Identification of three interferon-inducible cellular enzymes that inhibit the replication of hepatitis C virus.
Hepatitis C virus (HCV) infection is a common cause of chronic hepatitis and is currently treated with alpha interferon (IFN-alpha)-based therapies. However, the underlying mechanism of IFN-alpha therapy remains to be elucidated. To identify the cellular proteins that mediate the antiviral effects of IFN-alpha, we created a HEK293-based cell culture system to inducibly express individual interferon-stimulated genes (ISGs) and determined their antiviral effects against HCV. By screening 29 ISGs that are induced in Huh7 cells by IFN-alpha and/or up-regulated in HCV-infected livers, we discovered that viperin, ISG20, and double-stranded RNA-dependent protein kinase (PKR) noncytolytically inhibited the replication of HCV replicons. Mechanistically, inhibition of HCV replication by ISG20 and PKR depends on their 3'-5' exonuclease and protein kinase activities, respectively. Moreover, our work, for the first time, provides strong evidence suggesting that viperin is a putative radical S-adenosyl-l-methionine (SAM) enzyme. In addition to demonstrating that the antiviral activity of viperin depends on its radical SAM domain, which contains conserved motifs to coordinate [4Fe-4S] cluster and cofactor SAM and is essential for its enzymatic activity, mutagenesis studies also revealed that viperin requires an aromatic amino acid residue at its C terminus for proper antiviral function. Furthermore, although the N-terminal 70 amino acid residues of viperin are not absolutely required, deletion of this region significantly compromises its antiviral activity against HCV. Our findings suggest that viperin represents a novel antiviral pathway that works together with other antiviral proteins, such as ISG20 and PKR, to mediate the IFN response against HCV infection.
Jiang
Dong
D
Drexel Institute for Biotechnology and Virology Research, Department of Microbiology and Immunology, Drexel University College of Medicine, 3805 Old Easton Road, Doylestown, PA 18902, USA.
Guo
Haitao
H
Xu
Chunxiao
C
Chang
Jinhong
J
Gu
Baohua
B
Wang
Lijuan
L
Block
Timothy M
TM
Guo
Ju-Tao
JT
D000595:Amino Acid Sequence; D000998:Antiviral Agents / Q000494:pharmacology*; D002460:Cell Line; D005092:Exonucleases / Q000235:genetics / Q000378:metabolism*; D005095:Exoribonucleases; D015870:Gene Expression / Q000187:drug effects; D016679:Genome, Viral; D016174:Hepacivirus / Q000187:drug effects* / Q000502:physiology; D006801:Humans; D016898:Interferon-alpha / Q000494:pharmacology*; D008969:Molecular Sequence Data; D044925:Oxidoreductases Acting on CH-CH Group Donors; D011506:Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D012367:RNA, Viral / Q000235:genetics; D012093:Replicon / Q000187:drug effects / Q000502:physiology; D012436:S-Adenosylmethionine / Q000378:metabolism; D014162:Transfection; D014779:Virus Replication / Q000187:drug effects*; D019892:eIF-2 Kinase / Q000235:genetics / Q000378:metabolism*; D000090123:Viperin Protein
D000998:Antiviral Agents; D005092:Exonucleases; D005095:Exoribonucleases; D016898:Interferon-alpha; D044925:Oxidoreductases Acting on CH-CH Group Donors; D011506:Proteins; D012367:RNA, Viral; D012436:S-Adenosylmethionine; D019892:eIF-2 Kinase; C444427:RSAD2 protein, human; D000090123:Viperin Protein; C112346:ISG20 protein, human