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http://dx.doi.org/10.14348/molcells.2022.0018

Hepatitis C Virus Nonstructural Protein 5A Interacts with Immunomodulatory Kinase IKKε to Negatively Regulate Innate Antiviral Immunity  

Kang, Sang-Min (Laboratory for Infectious Disease Prevention, Korea Zoonosis Research Institute, Jeonbuk National University)
Park, Ji-Young (Division of Chronic Viral Disease, Center for Emerging Virus Research, National Institute of Infectious Diseases, National Institute of Health, Korea Disease Control and Prevention Agency)
Han, Hee-Jeong (Laboratory for Infectious Disease Prevention, Korea Zoonosis Research Institute, Jeonbuk National University)
Song, Byeong-Min (Laboratory for Infectious Disease Prevention, Korea Zoonosis Research Institute, Jeonbuk National University)
Tark, Dongseob (Laboratory for Infectious Disease Prevention, Korea Zoonosis Research Institute, Jeonbuk National University)
Choi, Byeong-Sun (Division of Chronic Viral Disease, Center for Emerging Virus Research, National Institute of Infectious Diseases, National Institute of Health, Korea Disease Control and Prevention Agency)
Hwang, Soon B. (Laboratory of RNA Viral Diseases, Korea Zoonosis Research Institute, Jeonbuk National University)
Publication Information
Molecules and Cells / v.45, no.10, 2022 , pp. 702-717 More about this Journal
Abstract
Hepatitis C virus (HCV) infection can lead to chronic hepatitis, liver cirrhosis, and hepatocellular carcinoma. HCV employs diverse strategies to evade host antiviral innate immune responses to mediate a persistent infection. In the present study, we show that nonstructural protein 5A (NS5A) interacts with an NF-κB inhibitor immunomodulatory kinase, IKKε, and subsequently downregulates beta interferon (IFN-β) promoter activity. We further demonstrate that NS5A inhibits DDX3-mediated IKKε and interferon regulatory factor 3 (IRF3) phosphorylation. We also note that hyperphosphorylation of NS5A mediates protein interplay between NS5A and IKKε, thereby contributing to NS5A mediated modulation of IFN-β signaling. Lastly, NS5A inhibits IKKε-dependent p65 phosphorylation and NF-κB activation. Based on these findings, we propose NS5A as a novel regulator of IFN signaling events, specifically by inhibiting IKKε downstream signaling cascades through its interaction with IKKε. Taken together, these data suggest an additional mechanistic means by which HCV modulates host antiviral innate immune responses to promote persistent viral infection.
Keywords
DDX3; hepatitis C virus; IFN-${\beta}$; $IKK{\varepsilon}$; IRF3; NS5A;
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1 Ng, S.L., Friedman, B.A., Schmid, S., Gertz, J., Myers, R.M., Tenoever, B.R., and Maniatis, T. (2011). IkappaB kinase epsilon (IKK(epsilon)) regulates the balance between type I and type II interferon responses. Proc. Natl. Acad. Sci. U. S. A. 108, 21170-21175.   DOI
2 Oshiumi, H., Sakai, K., Matsumoto, M., and Seya, T. (2010). DEAD/H BOX 3 (DDX3) helicase binds the RIG-I adaptor IPS-1 to up-regulate IFN-betainducing potential. Eur. J. Immunol. 40, 940-948.   DOI
3 Perwitasari, O., Cho, H., Diamond, M.S., and Gale, M., Jr. (2011). Inhibitor of kappaB kinase epsilon (IKK(epsilon)), STAT1, and IFIT2 proteins define novel innate immune effector pathway against West Nile virus infection. J. Biol. Chem. 286, 44412-44423.   DOI
4 Reed, K.E. and Rice, C.M. (2000). Overview of hepatitis C virus genome structure, polyprotein processing, and protein properties. Curr. Top. Microbiol. Immunol. 242, 55-84.
