Abstract 148: Control of NADPH Oxidases Stability and Inflammatory Response by Heat Shock Protein 90
Previously we have shown that Hsp90 binds to NADPH oxidases (Nox) and regulates ROS production. The goals of this study were to determine the mechanisms regulating Hsp90-dependent Nox protein stability and whether inhibition of ROS production accounts for the anti-inflammatory properties of Hsp90 inhibitors. The expression of Nox 1, 2 and 5 proteins were decreased by Hsp90 inhibitors in different cell types (COS-7, HPAEC, HAEC, HL-60, murine macrophages and RAW264.7) and also in intact mouse lung. To determine the mechanisms underlying the decrease in Nox protein stability in the presence of Hsp90 inhibitors, we employed COS-7 cells expressing Nox5. The expression of Nox5 protein, but not mRNA, was robustly decreased by Hsp90 inhibitors. Hsp90 inhibitors promoted Nox5 ubiquitination and degradation which was prevented by inhibitors of the proteasome. Inhibition of Hsp90 upregulates the expression of Hsp70 and we found that Hsp70 binds to Nox5 and can promotes its degradation. The Hsp70-regulated ubiquitin ligase, CHIP associated with Nox5 and promoted increased ubiquitination and degradation. In isolated murine macrophages in vitro, we found that Hsp90 inhibitors robustly decreased the expression of ROS-sensitive inflammatory genes, including MCP-1, TNF-alpha, Cox2, M-CSF and STAT3. In vivo, Hsp90 inhibitors prevented the LPS-induced upregulation of MCP-1 and TNF-alpha in mouse lung. We conclude that Hsp90 binds to Nox and facilities its proper folding and ROS production. In contrast, the binding of Hsp70 and CHIP to Nox proteins induces degradation and reduced ROS. The effects of Hsp90 inhibitors on ROS-sensitive inflammatory proteins suggest that this pathway is a significant mechanism underlying the anti-inflammatory actions and may have important implications in controlling inflammation in cardiovascular diseases.
- © 2012 by American Heart Association, Inc.