Involvement of oxidative and nitrosative stress in modulation of gene expression and functional responses by IFNgamma.

IFNgamma is a potent immunomodulator which plays important roles in host defense. IFNgamma modulates transcription of growth-related genes [N-myc downstream regulator 1, growth arrest and DNA damage inducible gamma and inhibitor of DNA binding 2 (Id2)], which is followed by increased growth suppression in the mouse hepatoma cell line, H6. Further studies revealed modulation of genes involved in oxidative and nitrosative stress (iNos, gp91phox and Catalase) and increased generation of reactive oxygen species (ROS) and reactive nitrogen intermediates (RNIs) upon IFNgamma treatment. High amounts of ROS and RNI are responsible for IFNgamma-mediated reduction in cell growth as this process is blocked, using either diphenylene iodonium (DPI), an inhibitor of flavin-containing NADPH oxidases, or N-methyl L-arginine (LNMA), an inhibitor of nitric oxide synthase. Based on studies with LNMA and DPI, IFNgamma-modulated genes can be categorized into two distinct sets: oxidative and nitrosative stress independent (transporter associated with antigen processing 2, Cd80, Lmp10 and Icosl) and oxidative and nitrosative stress dependent (iNos, gp91phox, Catalase and Id2). In addition, DPI or LNMA blocked IFNgamma-induced activation of Ras, demonstrating the involvement of oxidative and nitrosative stress. Manumycin A, a farnesyl transferase inhibitor, blocked Ras activation and reduced NADPH oxidase activity and ROS amounts leading to increased cell growth in the presence of IFNgamma. Notably, the IFNgamma-induced MHC class I levels are not modulated in cells treated with DPI, LNMA or manumycin A. Together, these results delineate the role of high amounts of ROS, RNI and Ras activation in modulating expression of some genes and, thereby, function by IFNgamma. The implications of these results during modulation of immune responses by IFNgamma are discussed.