Reticulon 3 interacts with NS4B of the hepatitis C virus and negatively regulates viral replication by disrupting NS4B self-interaction

Ming-Jhan Wu, Po Yuan Ke, John T A Hsu, Chau Ting Yeh, Jim Tong Horng

Research output: Contribution to journalArticle

14 Citations (Scopus)

Abstract

Summary: The non-structural protein 4B (NS4B) of the hepatitis C virus (HCV) is an endoplasmic reticulum (ER) membrane protein comprising two consecutive amphipathic α-helical domains (AH1 and AH2). Its self-oligomerization via the AH2 domain is required for the formation of the membranous web that is necessary for viral replication. Previously, we reported that the host-encoded ER-associated reticulon 3 (RTN3) protein is involved in the formation of the replication-associated membranes of (+)RNA enteroviruses during viral replication. In this study, we demonstrated that the second transmembrane region of RTN3 competed for, and bound to, the AH2 domain of NS4B, thus abolishing NS4B self-interaction and leading to the downregulation of viral replication. This interaction was mediated by two crucial residues, lysine 52 and tyrosine 63, of AH2, and was regulated by the AH1 domain. The silencing of RTN3 in Huh7 and AVA5 cells harbouring an HCV replicon enhanced the replication of HCV, which was counteracted by the overexpression of recombinant RTN3. The synthesis of viral RNA was also increased in siRNA-transfected human primary hepatocytes infected with HCV derived from cell culture. Our results demonstrated that RTN3 acted as a restriction factor to limit the replication of HCV. The NS4B of the HCV is an ER membrane protein comprising two amphipathic α-helical domains (AH1 and AH2). Its self-oligomerization via the AH2 domain is required for the formation of the membranous web that is necessary for viral replication. In this study, we demonstrate that the host-encoded reticulon 3 (RTN3) competes for, and binds to, the AH2 domain of NS4B, thus abolishing NS4B self-interaction and leading to the downregulation of viral replication. This interaction was mediated by two crucial residues, lysine 52 and tyrosine 63, of AH2, and was regulated by the AH1 domain. The silencing of RTN3 in Huh7 and AVA5 cells harbouring an HCV replicon enhanced the replication of HCV, which was counteracted by the overexpression of recombinant RTN3. The synthesis of viral RNA was also increased in siRNA-transfected human primary hepatocytes infected with HCV derived from cell culture. Our results demonstrated that RTN3 acted as a restriction factor to limit the replication of HCV.

Original languageEnglish (US)
Pages (from-to)1603-1618
Number of pages16
JournalCellular Microbiology
Volume16
Issue number11
DOIs
StatePublished - Nov 1 2014
Externally publishedYes

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Hepacivirus
Proteins
Endoplasmic Reticulum
Replicon
Viral RNA
Small Interfering RNA
Lysine
Tyrosine
Hepatocytes
Membrane Proteins
Down-Regulation
Cell Culture Techniques
Enterovirus
RNA
Membranes

ASJC Scopus subject areas

  • Microbiology
  • Immunology
  • Virology
  • Medicine(all)

Cite this

Reticulon 3 interacts with NS4B of the hepatitis C virus and negatively regulates viral replication by disrupting NS4B self-interaction. / Wu, Ming-Jhan; Ke, Po Yuan; Hsu, John T A; Yeh, Chau Ting; Horng, Jim Tong.

In: Cellular Microbiology, Vol. 16, No. 11, 01.11.2014, p. 1603-1618.

Research output: Contribution to journalArticle

Wu, Ming-Jhan ; Ke, Po Yuan ; Hsu, John T A ; Yeh, Chau Ting ; Horng, Jim Tong. / Reticulon 3 interacts with NS4B of the hepatitis C virus and negatively regulates viral replication by disrupting NS4B self-interaction. In: Cellular Microbiology. 2014 ; Vol. 16, No. 11. pp. 1603-1618.
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