Type I interferon system is essential for anti-viral immune response and is a primary target of viral immune evasion strategies. Influenza A viruses mainly utilize the non-structural protein 1 (NS1) to suppress host immune responses, especially the type I interferon system. Here, we showed influenza infection induces the expression of MAP kinase phosphatase 5 (MKP5), a member of MAP kinase phosphatase (MKP) family, in host cells in a NS1 dependent manner. Mice deficient in MKP5 were resistant to H1N1 influenza virus infection, which was associated with increased IRF3 activation and type I interferon expression compared with WT mice. We further found that MKP5 interacts with and dephosphorylates IRF3 to inhibit its activation. Interestingly, the inhibitory effect of MKP5 on type I interferon expression does not require NS1. Our study, therefore, revealed MKP5 as a negative regulator of IRF3 and a novel mechanism by which influenza viruses inhibit type I interferon expression in host through MKP5.