011, p. 614-622.

Research output: Contribution to journalArticle

Lima, VV, Giachini, FR, Carneiro, FS, Carvalho, MHC, Fortes, ZB, Webb, RC & Tostes, RC 2011, 'O-GlcNAcylation contributes to the vascular effects of ET-1 via activation of the RhoA/Rho-kinase pathway', Cardiovascular Research, vol. 89, no. 3, pp. 614-622. https://doi.org/10.1093/cvr/cvq338
Lima, Victor V. ; Giachini, Fernanda R. ; Carneiro, Fernando S. ; Carvalho, Maria Helena C. ; Fortes, Zuleica B. ; Webb, R. Clinton ; Tostes, Rita C. / O-GlcNAcylation contributes to the vascular effects of ET-1 via activation of the RhoA/Rho-kinase pathway. In: Cardiovascular Research. 2011 ; Vol. 89, No. 3. pp. 614-622.
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abstract = "AimsGlycosylation 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 resultsIncubation 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 (ETA 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.ConclusionWe suggest that ET-1 augments O-GlcNAcylation and this modification contributes to increased vascular contractile responses via activation of the RhoA/Rho-kinase pathway.",
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T1 - O-GlcNAcylation contributes to the vascular effects of ET-1 via activation of the RhoA/Rho-kinase pathway

AU - Lima, Victor V.

AU - Giachini, Fernanda R.

AU - Carneiro, Fernando S.

AU - Carvalho, Maria Helena C.

AU - Fortes, Zuleica B.

AU - Webb, R. Clinton

AU - Tostes, Rita C.

PY - 2011/2/15

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N2 - AimsGlycosylation 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 resultsIncubation 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 (ETA 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.ConclusionWe suggest that ET-1 augments O-GlcNAcylation and this modification contributes to increased vascular contractile responses via activation of the RhoA/Rho-kinase pathway.

AB - AimsGlycosylation 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 resultsIncubation 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 (ETA 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.ConclusionWe suggest that ET-1 augments O-GlcNAcylation and this modification contributes to increased vascular contractile responses via activation of the RhoA/Rho-kinase pathway.

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KW - Vascular reactivity

KW - β-N-acetylglucosamine

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