Genomes and Genes
Roles for microRNA-122 in hepatitis C virus RNA amplification
Principal Investigator: Peter Sarnow
Abstract: DESCRIPTION (provided by applicant): An estimated 170 million people worldwide and 4 million people in the United States are infected with hepatitis C virus (HCV). The majorities of patients do not resolve the infection and develop chronic infections that often lead to end-stage liver disease and hepatocellular carcinoma. Current treatment is limited to a combination therapy of ribavirin and interferon 1. This therapy is expensive and ineffective in 50% of infected individuals. Thus, there is an urgent need to identify viral or cellular molecules that can be used as novel targets in antiviral therapy. It was discovered that HCV binds two molecules of a liver-specific microRNA, miR-122, resulting in a novel, unprecedented upregulation of the viral genome. Sequestration of miR-122 in HCV-infected cultured cells or in livers of infected chimpanzees leads to a dramatic loss of infectious virus without emergence of resistant virus. Therefore, the dependence of HCV on miR-122 presents an Achilles heel of the virus that can be explored for antiviral intervention. This application proposes to study the roles for miR-122 in the viral life cycle and in cholesterol biosynthesis using a novel class of antisense molecules, locked nucleic acids (LNAs) that can easily be delivered to the liver in animals where it sequesters miR-122 in an inactive small duplex RNA. In particular, the first aim will characterize the RNA-RNA interactions in the miR-122/HCV complex, using genetic and biochemical approaches. Aim 2 will test the hypothesis that miR-122 protects the 5'end sequences of the HCV RNA from degradation by ribonucleases or RNA modification enzymes, or aids in the avoidance of activation of double-stranded RNA sensors such as the retinoic acid inducible gene I. These studies will be performed in specific and genome-wide siRNA-mediated gene knockdown experiments. Aim 3 proposes to examine roles for the known isoforms of miR-122 that contain extra 3'terminal adenosine or guanosine residues, on HCV RNA abundance. Deep sequencing analysis and gene knockdown of suspected nucleotidyl transferases will aid in this analysis. The final aim will characterize in detail the mechanism by which miR-122 regulates the expression of Insig1, the major negative regulator of cholesterol and fatty acid metabolism in the liver. In particular, the miR-122-mediated down-regulation of a distinct polyadenylation/cleavage site in a specific Insig1 isoform mRNAs will be examined. Overall, this application will address fundamental aspects about the functions of miR-122 in the HCV life cycle and cholesterol metabolism. The outcomes from these studies will detail novel mechanisms of gene expression mediated by microRNAs in eukaryotic cells and will point to new venues for antiviral therapies. PUBLIC HEALTH RELEVANCE: An estimated 170 million people worldwide and 4 million in the United States are infected with hepatitis C virus (HCV). The majority of patients do not resolve the infection and become chronic carriers, ultimately needing expensive liver transplants. There is no vaccine for HCV, and current treatments, which include ribavirin and interferon 1, are expensive and relatively ineffective. It was discovered that HCV binds two molecules of a liver- specific microRNA, miR-122, resulting in a novel, unprecedented upregulation of the viral genome. This proposal explores the mechanisms by which miR-122 protects HCV RNA in the liver. The dependence of HCV on miR-122 presents an Achilles heel of the virus that can be used for antiviral intervention. We will study the roles for miR-122 in the viral life cycle and in cholesterol biosynthesis using a novel class of antisense molecules (locked nucleic acids) that can easily bind and inactivate miR-122 in the liver of animals. This is a highly significant approach, because LNA-mediated sequestration of miR-122 in the liver of HCV-infected chimpanzees resulted in a 2.5 fold drop in viral load without any emergence of resistant virus (Lanford et al. 2010. Science: 327:198-201).
Funding Period: 2006-02-15 - 2016-01-31
more information: NIH RePORT
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Department of Biochemistry, University of Cambridge, Cambridge CB2 1GA, UK Department of Microbiology and Immunology, Stanford University School of Medicine, Stanford, CA 94305, USA
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Department of Microbiology and Immunology, Stanford University School of Medicine, Stanford, CA 94305, USAA
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Department of Microbiology and Immunology, Stanford University School of Medicine, Stanford, California, United States of America
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Department of Microbiology and Immunology, Stanford University School of Medicine, Stanford, CA 94305, USA
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Department of Medicine, University of Massachusetts Medical School, MA 01605, United States
J Hepatol 57:462-6. 2012..In this review, we highlight the latest advances in the field of microRNA biology and their potential as emerging therapeutic targets in liver disease...
- Combating hepatitis C virus by targeting microRNA-122 using locked nucleic acidsErica S Machlin
Department of Microbiology and Immunology, Stanford University School of Medicine, CA 94305, USA
Curr Gene Ther 12:301-6. 2012..This review summarizes the success of sequestration of liver-specific microRNA miR-122 by antisense locked nucleic acids and their use in combating hepatitis C virus in clinical trials...
- Modulation of hepatitis C virus RNA abundance and virus release by dispersion of processing bodies and enrichment of stress granulesCara T Pager
Department of Microbiology and Immunology, Stanford University, Stanford, CA 94305 5124, United States
Virology 435:472-84. 2013..These data argue that HCV subverts P-body and stress granule components to aid in viral gene expression at particular sites in the cytoplasm...
- Modulation of GB virus B RNA abundance by microRNA-122: dependence on and escape from microRNA-122 restrictionSelena M Sagan
Department of Microbiology and Immunology, Stanford University School of Medicine, Stanford, California, USA
J Virol 87:7338-47. 2013..This finding suggests that structural features at the end of the viral genome dictate whether miR-122 is required to aid in maintaining viral RNA abundance...
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