Many viruses target signal transducer and activator of transcription (STAT) 1 and STAT2 to antagonise antiviral interferon signalling, but targeting of other STATs such as STAT3, a pleiotropic molecule that mediates signalling by diverse cytokines, is poorly understood. Here, using virus infection, quantitative
live cell imaging, innate immune signalling and protein interaction assays, and complementation/depletion of STAT expression, we show that STAT3 antagonism is conserved among P-proteins of diverse pathogenic lyssaviruses and correlates with pathogenesis. We also find that another negative sense RNA virus, Ebola virus, targets STAT3. Importantly, targeting of STAT3 differs between viruses, such that some viruses use highly selective targeting of specific cytokine-activated dimers, variously using mechanisms such as direct interaction with specific STATs, antagonism of nuclear trafficking pathways, or combinations of both. RT-qPCR and reporter gene assays indicate that this results in specific modulation of cytokine pathways, effecting differential antagonism of target genes. These data provide novel insights into mechanisms by which viruses can modulate cellular function to support infection through discriminatory targeting of immune signalling complexes. The findings also highlight the potential application of selective interferon-antagonists as tools to delineate signalling by particular STAT complexes, significant not only to pathogen-host interactions but also cell physiology, development and cancer.