Interferons are a diverse family of cytokines which possess

Interferons are a diverse family of cytokines which possess antiviral and immunomodulatory properties (Isaacs and Lindenmann, 1957; Stark et al., 1998; Theofilopoulos et al., 2005). These specialized proteins are produced by cells in response to viral infection. Cellular sensors such as retinoic tropisetron inducible gene I (RIG-I or DDX58) and toll like receptors (TLR3, TLR7 and TLR9) can recognize pathogen associated molecular patterns (PAMPs) that can trigger an innate immune response (Thompson and Locarnini, 2007). Viral specific DNA, RNA and proteins, and microbial products are sensed by TLR and RIG-I resulting in the phosphorylation and activation of interferon regulatory factor 3 (IRF3) and IRF7 (Diebold et al., 2004; Gale and Foy, 2005; Perry et al., 2005). Phosphorylated IRFs and NF-κB translocate to the nucleus and transactivate the expression of IFN-α and IFN-β genes that are critical in the elimination of invading pathogens. By autocrine and paracrine means, the produced interferons serve as ligands for the Janus activated kinase (JAK)–signal transducer and activator of transcription (STAT) pathway (Horvath, 2004). Type I interferons bind to the IFN alpha/beta receptors associated with two tyrosine kinases, JAK1 and TYK2 which phosphorylate STAT1 and STAT2 (Horvath, 2004). Both STAT proteins bind with IRF9 generating the interferon stimulating gene 3 (ISGF3) complex (Kessler et al., 1990; Veals et al., 1993; Darnell et al., 1994; Li et al., 1996). This complex is translocated to the nucleus binding to the IFN-stimulated response element (ISRE) directing the expression of IFN stimulated genes (ISGs) and converting the host cell to an antiviral state (Muller et al., 1994; Haque and Williams, 1998; Liu et al., 2012). ISGs are involved in executing antiviral response against pathogen like hepatitis C virus (Saito et al., 2008). ISGs function in modulating various cellular processes, which includes altering the cell cycle, apoptosis, angiogenesis, metabolism and transcriptional regulation (Balachandran et al., 1998; Tan and Katze, 1999; de Veer et al., 2001). There are over 300 canonical ISGs described and recently genetic screens have been used for identification of ISGs with antiviral functions (Schoggins et al., 2011, 2014). Antiviral ISGs including GBP1, MXA, CNP, cGAS, TAP1, RIG-I, MDA5 (IFIH1), IRF1 and LGP2 (DHX58) are well characterized (Anderson et al., 1999; Kochs and Haller, 1999; Yoneyama et al., 2005; Schoggins et al., 2011; Liu et al., 2012; Wilson et al., 2012; Schoggins et al., 2014). Due to its antiviral properties, IFN-α is a widely used therapeutic agent against HCV and hepatitis B virus (HBV) that cause hepatitis, fibrosis and cirrhosis (Schalm et al., 2000; Manns et al., 2001). Pluripotent stem cell-derived endodermal cells and hepatic lineage cells are currently being tested for regenerating damaged liver in preclinical animal models (Duan et al., 2007; Liu et al., 2011; Liu et al., 2012). During embryonic development, hepatic lineage cells originate from endoderm layer. Growth factors HGF, BMP, Wnt and FGF signaling by mesenchymal cells are critical for liver development (Schmidt et al., 1995; Uehara et al., 1995; Weinstein et al., 2001; Michalopoulos, 2010). Using a combination of these growth factors, pluripotent stem cells can be differentiated into hepatocytes via endoderm (Cai et al., 2007; Rashid et al., 2010; Chen et al., 2012; Cheng et al., 2012). This in vitro platform is useful for understanding the differentiation stage-specific cellular perturbations for external stimuli. Our objective is to investigate the innate immune response exhibited by cells at different phases of hepatic differentiation upon exposure to interferon (IFN-α) stimuli. We used an unbiased discovery based approach to profile ISGs during various hepatic differentiation phases of pluripotent stem cells. We identified both canonical and non-canonical ISGs, including novel ISGs. Functional relevance of selected ISGs was verified during HCV replication in differentiated hepatic cells.