Although the role of human IRF-5 in antiviral and inflammatory responses in vitro has been well characterized, much remains to be elucidated about murine IRF-5. Murine IRF-5, unlike the heavily spliced human gene, is primarily expressed as a full-length transcript, with only a single splice variant that was detected in very low levels in the bone marrow of C57BL/6J mice. This bone marrow variant contains a 288-nucleotide deletion from exons 4-6 and exhibits impaired transcriptional activity. The murine IRF-5 can be activated by both TBK1 and MyD88 to form homodimers and bind to and activate transcription of type I interferon and inflammatory cytokine genes. The importance of IRF-5 in the antiviral and inflammatory response in vivo is highlighted by marked reductions in serum levels of type I interferon and tumor necrosis factor alpha (TNFalpha) in Newcastle disease virus-infected Irf5(-)(/)(-) mice. IRF-5 is critical for TLR3-, TLR4-, and TLR9-dependent induction of TNFalpha in CD11c(+) dendritic cells. In contrast, TLR9, but not TLR3/4-mediated induction of type I IFN transcription, is dependent on IRF-5 in these cells. In addition, IRF-5 regulates TNFalpha but not type I interferon gene transcription in Newcastle disease virus-infected peritoneal macrophages. Altogether, these data reveal the cell type-specific importance of IRF-5 in MyD88-mediated antiviral pathways and the widespread role of IRF-5 in the regulation of inflammatory cytokines.