5 Adli, M. and Baldwin, A.S. (2006). IKK-i/IKKepsilon controls constitutive, cancer cell-associated NF-kappaB activity via regulation of Ser-536 p65/ RelA phosphorylation. J. Biol. Chem. 281, 26976-26984.   DOI
6 Schroder, M., Baran, M., and Bowie, A.G. (2008). Viral targeting of DEAD box protein 3 reveals its role in TBK1/IKKepsilon-mediated IRF activation. EMBO J. 27, 2147-2157.   DOI
7 Reed, K.E., Xu, J., and Rice, C.M. (1997). Phosphorylation of the hepatitis C virus NS5A protein in vitro and in vivo: properties of the NS5A-associated kinase. J. Virol. 71, 7187-7197.   DOI
8 Reyes, G.R. (2002). The nonstructural NS5A protein of hepatitis C virus: an expanding, multifunctional role in enhancing hepatitis C virus pathogenesis. J. Biomed. Sci. 9, 187-197.   DOI
9 Tanji, Y., Kaneko, T., Satoh, S., and Shimotohno, K. (1995). Phosphorylation of hepatitis C virus-encoded nonstructural protein NS5A. J. Virol. 69, 3980-3986.   DOI
10 Wong, A.H., Tam, N.W., Yang, Y.L., Cuddihy, A.R., Li, S., Kirchhoff, S., Hauser, H., Decker, T., and Koromilas, A.E. (1997). Physical association between STAT1 and the interferon-inducible protein kinase PKR and implications for interferon and double-stranded RNA signaling pathways. EMBO J. 16, 1291-1304.   DOI
11 Sharma, S., tenOever, B.R., Grandvaux, N., Zhou, G.P., Lin, R., and Hiscott, J. (2003). Triggering the interferon antiviral response through an IKK-related pathway. Science 300, 1148-1151.   DOI
12 Saito, T., Owen, D.M., Jiang, F., Marcotrigiano, J., and Gale, M., Jr. (2008). Innate immunity induced by composition-dependent RIG-I recognition of hepatitis C virus RNA. Nature 454, 523-527.   DOI
13 Kawai, T. and Akira, S. (2006). Innate immune recognition of viral infection. Nat. Immunol. 7, 131-137.   DOI
14 Kumthip, K., Chusri, P., Jilg, N., Zhao, L., Fusco, D.N., Zhao, H., Goto, K., Cheng, D., Schaefer, E.A., Zhang, L., et al. (2012). Hepatitis C virus NS5A disrupts STAT1 phosphorylation and suppresses type I interferon signaling. J. Virol. 86, 8581-8591.   DOI
15 Li, K., Foy, E., Ferreon, J.C., Nakamura, M., Ferreon, A.C., Ikeda, M., Ray, S.C., Gale, M., Jr., and Lemon, S.M. (2005a). Immune evasion by hepatitis C virus NS3/4A protease-mediated cleavage of the Toll-like receptor 3 adaptor protein TRIF. Proc. Natl. Acad. Sci. U. S. A. 102, 2992-2997.   DOI
16 Masaki, T., Matsunaga, S., Takahashi, H., Nakashima, K., Kimura, Y., Ito, M., Matsuda, M., Murayama, A., Kato, T., Hirano, H., et al. (2014). Involvement of hepatitis C virus NS5A hyperphosphorylation mediated by casein kinase I-alpha in infectious virus production. J. Virol. 88, 7541-7555.   DOI
17 Yu, M. and Levine, S.J. (2011). Toll-like receptor, RIG-I-like receptors and the NLRP3 inflammasome: key modulators of innate immune responses to double-stranded RNA viruses. Cytokine Growth Factor Rev. 22, 63-72.   DOI
18 Yu, S., Chen, J., Wu, M., Chen, H., Kato, N., and Yuan, Z. (2010). Hepatitis B virus polymerase inhibits RIG-I- and Toll-like receptor 3-mediated beta interferon induction in human hepatocytes through interference with interferon regulatory factor 3 activation and dampening of the interaction between TBK1/IKKepsilon and DDX3. J. Gen. Virol. 91, 2080-2090.   DOI
