Hepatitis C virus (HCV) infects approximately 2% of the world's population and is the primary cause of liver transplants in the United States. Based on lessons learned from diseases such as AIDS, HCV RNA replication is a promising target for antiviral development. However, the replication of all viruses with plus-strand RNA genomes is poorly understood, especially at the biochemical level. The overall goal of the research in the Kao lab is to understand mechanism of RNA virus replication. The goal for this project is to build knowledge about the subunits of the HCV replication complex, with emphasis on protein-RNA, and protein-protein interactions. This is an extension of the past six years of research in the Kao lab, where a number of basic properties of the HCV polymerase and the HCV protease-helicase have been examined using biochemical, biophysical, and cell-based methods. The research can be partitioned into several related subaims that will: 1. Identify and validate the biological importance of the residues in the HCV RdRp that interacts with the substrate NTPs, the template RNA, and during different stages of HCV RNA synthesis. 2. Elucidate the interactions between the HCV RdRp and the nascent RNA and identify and characterize the nascent RNA exit channel. 3. Examine the protein-protein and protein-RNA interactions with other replicase- associated subunits of the HCV replicase and examine effects of the interactions on HCV replicase formation in cells. 4. Obtain images of the protein complexes using electron microscopy and reconstruct their structures. Results from this proposal will advance the understanding of the mechanism of viral RNA-dependent RNA synthesis for ALL positive-strand RNA viruses. The knowledge can also be used to compare the mechanisms of action of all template-dependent (both viral and cellular) polymerases.

Agency
National Institute of Health (NIH)
Institute
National Institute of Allergy and Infectious Diseases (NIAID)
Type
Research Project (R01)
Project #
5R01AI073335-05
Application #
8204883
Study Section
Virology - B Study Section (VIRB)
Program Officer
Koshy, Rajen
Project Start
2008-01-01
Project End
2013-12-31
Budget Start
2012-01-01
Budget End
2013-12-31
Support Year
5
Fiscal Year
2012
Total Cost
$332,912
Indirect Cost
$113,564
Name
Indiana University Bloomington
Department
Biology
Type
Schools of Arts and Sciences
DUNS #
006046700
City
Bloomington
State
IN
Country
United States
Zip Code
47401
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Fan, Baochang; Lu, Kuan-Yi; Reymond Sutandy, F X et al. (2014) A human proteome microarray identifies that the heterogeneous nuclear ribonucleoprotein K (hnRNP K) recognizes the 5' terminal sequence of the hepatitis C virus RNA. Mol Cell Proteomics 13:84-92
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Yamane, Daisuke; McGivern, David R; Wauthier, Eliane et al. (2014) Regulation of the hepatitis C virus RNA replicase by endogenous lipid peroxidation. Nat Med 20:927-35
Vaughan, Robert; Li, Yi; Fan, Baochang et al. (2012) RNA binding by the NS3 protease of the hepatitis C virus. Virus Res 169:80-90
Shu, Chang; Yi, Guanghui; Watts, Tylan et al. (2012) Structure of STING bound to cyclic di-GMP reveals the mechanism of cyclic dinucleotide recognition by the immune system. Nat Struct Mol Biol 19:722-4
Vaughan, Robert; Fan, Baochang; You, Jin-Sam et al. (2012) Identification and functional characterization of the nascent RNA contacting residues of the hepatitis C virus RNA-dependent RNA polymerase. RNA 18:1541-52
Yi, Guanghui; Deval, Jerome; Fan, Baochang et al. (2012) Biochemical study of the comparative inhibition of hepatitis C virus RNA polymerase by VX-222 and filibuvir. Antimicrob Agents Chemother 56:830-7
Lu, Cheng; Ranjith-Kumar, C T; Hao, Lujiang et al. (2011) Crystal structure of RIG-I C-terminal domain bound to blunt-ended double-strand RNA without 5' triphosphate. Nucleic Acids Res 39:1565-75

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