IL-9-producing Th9 cells have been associated with autoimmune diseases, such as experimental autoimmune encephalitis. However, the factors that negatively regulate Th9 cells during autoimmune inflammation are unclear. In this article, we show that IFN- inhibits Th9 differentiation both in vitro and in vivo. This suppressive activity was dependent on the transcription factor STAT-1. In addition to its direct inhibitory effect on Th9 differentiation, IFN- suppressed Th9 cells through the induction of IL-27 from dendritic cells. In vitro, treatment of naive CD4(+) T cells with IL-27 suppressed the development of Th9 cells, which was partially dependent on the transcription factors STAT-1 and T-bet. Moreover, IL-27 treatment completely abrogated the encephalitogenicity of Th9 cells in the experimental autoimmune encephalomyelitis model. Thus, our results identify a previously unknown mechanism by which IFN- limits Th9-mediated autoimmune inflammation through dendritic cell modulation of IL-27.