We demonstrate that novel bat HL17NL10 and HL18NL11 influenza virus NS1 proteins are effective interferon-antagonists, but do not block general host gene expression. Solving the RNA-binding domain structures revealed the canonical NS1 symmetrical homodimer, and RNA-binding required conserved basic residues in this domain. Interferon-antagonism was strictly dependent on RNA-binding, and chimeric bat influenza viruses expressing NS1s defective in this activity were highly attenuated in interferon-competent cells, but not in cells unable to establish antiviral immunity