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25 Nitric Oxide-cGMP Signalling Promotes B-Catenin Nuclear Translocation and Transcriptional Activity in Endothelial Cells
  1. C M Warboys1,
  2. N Chen1,
  3. Q Zhang1,
  4. Y Shaifta2,
  5. G Vanderslott1,
  6. G Passacquale1,
  7. Y Hu1,
  8. Q Xu1,
  9. J P T Ward2,
  10. A Ferro1
  1. 1Cardiovascular Division, King’s College London, UK
  2. 2Division of Asthma, Allergy and Lung Biology, King’s College London, UK

Abstract

Nitric oxide (NO) derived from endothelial NO synthase (eNOS) exerts cardioprotective effects. eNOS binds to a number of proteins that regulate its function. Using mass spectroscopy to study eNOS immunoprecipitated from human umbilical vein endothelial cells (HUVEC), we identified β-catenin as a novel binding partner of eNOS. This previously unrecognised interaction was confirmed by western blot analyses of both eNOS and β-catenin immunoprecipitates. Further, eNOS activation (using adenosine, salbutamol, histamine or thrombin), application of an NO donor (spermine NONOate) or elevation of cGMP (using sildenafil or 8-bromo-cGMP) all increased nuclear translocation of β-catenin. Nuclear β-catenin activates T cell factor (TCF)/lymphoid enhancing factor (LEF) transcription factors. Application of spermine NONOate or elevation of cGMP increased β-catenin transcriptional activity, as assessed using a luciferase reporter assay in HUVEC transfected with TCF/LEF reporter plasmids. The role of β-catenin in regulating NO-mediated angiogenesis was assessed in wild type and β-catenin-/- mouse pulmonary endothelial cells (MPECs) using an in vitro Matrigel assay. Stimulation with vascular endothelial growth factor (VEGF; NO dependent), spermine NONOate or cGMP elevation increased tube length compared to untreated controls in wild type but not β-catenin-/- MPECs, although both exhibited similar responses to basic fibroblast growth factor (NO independent). Similarly, in C57BL/6 mice, subcutaneous Matrigel plugs containing VEGF and β-catenin siRNA contained fewer endothelial cells compared to plugs containing scrambled siRNA. We conclude that activation of NO-cGMP signalling induces nuclear translocation of β-catenin, which promotes angiogenesis. Whether this contributes to other physiological processes involving NO-mediated transcription remains to be determined.

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