Molecules and Cells

Korean Society for Molecular and Cellular Biology (한국분자세포생물학회)
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Hepatitis C Virus Nonstructural 5A Protein Interacts with Telomere Length Regulation Protein: Implications for Telomere Shortening in Patients Infected with HCV

  • Lim, Yun-Sook (Laboratory of RNA Viral Diseases, Korea Zoonosis Research Institute, Jeonbuk National University) ;
  • Nguyen, Men T.N. (Ilsong Institute of Life Science, Hallym University) ;
  • Pham, Thuy X. (Laboratory of RNA Viral Diseases, Korea Zoonosis Research Institute, Jeonbuk National University) ;
  • Huynh, Trang T.X. (Laboratory of RNA Viral Diseases, Korea Zoonosis Research Institute, Jeonbuk National University) ;
  • Park, Eun-Mee (Center for Immunology and Pathology, National Institute of Health, Korea Center for Disease Control & Prevention) ;
  • Choi, Dong Hwa (Biocenter, Gyeonggido Business & Science Accelerator) ;
  • Kang, Sang 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) ;
  • Hwang, Soon B. (Laboratory of RNA Viral Diseases, Korea Zoonosis Research Institute, Jeonbuk National University)
  • Received : 2021.06.23
  • Accepted : 2021.10.27
  • Published : 2022.03.31
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Abstract

Hepatitis C virus (HCV) is a major cause of chronic liver disease and is highly dependent on cellular proteins for viral propagation. Using protein microarray analysis, we identified 90 cellular proteins as HCV nonstructural 5A (NS5A) interacting partners, and selected telomere length regulation protein (TEN1) for further study. TEN1 forms a heterotrimeric complex with CTC and STN1, which is essential for telomere protection and maintenance. Telomere length decreases in patients with active HCV, chronic liver disease, and hepatocellular carcinoma. However, the molecular mechanism of telomere length shortening in HCV-associated disease is largely unknown. In the present study, protein interactions between NS5A and TEN1 were confirmed by immunoprecipitation assays. Silencing of TEN1 reduced both viral RNA and protein expression levels of HCV, while ectopic expression of the siRNA-resistant TEN1 recovered the viral protein level, suggesting that TEN1 was specifically required for HCV propagation. Importantly, we found that TEN1 is re-localized from the nucleus to the cytoplasm in HCV-infected cells. These data suggest that HCV exploits TEN1 to promote viral propagation and that telomere protection is compromised in HCV-infected cells. Overall, our findings provide mechanistic insight into the telomere shortening in HCV-infected cells.

Keywords

Acknowledgement

We thank Dr. Fuyuki Ishikawa (Kyoto University) for providing rabbit anti-TEN1 antibody. This work was supported by the National Research Foundation of Korea (NRF) grant funded by the Korea government (MSIT) (2021R1A2C2003275 for S.B.H.). This work was also supported by the National Research Foundation of Korea (NRF) grant funded by the Korea government (MSIT) (2019R1A2C1086914 for Y.S.L.).

