H epatitis C virus (HCV) is one of the most important pathogens causing liver-related morbidity and mortality. 1 Hepatitis C is characterized by persistent infection of the liver, leading to the development of chronic hepatitis, liver cirrhosis, and hepatocellular carcinoma. Type-I interferon (IFN) plays a central role in eliminating viruses, not only by way of therapeutic applications 2 but also as a natural cellular antiviral mechanism. 3,4 Interferons are produced naturally in response to virus infection and to cellular exposure to IFN itself. Binding of the IFNs to their receptors activates the Jak-STAT pathway to form a complex with IFN-stimulated gene factor-3 (ISGF3), which translocates to the nucleus, binds the IFN-stimulated response element (ISRE) located in the promoter/enhancer region of the IFN-stimulated genes (ISGs), and activates expression of ISGs.HCV subgenomic replicons constitute in-vitro models that simulate cellular autonomous replication of HCV

Gene alterations in TERT promoter, TP53, CTNNB1, and HBV integration were closely associated with HCC development, and mutations in TERT promoter are related to poor prognosis. These results are useful for understanding the underlying mechanism of hepatocarcinogenesis, diagnosis, and predicting outcomes of patients with HCC.

Hepatitis C virus (HCV) infection blocks cellular interferon (IFN)-mediated antiviral signaling through cleavage of Cardif by HCV-NS3/4A serine protease. Like NS3/4A, NS4B protein strongly blocks IFN-b production signaling mediated by retinoic acid-inducible gene I (RIG-I); however, the underlying molecular mechanisms are not well understood. Recently, the stimulator of interferon genes (STING) was identified as an activator of RIG-I signaling. STING possesses a structural homology domain with flaviviral NS4B, which suggests a direct protein-protein interaction. In the present study, we investigated the molecular mechanisms by which NS4B targets RIG-I-induced and STING-mediated IFN-b production signaling. IFN-b promoter reporter assay showed that IFN-b promoter activation induced by RIG-I or Cardif was significantly suppressed by both NS4B and NS3/4A, whereas STING-induced IFN-b activation was suppressed by NS4B but not by NS3/4A, suggesting that NS4B had a distinct point of interaction. Immunostaining showed that STING colocalized with NS4B in the endoplasmic reticulum. Immunoprecipitation and bimolecular fluorescence complementation (BiFC) assays demonstrated that NS4B specifically bound STING. Intriguingly, NS4B expression blocked the protein interaction between STING and Cardif, which is required for robust IFN-b activation. NS4B truncation assays showed that its N terminus, containing the STING homology domain, was necessary for the suppression of IFN-b promoter activation. NS4B suppressed residual IFN-b activation by an NS3/4A-cleaved Cardif (Cardif1-508), suggesting that NS3/4A and NS4B may cooperate in the blockade of IFN-b production. Conclusion: NS4B suppresses RIG-I-mediated IFN-b production signaling through a direct protein interaction with STING. Disruption of that interaction may restore cellular antiviral responses and may constitute a novel therapeutic strategy for the eradication of HCV. (HEPATOLOGY 2013;57:46-58)

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Splicing of messenger RNAs is regulated by site-specific binding of members of the serine-arginine-rich (SR) protein family, and SR protein kinases (SRPK) 1 and 2 regulate overall activity of the SR proteins by phosphorylation of their RS domains. We have reported that specifically designed SRPK inhibitors suppressed effectively several DNA and RNA viruses in vitro and in vivo. Here, we show that an SRPK inhibitor, SRPIN340, suppressed in a dose-dependent fashion expression of a hepatitis C virus (HCV) subgenomic replicon and replication of the HCV-JFH1 clone in vitro. The inhibitory effects were not associated with antiproliferative or nonspecific cytotoxic effects on the host cells. Overexpression of SRPK1 or SRPK2 resulted in augmentation of HCV replication, while small interfering RNA (siRNA) knockdown of the SRPKs suppressed HCV replication significantly. Immunocytochemistry showed that SRPKs and the HCV core and NS5A proteins colocalized to some extent in the perinuclear area. Our results demonstrate that SRPKs are host factors essential for HCV replication and that functional inhibitors of these kinases may constitute a new class of antiviral agents against HCV infection.Hepatitis C virus (HCV) infects up to 170 million people worldwide, and these infections frequently are characterized by chronic liver inflammation, leading to decompensated liver cirrhosis and hepatocellular cancers (1). Alpha and beta interferons are the mainstay of HCV therapeutics. However, the most effective pegylated interferon plus ribavirin combination therapies can eliminate HCV from around half of the patients only (6). These difficulties in eradicating HCV are compounded by the limited treatment options. For this reason, the development of safe and effective therapeutic agents against HCV has been a strong motivation in academia and industry (23).Serine-arginine-rich (SR) proteins are a family of non-small nuclear ribonucleoprotein particle (non-snRNP) splicing factors that are highly conserved throughout the eukaryotes. They harbor one or two RNA recognition motifs and an RS domain at the amino and carboxyl termini, respectively (29). RS domains consist of multiple consecutive Arg-Ser/Ser-Arg dipeptide repeats, in which the Ser residues are extensively phosphorylated by several kinases, including SR protein kinases (SRPKs). SRPK1 was the first SR protein kinase to be cloned, on the basis of its ability to phosphorylate SR proteins in vitro (8, 9), and two other structurally related kinases, SRPK2 and SRPK3, also have been shown to phosphorylate SR proteins (16,31). Although the precise physiological role of this phosphorylation remains unknown, it is expected that phosphorylation of SR proteins affects their protein-protein and protein-RNA interactions, intracellular localization and trafficking, and alternative splicing of pre-mRNA (21).As SRPK-dependent herpes simplex virus (HSV) splicing and SRPK-mediated phosphorylation of hepatitis B virus (HBV) core protein have been reported (4,25,33), it is reasonable to expect that SR...