Research ArticleVASCULAR BIOLOGY

Stromal cell–derived factor 2 is critical for Hsp90-dependent eNOS activation

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Sci. Signal.  18 Aug 2015:
Vol. 8, Issue 390, pp. ra81
DOI: 10.1126/scisignal.aaa2819

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Maximizing nitric oxide production

The gas nitric oxide causes blood vessels to relax and blood pressure to drop. In endothelial cells, the enzyme eNOS produces nitric oxide. The chaperone protein Hsp90 promotes the activation of eNOS by enhancing its phosphorylation. Siragusa et al. found that SDF2 bound eNOS, enhancing the phosphorylation of eNOS and NO production by promoting the binding of eNOS to Hsp90.

Abstract

Endothelial nitric oxide synthase (eNOS) catalyzes the conversion of l-arginine and molecular oxygen into l-citrulline and nitric oxide (NO), a gaseous second messenger that influences cardiovascular physiology and disease. Several mechanisms regulate eNOS activity and function, including phosphorylation at Ser and Thr residues and protein-protein interactions. Combining a tandem affinity purification approach and mass spectrometry, we identified stromal cell–derived factor 2 (SDF2) as a component of the eNOS macromolecular complex in endothelial cells. SDF2 knockdown impaired agonist-stimulated NO synthesis and decreased the phosphorylation of eNOS at Ser1177, a key event required for maximal activation of eNOS. Conversely, SDF2 overexpression dose-dependently increased NO synthesis through a mechanism involving Akt and calcium (induced with ionomycin), which increased the phosphorylation of Ser1177 in eNOS. NO synthesis by iNOS (inducible NOS) and nNOS (neuronal NOS) was also enhanced upon SDF2 overexpression. We found that SDF2 was a client protein of the chaperone protein Hsp90, interacting preferentially with the M domain of Hsp90, which is the same domain that binds to eNOS. In endothelial cells exposed to vascular endothelial growth factor (VEGF), SDF2 was required for the binding of Hsp90 and calmodulin to eNOS, resulting in eNOS phosphorylation and activation. Thus, our data describe a function for SDF2 as a component of the Hsp90-eNOS complex that is critical for signal transduction in endothelial cells.

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