19 Li, X.D., Sun, L., Seth, R.B., Pineda, G., and Chen, Z.J. (2005b). Hepatitis C virus protease NS3/4A cleaves mitochondrial antiviral signaling protein off the mitochondria to evade innate immunity. Proc. Natl. Acad. Sci. U. S. A. 102, 17717-17722.   DOI
20 Lin, W., Kim, S.S., Yeung, E., Kamegaya, Y., Blackard, J.T., Kim, K.A., Holtzman, M.J., and Chung, R.T. (2006). Hepatitis C virus core protein blocks interferon signaling by interaction with the STAT1 SH2 domain. J. Virol. 80, 9226-9235.   DOI
21 Matsumoto, M., Hwang, S.B., Jeng, K.S., Zhu, N., and Lai, M.M. (1996). Homotypic interaction and multimerization of hepatitis C virus core protein. Virology 218, 43-51.   DOI
22 Morikawa, K., Lange, C.M., Gouttenoire, J., Meylan, E., Brass, V., Penin, F., and Moradpour, D. (2011). Nonstructural protein 3-4A: the Swiss army knife of hepatitis C virus. J. Viral Hepat. 18, 305-315.   DOI
23 Fitzgerald, K.A., McWhirter, S.M., Faia, K.L., Rowe, D.C., Latz, E., Golenbock, D.T., Coyle, A.J., Liao, S.M., and Maniatis, T. (2003). IKKepsilon and TBK1 are essential components of the IRF3 signaling pathway. Nat. Immunol. 4, 491-496.
24 Nakatsu, Y., Matsuoka, M., Chang, T.H., Otsuki, N., Noda, M., Kimura, H., Sakai, K., Kato, H., Takeda, M., and Kubota, T. (2014). Functionally distinct effects of the C-terminal regions of IKKepsilon and TBK1 on type I IFN production. PLoS One 9, e94999.   DOI
25 Nguyen, L.P., Nguyen, T.T.T., Nguyen, H.C., Pham, H.T., Han, K.M., Choi, D.H., Park, E.M., Kang, S.M., Tark, D., Lim, Y.S., et al. (2020). Cortactin interacts with hepatitis C virus core and NS5A proteins: implications for virion assembly. J. Virol. 94, e01306-20.
26 Noguchi, T., Satoh, S., Noshi, T., Hatada, E., Fukuda, R., Kawai, A., Ikeda, S., Hijikata, M., and Shimotohno, K. (2001). Effects of mutation in hepatitis C virus nonstructural protein 5A on interferon resistance mediated by inhibition of PKR kinase activity in mammalian cells. Microbiol. Immunol. 45, 829-840.   DOI
27 Park, K.J., Choi, S.H., Choi, D.H., Park, J.M., Yie, S.W., Lee, S.Y., and Hwang, S.B. (2003). Hepatitis C virus NS5A protein modulates c-Jun N-terminal kinase through interaction with tumor necrosis factor receptor-associated factor 2. J. Biol. Chem. 278, 30711-30718.   DOI
28 Lan, K.H., Lan, K.L., Lee, W.P., Sheu, M.L., Chen, M.Y., Lee, Y.L., Yen, S.H., Chang, F.Y., and Lee, S.D. (2007). HCV NS5A inhibits interferon-alpha signaling through suppression of STAT1 phosphorylation in hepatocytederived cell lines. J. Hepatol. 46, 759-767.
29 Asabe, S.I., Tanji, Y., Satoh, S., Kaneko, T., Kimura, K., and Shimotohno, K. (1997). The N-terminal region of hepatitis C virus-encoded NS5A is important for NS4A-dependent phosphorylation. J. Virol. 71, 790-796.   DOI
30 Choi, J.W., Kim, J.W., Nguyen, L.P., Nguyen, H.C., Park, E.M., Choi, D.H., Han, K.M., Kang, S.M., Tark, D., Lim, Y.S., et al. (2020). Nonstructural NS5A protein regulates LIM and SH3 domain protein 1 to promote hepatitis C virus propagation. Mol. Cells 43, 469-478.
