Abstract
Background: S-Nitrosoglutathione (GSNO) is an endogenous bronchodilator. GSNO reductase (GSNOR) depletes airway GSNO; inhibitors are in development for asthma. However, GSNO can be tumorigenic by activating wild-type (wt) Ras though S-nitrosylation. We studied whether decreased GSNO reductase promotes wt Ras S-nitrosylation and activation in the lung.
Methods: Lung non-small cell cancer lines (H157; Calu-1; H226;Calu-3; H1650; A549) were studied. Ras S-Nitrosylation was studied by biotin substitution and immunoblot. Active GTP-Ras was assayed in a Raf-1RBD-binding ELISA. Certain cells were exposed to 1) 30 ppm NO in a sealed incubator; or 2) S-nitrosocysteine (CSNO, 10 μM), GSH (2mM) and NADH (300 μM) with or without the GSNO reductase inhibitor, C3 (100 μM; 10 min; from P. Sanghani). GSNO reductase -/- (from J.S. Stamler) were compared with C57Black6 mice with and without intratracheal ethyl nitrite (EtONO, 10 μM).
Results: Wt Ras S-nitrosylation was increased relative to expression in cancer cell lines, though expression of was low. NO exposure for 5d, but not 2d, increased Ras S-nitrosylation relative to expression in vitro. S-Nitrosylation of Ras relative to total Ras expression was increased in the lungs of GSNO reductase -/- mice; and was increased in wt mice by EtONO. CSNO increased Ras activity. In the H1650 cells, this increase was partially augmented by the C3, though baseline expression and activity of GSNO reductase were low.
Conclusions: Though loss of GSNO reductase function may benefit asthma, it prevents wt Ras denitrosylation and inactivation. This effect is associated with tumorigenesis and may not be a good strategy for asthma treatment, particularly in smokers.
- © 2011 ERS