title: Control of innate immune signaling and membrane targeting by the Hepatitis C virus NS3/4A protease are governed by the NS3 helix α0.
abstract: Hepatitis C virus (HCV) infection is sensed in the host cell by the cytosolic pathogen recognition receptor RIG-I. RIG-I signaling is propagated through its signaling adaptor protein MAVS to drive activation of innate immunity. However, HCV blocks RIG-I signaling through viral NS3/4A protease cleavage of MAVS on the mitochondrion-associated endoplasmic reticulum (ER) membrane (MAM). The multifunctional HCV NS3/4A serine protease is associated with intracellular membranes, including the MAM, through membrane-targeting domains within NS4A and also at the amphipathic helix α(0) of NS3. The serine protease domain of NS3 is required for both cleavage of MAVS, a tail-anchored membrane protein, and processing the HCV polyprotein. Here, we show that hydrophobic amino acids in the NS3 helix α(0) are required for selective cleavage of membrane-anchored portions of the HCV polyprotein and for cleavage of MAVS for control of RIG-I pathway signaling of innate immunity. Further, we found that the hydrophobic composition of NS3 helix α(0) is essential to establish HCV replication and infection. Alanine substitution of individual hydrophobic amino acids in the NS3 helix α(0) impaired HCV RNA replication in cells with a functional RIG-I pathway, but viral RNA replication was rescued in cells lacking RIG-I signaling. Therefore, the hydrophobic amphipathic helix α(0) of NS3 is required for NS3/4A control of RIG-I signaling and HCV replication by directing the membrane targeting of both viral and cellular substrates.
authors.affiliation: Department of Immunology, University of Washington, Seattle, Washington, USA.
authors.forename: Stacy M
authors.initials: SM
authors.lastname: Horner
authors.forename: Hae Soo
authors.initials: HS
authors.lastname: Park
authors.forename: Michael
authors.initials: M
authors.lastname: Gale
mesh_terms: D048868:Adaptor Proteins, Signal Transducing / Q000276:immunology / Q000378:metabolism; D000595:Amino Acid Sequence; D002352:Carrier Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D002460:Cell Line; D000071457:DEAD Box Protein 58; D053487:DEAD-box RNA Helicases / Q000276:immunology; D004721:Endoplasmic Reticulum / Q000378:metabolism; D016174:Hepacivirus / Q000737:chemistry / Q000201:enzymology* / Q000235:genetics; D006526:Hepatitis C / Q000276:immunology / Q000378:metabolism* / Q000821:virology; D006801:Humans; D007113:Immunity, Innate; D007425:Intracellular Membranes / Q000378:metabolism / Q000821:virology; D047908:Intracellular Signaling Peptides and Proteins; D008928:Mitochondria / Q000378:metabolism; D008969:Molecular Sequence Data; D011485:Protein Binding; D017433:Protein Structure, Secondary; D011971:Receptors, Immunologic; D015398:Signal Transduction; D017361:Viral Nonstructural Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D012697:Serine Endopeptidases; D043583:Nucleoside-Triphosphatase; D000085242:Viral Proteases
chemical_list: D048868:Adaptor Proteins, Signal Transducing; D002352:Carrier Proteins; D000071457:DEAD Box Protein 58; D053487:DEAD-box RNA Helicases; D047908:Intracellular Signaling Peptides and Proteins; D011971:Receptors, Immunologic; D017361:Viral Nonstructural Proteins; C505371:MAVS protein, human; C084422:NS3 protein, hepatitis C virus; D012697:Serine Endopeptidases; D043583:Nucleoside-Triphosphatase; D000085242:Viral Proteases; C455041:NS4A cofactor peptide, Hepatitis C virus; C452820:RIGI protein, human