Persistent infection with hepatitis C virus (HCV) can lead to chronic hepatitis, liver cirrhosis, and hepatocellular carcinoma. All current therapies of hepatitis C include interferon‐alpha (IFN‐α). Moreover, IFN‐gamma (IFN‐γ), the only type II IFN, strongly inhibits HCV replication in vitro and is the primary mediator of HCV‐specific antiviral T‐cell responses. However, for both cytokines the precise set of effector protein(s) responsible for replication inhibition is not known. The aim of this study was the identification of IFN‐α and IFN‐γ stimulated genes (ISGs) responsible for controlling HCV replication. We devised an RNA interference (RNAi)‐based “gain of function” screen and identified, in addition to known ISGs earlier reported to suppress HCV replication, several new ones with proven antiviral activity. These include IFIT3 (IFN‐induced protein with tetratricopeptide repeats 3), TRIM14 (tripartite motif containing 14), PLSCR1 (phospholipid scramblase 1), and NOS2 (nitric oxide synthase 2, inducible). All ISGs identified in this study were up‐regulated both by IFN‐α and IFN‐γ, demonstrating a substantial overlap of HCV‐specific effectors induced by either cytokine. Nevertheless, some ISGs were more specific for IFN‐α or IFN‐γ, which was most pronounced in case of PLSCR1 and NOS2 that were identified as main effectors of IFN‐γ‐mediated anti‐HCV activity. Combinatorial knockdowns of ISGs suggest additive or synergistic effects demonstrating that with either IFN, inhibition of HCV replication is caused by the combined action of multiple ISGs. Conclusion: Our study identifies a number of novel ISGs contributing to the suppression of HCV replication by type I and type II IFN. We demonstrate a substantial overlap of antiviral programs triggered by either cytokine and show that suppression of HCV replication is mediated by the concerted action of multiple effectors. (HEPATOLOGY 2012;56:2082–2093)