31 Gu, L., Fullam, A., Brennan, R., and Schroder, M. (2013). Human DEAD box helicase 3 couples IkappaB kinase epsilon to interferon regulatory factor 3 activation. Mol. Cell. Biol. 33, 2004-2015.   DOI
32 Lim, Y.S., Nguyen, M.T.N., Pham, T.X., Huynh, T.T.X., Park, E.M., Choi, D.H., Kang, S.M., Tark, D., and Hwang, S.B. (2022). Hepatitis C virus NS5A protein interacts with telomere length regulation protein: implications for telomere shortening in patients infected with HCV. Mol. Cells 45, 148-157.   DOI
33 Lau, D.T., Fish, P.M., Sinha, M., Owen, D.M., Lemon, S.M., and Gale, M., Jr. (2008). Interferon regulatory factor-3 activation, hepatic interferonstimulated gene expression, and immune cell infiltration in hepatitis C virus patients. Hepatology 47, 799-809.   DOI
34 Li, Q., Brass, A.L., Ng, A., Hu, Z., Xavier, R.J., Liang, T.J., and Elledge, S.J. (2009). A genome-wide genetic screen for host factors required for hepatitis C virus propagation. Proc. Natl. Acad. Sci. U. S. A. 106, 16410-16415.   DOI
35 Li, S., Wang, L., Berman, M., Kong, Y.Y., and Dorf, M.E. (2011). Mapping a dynamic innate immunity protein interaction network regulating type I interferon production. Immunity 35, 426-440.   DOI
36 Lin, R., Heylbroeck, C., Pitha, P.M., and Hiscott, J. (1998). Virus-dependent phosphorylation of the IRF-3 transcription factor regulates nuclear translocation, transactivation potential, and proteasome-mediated degradation. Mol. Cell. Biol. 18, 2986-2996.   DOI
37 Lindenbach, B.D. and Rice, C.M. (2005). Unravelling hepatitis C virus replication from genome to function. Nature 436, 933-938.   DOI
38 Metz, P., Reuter, A., Bender, S., and Bartenschlager, R. (2013). Interferonstimulated genes and their role in controlling hepatitis C virus. J. Hepatol. 59, 1331-1341.   DOI
39 Tenoever, B.R., Ng, S.L., Chua, M.A., McWhirter, S.M., Garcia-Sastre, A., and Maniatis, T. (2007). Multiple functions of the IKK-related kinase IKKepsilon in interferon-mediated antiviral immunity. Science 315, 1274-1278.   DOI
40 Jensen, S. and Thomsen, A.R. (2012). Sensing of RNA viruses: a review of innate immune receptors involved in recognizing RNA virus invasion. J. Virol. 86, 2900-2910.   DOI
41 Pavio, N., Taylor, D.R., and Lai, M.M. (2002). Detection of a novel unglycosylated form of hepatitis C virus E2 envelope protein that is located in the cytosol and interacts with PKR. J. Virol. 76, 1265-1272.   DOI
42 Polyak, S.J., Khabar, K.S., Paschal, D.M., Ezelle, H.J., Duverlie, G., Barber, G.N., Levy, D.E., Mukaida, N., and Gretch, D.R. (2001). Hepatitis C virus nonstructural 5A protein induces interleukin-8, leading to partial inhibition of the interferon-induced antiviral response. J. Virol. 75, 6095-6106.   DOI
43 Ray, R.B. and Ray, R. (2001). Hepatitis C virus core protein: intriguing properties and functional relevance. FEMS Microbiol. Lett. 202, 149-156.   DOI
44 Brownell, J., Bruckner, J., Wagoner, J., Thomas, E., Loo, Y.M., Gale, M., Jr., Liang, T.J., and Polyak, S.J. (2014). Direct, interferon-independent activation of the CXCL10 promoter by NF-kappaB and interferon regulatory factor 3 during hepatitis C virus infection. J. Virol. 88, 1582-1590.   DOI
45 Saito, I., Miyamura, T., Ohbayashi, A., Harada, H., Katayama, T., Kikuchi, S., Watanabe, Y., Koi, S., Onji, M., and Ohta, Y. (1990). Hepatitis C virus infection is associated with the development of hepatocellular carcinoma. Proc. Natl. Acad. Sci. U. S. A. 87, 6547-6549.   DOI
