Viral infection of mammalian cells triggers the synthesis and secretion of type I interferons (i.e. IFN-α/β), which induce the transcription of genes that cause cells to adopt an antiviral state. Many viruses have adapted mechanisms to evade IFN-α/β-mediated responses. The leader protein of mengovirus, a picornavirus, has been implicated as an IFN-α/β antagonist. Here, we show that the leader inhibits the transcription of IFN-α/β and that both the presence of a zinc finger motif in its N-terminus and phosphorylation of threonine-47 are required for this function. Transcription of IFN-α/β genes relies on the activity of a number of transcription factors, including interferon regulatory factor 3 (IRF-3). We show that the leader interferes with the transactivation activity of IRF-3 by interfering with its dimerization. Accordingly, mutant viruses with a disturbed leader function were impaired in their ability to suppress IFN-α/β transcription in vivo. By consequence, the leader mutant viruses had an impaired ability to replicate and spread in normal mice but not in IFNAR-KO mice, which are incapable of mounting an IFN-α/β-dependent antiviral response. These results suggest that the leader, by suppressing IRF3-mediated IFN-α/β production, plays an important role in replication and dissemination of mengovirus in its host.