References

  1. 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. https://doi.org/10.1371/journal.ppat.1000035
  2. Biron-Shental, T., Amiel, A., Anchidin, R., Sharony, R., Hadary, R., and Kitay-Cohen, Y. (2013). Telomere length and telomerase reverse transcriptase mRNA expression in patients with hepatitis C. Hepatogastroenterology 60, 1713-1716.
  3. Casteel, D.E., Zhuang, S., Zeng, Y., Perrino, F.W., Boss, G.R., Goulian, M., and Pilz, R.B. (2009). A DNA polymerase-α.primase cofactor with homology to replication protein A-32 regulates DNA replication in mammalian cells. J. Biol. Chem. 284, 5807-5818. https://doi.org/10.1074/jbc.M807593200
  4. Cawthon, R.M. (2009). Telomere length measurement by a novel monochrome multiplex quantitative PCR method. Nucleic Acids Res. 37, e21. https://doi.org/10.1093/nar/gkn1027
  5. Chen, L.Y., Redon, S., and Lingner, J. (2012). The human CST complex is a terminator of telomerase activity. Nature 488, 540-544. https://doi.org/10.1038/nature11269
  6. 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. https://doi.org/10.14348/molcells.2020.0018
  7. Feng, X., Hsu, S.J., Kasbek, C., Chaiken, M., and Price, C.M. (2017). CTC1-mediated C-strand fill-in is an essential step in telomere length maintenance. Nucleic Acids Res. 45, 4281-4293. https://doi.org/10.1093/nar/gkx125
  8. Giannini, C. and Brechot, C. (2003). Hepatitis C virus biology. Cell Death Differ. 10 Suppl 1, S27-S38. https://doi.org/10.1038/sj.cdd.4401121
  9. Gu, P., Min, J.N., Wang, Y., Huang, C., Peng, T., Chai, W., and Chang, S. (2012). CTC1 deletion results in defective telomere replication, leading to catastrophic telomere loss and stem cell exhaustion. EMBO J. 31, 2309-2321. https://doi.org/10.1038/emboj.2012.96
  10. Huang, C., Dai, X., and Chai, W. (2012). Human Stn1 protects telomere integrity by promoting efficient lagging-strand synthesis at telomeres and mediating C-strand fill-in. Cell Res. 22, 1681-1695. https://doi.org/10.1038/cr.2012.132
  11. Kasbek, C., Wang, F., and Price, C.M. (2013). Human TEN1 maintains telomere integrity and functions in genome-wide replication restart. J. Biol. Chem. 288, 30139-30150. https://doi.org/10.1074/jbc.M113.493478
  12. Kelich, J.M., Papaioannou, H., and Skordalakes, E. (2021). Pol α-primase dependent nuclear localization of the mammalian CST complex. Commun. Biol. 4, 349. https://doi.org/10.1038/s42003-021-01845-4
  13. Kim, S., Welsch, C., Yi, M., and Lemon, S.M. (2011). Regulation of the production of infectious genotype 1a hepatitis C virus by NS5A domain III. J. Virol. 85, 6645-6656. https://doi.org/10.1128/JVI.02156-10
  14. Kitada, T., Seki, S., Kawakita, N., Kuroki, T., and Monna, T. (1995). Telomere shortening in chronic liver diseases. Biochem. Biophys. Res. Commun. 211, 33-39. https://doi.org/10.1006/bbrc.1995.1774
  15. Kitay-Cohen, Y., Goldberg-Bittman, L., Hadary, R., Fejgin, M.D., and Amiel, A. (2008). Telomere length in Hepatitis C. Cancer Genet. Cytogenet. 187, 34-38. https://doi.org/10.1016/j.cancergencyto.2008.08.006
  16. Lim, C.J. and Cech, T.R. (2021). Shaping human telomeres: from shelterin and CST complexes to telomeric chromatin organization. Nat. Rev. Mol. Cell Biol. 22, 283-298. https://doi.org/10.1038/s41580-021-00328-y
  17. Lim, Y.S. and Hwang, S.B. (2011). Hepatitis C virus NS5A protein interacts with phosphatidylinositol 4-kinase type IIIα and regulates viral propagation. J. Biol. Chem. 286, 11290-11298. https://doi.org/10.1074/jbc.M110.194472
  18. Lim, Y.S., Tran, H.T., Park, S.J., Yim, S.A., and Hwang, S.B. (2011). Peptidyl-prolyl isomerase Pin1 is a cellular factor required for hepatitis C virus propagation. J. Virol. 85, 8777-8788. https://doi.org/10.1128/JVI.02533-10