46 Seth, R.B., Sun, L., Ea, C.K., and Chen, Z.J. (2005). Identification and characterization of MAVS, a mitochondrial antiviral signaling protein that activates NF-kappaB and IRF 3. Cell 122, 669-682.   DOI
47 Appel, N., Pietschmann, T., and Bartenschlager, R. (2005). Mutational analysis of hepatitis C virus nonstructural protein 5A: potential role of differential phosphorylation in RNA replication and identification of a genetically flexible domain. J. Virol. 79, 3187-3194.   DOI
48 Appel, N., Zayas, M., Miller, S., Krijnse-Locker, J., Schaller, T., Friebe, P., Kallis, S., Engel, U., and Bartenschlager, R. (2008). Essential role of domain III of nonstructural protein 5A for hepatitis C virus infectious particle assembly. PLoS Pathog. 4, e1000035.   DOI
49 Balka, K.R., Louis, C., Saunders, T.L., Smith, A.M., Calleja, D.J., D'Silva, D.B., Moghaddas, F., Tailler, M., Lawlor, K.E., Zhan, Y., et al. (2020). TBK1 and IKKε act redundantly to mediate STING-induced NF-κB responses in myeloid cells. Cell Rep. 31, 107492.   DOI
50 Choi, S.H. and Hwang, S.B. (2006). Modulation of TGF-beta signal transduction pathway by hepatitis C virus nonstructural 5A protein. J. Biol. Chem. 281, 7468-7478.   DOI
51 Ferreon, J.C., Ferreon, A.C., Li, K., and Lemon, S.M. (2005). Molecular determinants of TRIF proteolysis mediated by the hepatitis C virus NS3/4A protease. J. Biol. Chem. 280, 20483-20492.   DOI
52 Indukuri, H., Castro, S.M., Liao, S.M., Feeney, L.A., Dorsch, M., Coyle, A.J., Garofalo, R.P., Brasier, A.R., and Casola, A. (2006). Ikkepsilon regulates viral-induced interferon regulatory factor-3 activation via a redox-sensitive pathway. Virology 353, 155-165.   DOI
53 Gale, M., Jr., Blakely, C.M., Kwieciszewski, B., Tan, S.L., Dossett, M., Tang, N.M., Korth, M.J., Polyak, S.J., Gretch, D.R., and Katze, M.G. (1998). Control of PKR protein kinase by hepatitis C virus nonstructural 5A protein: molecular mechanisms of kinase regulation. Mol. Cell. Biol. 18, 5208-5218.   DOI
54 Gale, M.J., Jr., Korth, M.J., Tang, N.M., Tan, S.L., Hopkins, D.A., Dever, T.E., Polyak, S.J., Gretch, D.R., and Katze, M.G. (1997). Evidence that hepatitis C virus resistance to interferon is mediated through repression of the PKR protein kinase by the nonstructural 5A protein. Virology 230, 217-227.   DOI
55 Hoofnagle, J.H. (2002). Course and outcome of hepatitis C. Hepatology 36(5 Suppl 1), S21-S29.
56 Kanda, T., Steele, R., Ray, R., and Ray, R.B. (2007). Hepatitis C virus infection induces the beta interferon signaling pathway in immortalized human hepatocytes. J. Virol. 81, 12375-12381.   DOI
57 Kaneko, T., Tanji, Y., Satoh, S., Hijikata, M., Asabe, S., Kimura, K., and Shimotohno, K. (1994). Production of two phosphoproteins from the NS5A region of the hepatitis C viral genome. Biochem. Biophys. Res. Commun. 205, 320-326.   DOI
58 Kawai, T. and Akira, S. (2008). Toll-like receptor and RIG-I-like receptor signaling. Ann. N. Y. Acad. Sci. 1143, 1-20.   DOI
59 Kawai, T., Takahashi, K., Sato, S., Coban, C., Kumar, H., Kato, H., Ishii, K.J., Takeuchi, O., and Akira, S. (2005). IPS-1, an adaptor triggering RIG-I- and Mda5-mediated type I interferon induction. Nat. Immunol. 6, 981-988.   DOI