The transcription factors interferon regulatory factor 3 (IRF3) and NF-κB are required for the expression of many genes involved in the innate immune response. Viral infection, or the binding of double-stranded RNA to Toll-like receptor 3, results in the coordinate activation of IRF3 and NF-κB. Activation of IRF3 requires signal-dependent phosphorylation, but little is known about the signaling pathway or kinases involved. Here we report that the noncanonical IκB kinase homologs, IκB kinase-ε (IKKε) and TANK-binding kinase-1 (TBK1), which were previously implicated in NF-κB activation, are also essential components of the IRF3 signaling pathway. Thus, IKKε and TBK1 have a pivotal role in coordinating the activation of IRF3 and NF-κB in the innate immune response.