While hundreds of genes are induced by type I interferons, their roles in restricting the influenza life cycle remain mostly unknown. Using a loss-of-function CRISPR screen in cells pre-stimulated with IFNβ, we identified a small number of factors required for restricting influenza A virus replication. In addition to known components of the interferon signaling pathway, we found that Replication Termination Factor 2 (RTF2) restricts influenza at the nuclear stage (and perhaps other stages) of the viral life cycle based on several lines of evidence. First, a deficiency in RTF2 leads to higher levels of viral primary transcription, even in the presence of cycloheximide to block genome replication and secondary transcription. Second, cells that lack RTF2 have enhanced activity of a viral reporter that depends solely on four viral proteins that carry out replication and transcription in the nucleus. Third, when RTF2 protein is mislocalized outside the nucleus, it is not able to restrict replication. Finally, the absence of RTF2 not only leads to enhanced viral transcription but also to reduced expression of anti-viral factors in response to interferon. RTF2 thus inhibits primary influenza transcription, likely acts in the nucleus, and contributes to upregulation of antiviral effectors in response to type I interferons.IMPORTANCE Viral infection triggers the secretion of type I interferons, which in turn induce expression of hundreds of anti-viral genes. However, the roles of these induced genes in controlling viral infections remain largely unknown, limiting our ability to develop host-based antiviral therapeutics against pathogenic viruses such as influenza virus. Here, we performed a loss-of-function genetic CRISPR screen in cells pre-stimulated with type I interferon to identify antiviral genes that restrict influenza A virus replication. Besides finding key components of the interferon signaling pathway, we discovered a new restriction factor, RTF2, which acts in the nucleus, restricts influenza virus transcription, and contributes to the interferon-induced upregulation of known restriction factors. Our work contributes to the field of antiviral immunology by discovering and characterizing a novel restriction factor of influenza, and may ultimately be useful for understanding how to control a virus that causes significant morbidity and mortality worldwide.