The Role of Signal Peptidase Complex Subunit 1 (SPCS1) in infectious Hepatitis C virus (HCV) Assembly

Abstract

Hepatitis C virus (HCV) is a single stranded RNA virus belonging to the hepacivirus genus in the Flaviviridae family. It causes severe liver diseases, including chronic hepatitis, cirrhosis, and hepatocellular carcinoma, leading to global health burden for 70 million chronically infected people. Despite availability of highly effective therapy, new HCV infections continue to increase in the US and worldwide due to barriers including lack of vaccine. Recent studies identified signal peptidase complex subunit I (SPCS1) as a proviral host factor specific for Flaviviridae viruses, including HCV. SPCS1 is a regulatory subunit of signal peptidase complex (SPC). SPCS1 was shown to regulate flavivirus C-prM processing, but in HCV, its role was attributed to the enhancement of NS2 and E2 interaction unrelated to SPC processing. Since our previous study determined that inhibiting HCV E2-p7 processing impairs NS2 and E2 interaction, we aim to assess whether SPCS1’s fundamental role in HCV propagation is to regulate E2-p7 processing. HCV E2-p7 processing is mediated by the SPC but suboptimal. We previously showed that either further facilitating this processing or inhibiting it impaired HCV assembly indicating that processing at this site requires exquisite regulation. However, the regulatory mechanism of E2-p7 processing remains incompletely defined. In this study, we determined that loss of SPCS1 impairs HCV E2-p7 processing. We also determined that making E2 and p7 separation independent of SPC-mediated cleavage or augmenting the SPC-mediated cleavage of E2-p7 junction by inserting HA tag to p7 N-terminus canceled SPCS1’s role in HCV assembly. Structural modeling suggested that HA tag relieves the structural constraint imposed on E2-p7 junction site by the p7 sequence. Based on these results we conclude that the mechanism of action of SPCS1 in HCV assembly is to promote E2-p7 processing by enhancing E2-p7 junction site presentation to the signal peptidase active site. By providing evidence that SPCS1 facilitates HCV assembly by regulating SPC-mediated cleavage of E2-p7 junction, similar to previously established role of this protein in C-prM junction processing in flavivirus, this dissertation establishes a common role for SPCS1 in Flaviviridae family virus propagation as to exquisitely regulate SPC-mediated processing of specific, suboptimal target sites.