Proteomic profiling of cellular proteins interacting with the hepatitis C virus core protein

Abstract

Hepatitis C virus (HCV) is a causative agent of chronic hepatitis and hepatocellular carcinoma. The core protein of HCV packages the viral RNA genome to form a nucleocapsid. In addition to its function as a structural protein, core protein is involved in regulation of cellular transcription, virus-induced transformation, and pathogenesis. To gain insights into cellular functions of the core protein by identification of cellular proteins interacting with the core protein, we employed a proteomic approach. Hepatocytes soluble cytoplasmic proteins were applied to the core proteins immobilized on Ni-nitrilotriacetic resin and total bound cellular proteins were resolved by 2-DE. Analyses of interacting proteins by matrix-assisted laser desorption/ionization-time of flight mass spectrometry allowed identification of 14 cellular proteins binding to the core protein. These proteins include DEAD-box polypeptide 5, similar in function to a known protein identified previously by yeast two-hybrid screening and 13 newly identified cellular proteins. Interestingly, nine protein spots were identified as intermediate microfilament proteins, including cytokeratins (five spots for cytokeratin 8, two for cytokeratin 19, and one for cytokeratin 18) and vimentin. Cytokeratin 8 and vimentin, which were previously shown to be involved in the infection processes of other viruses, were further analyzed to confirm their in vivo interactions with the core protein by immunoblotting and immunofluorescence microscopy. We discuss the functional implications of the interactions of the core protein with newly identified cellular proteins in HCV infection and pathogenesis.

Expression profiling reveals dysregulation of cellular cytoskeletal genes in HBx–induced hepatocarcinogenesis

“…35 Supportively, heterozygous mutations in K8 and K18 have been reported to be associated with various liver diseases, including cryptogenic cirrhosis, biliary atresia, hepatitis C or B, alcohol-induced cirrhosis, primary biliary cirrhosis and acute fulminant hepatitis. 36,37 In addition, proteomic analysis has revealed that K8 and K18 could interact with the core protein of hepatitis C virus, 38 and K8 induced autoantibodies among patients with HCC. 39 In this study, we provided the first evidence to show that the upregulation of K8 and K18 would contribute to the .…”

Functional Replacement of a Domain in the Rubella Virus P150 Replicase Protein by the Virus Capsid Protein

“…In addition to being structural components of the virion, capsid proteins play roles in the virus replication cycle distinct from virion assembly (1,4,6,11,14,15,18,20,27). RUBV C participates in RNA synthesis in an apparent variety of ways.…”

Host–virus interactions during hepatitis C virus infection: a complex and dynamic molecular biosystem

“…Using a combination of affinity-based pull down experiments and mass spectrometric analysis, Kang and co-workers were able to determine that HCV core protein interacts with 14 host cell factors. 134 Some proteins that were identified include heat shock protein 60, implicated in modulation of apoptosis as well as fibrous cytoskeletal proteins, vimentin and cytokeratin 8. Sato et al also examined the lipid droplet proteomes of cells expressing HCV core proteins and found a number of interesting lipid droplet proteins as well as differential expression of certain proteins such as adipocyte differentiationrelated protein (ADRP) and tail-interacting protein of 47 kDa (TIP47).…”

“…Recently proteomic approaches have been applied to study the differential protein expression during HCV protein expression, 61,134,135 replication and infection 136 as well as to discover biomarkers for HCC. [137][138][139][140] One of the first detailed proteomics study related to HCV biology was performed by Yan et al 141 In this study, they used isotope-coded affinity tags (ICAT) to examine differentially expressed proteins during interferon treatment of Huh-7 cells.…”