  19. Lindenbach, B.D. and Rice, C.M. (2005). Unravelling hepatitis C virus replication from genome to function. Nature 436, 933-938. https://doi.org/10.1038/nature04077
  20. Macdonald, A. and Harris, M. (2004). Hepatitis C virus NS5A: tales of a promiscuous protein. J. Gen. Virol. 85, 2485-2502. https://doi.org/10.1099/vir.0.80204-0
  21. McGivern, D.R. and Lemon, S.M. (2011). Virus-specific mechanisms of carcinogenesis in hepatitis C virus associated liver cancer. Oncogene 30, 1969-1983. https://doi.org/10.1038/onc.2010.594
  22. Miyake, Y., Nakamura, M., Navetani, A., Shimamura, S., Tamura, M., Yonehara, S., Saito, M., and Ishikawa, F. (2009). RPA-like mammalian Ctc1-Stn1-Ten1 complex binds to single-stranded DNA and protects telomeres independently of the Pot1 pathway. Mol. Cell 36, 193-206. https://doi.org/10.1016/j.molcel.2009.08.009
  23. Nakaoka, H., Nishiyama, A., Saito, M., and Ishikawa, F. (2012). Xenopus laevis Ctc1-Stn1-Ten1 (xCST) protein complex is involved in priming DNA synthesis on single-stranded DNA template in Xenopus egg extract. J. Biol. Chem. 287, 619-627. https://doi.org/10.1074/jbc.M111.263723
  24. Park, C., Min, S., Park, E.M., Lim, Y.S., Kang, S.M., Suzuki, T., Shin, E.C., and Hwang, S.B. (2015). Pim kinase interacts with nonstructural 5A protein and regulates hepatitis C virus entry. J. Virol. 89, 10073-10086. https://doi.org/10.1128/JVI.01707-15
  25. Saito, I., Miyamura, T., Ohbayashi, A., Harada, H., Katayama, T., Kikuchi, S., Watanabe, Y., Koi, S., Onji, M., Ohta, Y., et al. (1990). Hepatitis C virus infection is associated with the development of hepatocellular carcinoma. Proc. Natl. Acad. Sci. U. S. A. 87, 6547-6549. https://doi.org/10.1073/pnas.87.17.6547
  26. Simmonds, P. (2004). Genetic diversity and evolution of hepatitis C virus--15 years on. J. Gen. Virol. 85, 3173-3188. https://doi.org/10.1099/vir.0.80401-0
  27. Song, X., Leehy, K., Warrington, R.T., Lamb, J.C., Surovtseva, Y.V., and Shippen, D.E. (2008). STN1 protects chromosome ends in Arabidopsis thaliana. Proc. Natl. Acad. Sci. U. S. A. 105, 19815-19820. https://doi.org/10.1073/pnas.0807867105
  28. Srinivas, N., Rachakonda, S., and Kumar, R. (2020). Telomeres and telomere length: a general overview. Cancers (Basel) 12, 558. https://doi.org/10.3390/cancers12030558
  29. Surovtseva, Y.V., Churikov, D., Boltz, K.A., Song, X., Lamb, J.C., Warrington, R., Leehy, K., Heacock, M., Price, C.M., and Shippen, D.E. (2009). Conserved telomere maintenance component 1 interacts with STN1 and maintains chromosome ends in higher eukaryotes. Mol. Cell 36, 207-218. https://doi.org/10.1016/j.molcel.2009.09.017
  30. Tellinghuisen, T.L., Foss, K.L., and Treadaway, J. (2008). Regulation of hepatitis C virion production via phosphorylation of the NS5A protein. PLoS Pathog. 4, e1000032. https://doi.org/10.1371/journal.ppat.1000032
  31. Tran, G.V.Q., Luong, T.T.D., Park, E.M., Kim, J.W., Choi, J.W., Park, C., Lim, Y.S., and Hwang, S.B. (2016). Nonstructural 5A protein of hepatitis C virus regulates soluble resistance-related calcium-binding protein activity for viral propagation. J. Virol. 90, 2794-2805. https://doi.org/10.1128/JVI.02493-15
  32. Turner, K.J., Vasu, V., and Griffin, D.K. (2019). Telomere biology and human phenotype. Cells 8, 73. https://doi.org/10.3390/cells8010073
  33. Wiemann, S.U., Satyanarayana, A., Tsahuridu, M., Tillmann, H.L., Zender, L., Klempnauer, J., Flemming, P., Franco, S., Blasco, M.A., Manns, M.P., et al. (2002). Hepatocyte telomere shortening and senescence are general markers of human liver cirrhosis. FASEB J. 16, 935-942. https://doi.org/10.1096/fj.01-0977com