Bidirectional Cross-Regulation between the Endothelial Nitric Oxide Synthase and ?-Catenin Signaling Pathways.
Cardiovasc Res. 2014 Jul 25;
Authors: Warboys CM, Chen N, Zhang Q, Shaifta Y, Vanderslott G, Passacquale G, Hu Y, Xu Q, Ward JP, Ferro A
Abstract
AIMS: ?-catenin has been shown to be regulated by inducible nitric oxide synthase (NOS) in endothelial cells. We investigated here whether ?-catenin interacts with and regulates endothelial NOS (eNOS) and whether eNOS activation promotes ?-catenin signaling.
METHODS AND RESULTS: We identified ?-catenin as a novel eNOS binding protein in human umbilical vein endothelial cells (HUVECs) by mass spectroscopy and western blot analyses of ?-catenin and eNOS immunoprecipitates. This was confirmed by in situ proximity ligation assay. eNOS activity, assessed by cGMP production and eNOS phosphorylation (Ser1177), was enhanced in ?-catenin(-/-) mouse pulmonary endothelial cells (MPECs) relative to wild type MPECs. eNOS activation (using adenosine, salbutamol, thrombin or histamine), or application of an NO donor (spermine NONOate) or cGMP-analogue (8-bromo-cGMP) caused nuclear translocation of ?-catenin in HUVEC as shown by western blotting of nuclear extracts. Exposure to spermine NONOate, 8-bromo-cGMP or sildenafil (a phosphodiesterase type 5 inhibitor) also increased the expression of ?-catenin-dependent transcripts, IL-8 and cyclin D1. Stimulation of wild type MPECs with basic fibroblast growth factor (bFGF), vascular endothelial growth factor (VEGF), spermine NONOate, 8-bromo-cGMP or sildenafil increased tube length relative to controls in an angiogenesis assay. These responses were abrogated in ?-catenin(-/-) MPECs, with the exception of that to bFGF which is NO-independent. In C57BL/6 mice, subcutaneous VEGF-supplemented Matrigel plugs containing ?-catenin(-/-) MPECs exhibited reduced angiogenesis compared to plugs containing wild type MPECs. Angiogenesis was not altered in bFGF-supplemented Matrigel.
CONCLUSIONS: These data reveal bidirectional cross talk and regulation between the NO-cGMP and ?-catenin signaling pathways.
PMID: 25062958 [PubMed – as supplied by publisher]
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