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Application of Genetically Encoded Fluorescent Nitric Oxide (NO•) Probes, the geNOps, for Real-time Imaging of NO• Signals in Single Cells
Published on: March 16, 2017
AMP-activated protein kinase functionally phosphorylates endothelial nitric oxide synthase Ser633
Zhen Chen1, I-Chen Peng, Wei Sun
1Division of Biomedical Sciences, University of California, Riverside, CA 92521, USA.
Circulation Research
|January 10, 2009
Summary
AMP-activated protein kinase (AMPK) phosphorylates endothelial nitric oxide synthase (eNOS) at Ser633, a key event for nitric oxide (NO) production and cardiovascular homeostasis. This finding reveals a novel signaling pathway regulating vascular function.
Area of Science:
- Biochemistry
- Molecular Biology
- Cardiovascular Research
Background:
- Endothelial nitric oxide synthase (eNOS) regulates cardiovascular homeostasis through nitric oxide (NO) bioavailability.
- eNOS activity is modulated by posttranslational modifications, particularly phosphorylation.
- AMP-activated protein kinase (AMPK) activity increases in endothelial cells (ECs) during cardiovascular events.
Purpose of the Study:
- To investigate the phosphorylation of eNOS Ser633 by AMPK.
- To determine the functional relevance of this phosphorylation in ECs and mouse models.
- To elucidate the role of AMPK-eNOS signaling in NO production and cardiovascular health.
Main Methods:
- Utilized mouse models with genetic ablation or pharmacological inhibition of AMPK.
- Employed shear stress, atorvastatin, and adiponectin as stimuli.
- Generated gain-/loss-of-function eNOS mutants.
- Performed nano-liquid chromatography/tandem mass spectrometry (LC/MS/MS) for phosphorylation site confirmation.
Main Results:
- Shear stress, atorvastatin, and adiponectin increased AMPK and eNOS Ser633 phosphorylation, dependent on AMPK activity.
- Constitutively active AMPK or agonists induced sustained eNOS Ser633 phosphorylation.
- eNOS Ser633 phosphorylation was crucial for NO production.
- AMPKalpha2 deficiency attenuated atorvastatin-induced eNOS phosphorylation.
- LC/MS/MS confirmed eNOS Ser633 phosphorylation and competition with other sites.
Conclusions:
- AMPK directly phosphorylates eNOS at Ser633.
- This phosphorylation event is functionally significant for NO bioavailability in ECs.
- The AMPK-eNOS Ser633 pathway represents a novel target for modulating cardiovascular function.
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