Upregulation of immunity-related GTPase (IRG) proteins by TNF-alpha in murine astrocytes.

We examined the effect of tumor necrosis factor-alpha (TNF-alpha) on murine primary astrocytes. Proteomic analysis demonstrated that four new spots in the TNF-alpha-treated cells relative to untreated cells. Two of them were identified as Irgb6 and Irgd, members of immunity-related GTPase (IRG) proteins which are the key mediators of interferon-gamma (IFN-gamma)-induced resistance of pathogens in numerous cells. Gene expression analysis using RT-PCR showed that TNF-alpha dose-dependently increased the expression of both proteins. Immunocytochemical analysis showed that TNF-alpha increased the abundance of both proteins. A subcellular localization study demonstrated that TNF-alpha induced the partial colocalization of both proteins with the endoplasmic reticulum (ER) and Golgi apparatus, whereas IFN-gamma did not induce the colocalization of Irgd protein with the ER and Golgi. Combined stimulation with TNF-alpha and IFN-gamma had a synergistic effect on the expression of Irgb6 and an added effect on the expression of Irgd.