Hepatitis C virus (HCV) is a major cause of chronic liver disease and is highly dependent on cellular proteins for virus propagation. To identify the cellular factors involved in HCV propagation, we recently performed protein microarray assays using the HCV nonstructural 5A (NS5A) protein as a probe. Of 90 cellular protein candidates, we selected the soluble resistance-related calcium-binding protein (sorcin) for further characterization. Sorcin is a calcium-binding protein and is highly expressed in certain cancer cells. We verified that NS5A interacted with sorcin through domain I of NS5A, and phosphorylation of the threonine residue 155 of sorcin played a crucial role in protein interaction. Small interfering RNA (siRNA)-mediated knockdown of sorcin impaired HCV propagation. Silencing of sorcin expression resulted in a decrease of HCV assembly without affecting HCV RNA and protein levels. We further demonstrated that polo-like kinase 1 (PLK1)-mediated phosphorylation of sorcin was increased by NS5A. We showed that both phosphorylation and calcium-binding activity of sorcin were required for HCV propagation. These data indicate that HCV modulates sorcin activity via NS5A protein for its own propagation.
IMPORTANCESorcin is a calcium-binding protein and regulates intracellular calcium homeostasis. HCV NS5A interacts with sorcin, and phosphorylation of sorcin is required for protein interaction. Gene silencing of sorcin impaired HCV propagation at the assembly step of the HCV life cycle. Sorcin is phosphorylated by PLK1 via protein interaction. We showed that sorcin interacted with both NS5A and PLK1, and PLK1-mediated phosphorylation of sorcin was increased by NS5A. Moreover, calcium-binding activity of sorcin played a crucial role in HCV propagation. These data provide evidence that HCV regulates host calcium metabolism for virus propagation, and thus manipulation of sorcin activity may represent a novel therapeutic target for HCV.
Hepatitis C virus (HCV) is a major causative agent of non-A, non-B hepatitis. HCV infection often leads to chronic hepatitis, liver cirrhosis, and ultimately hepatocellular carcinoma (HCC) (1). HCV belongs to the member of the Hepacivirus genus within the Flaviviridae family. HCV is an enveloped virus with a positive-sense, single-stranded RNA genome of ϳ9.6 kb. Its genome encodes a single polyprotein precursor of more than 3,000 amino acids, which is cleaved by both host and viral proteases at the endoplasmic reticulum (ER), yielding structural (core, E1, and E2) and nonstructural (p7 and NS2 to NS5B) proteins (2). The nonstructural 5A (NS5A) protein is a phosphoprotein consisting of 447 amino acid residues. NS5A exists in two forms of polypeptide, p56 and p58, which are phosphorylated at serine residues by cellular kinase (3), and phosphorylation regulates the HCV life cycle (4). NS5A protein forms a part of the HCV RNA replication complex (5) and is involved in liver pathogenesis (6). NS5A is a multifunctional protein that regulates RNA replication, interferon (IFN) resis...