Comparison of U2OS and Huh-7 cells for identifying host factors that affect hepatitis C virus RNA replication

Jones, D.M., Domingues, P., Targett-Adams, P. and McLauchlan, J. (2010) Comparison of U2OS and Huh-7 cells for identifying host factors that affect hepatitis C virus RNA replication. Journal of General Virology, 91(9), pp. 2238-2248. (doi:10.1099/vir.0.022210-0) (PMID:20505011)

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Abstract

Host cell factors are critical to all stages of the hepatitis C virus (HCV) life cycle. While many cellular proteins that regulate HCV genome synthesis have been identified, the mechanisms engaged in this process are incompletely understood. To identify novel cellular proteins involved in HCV RNA replication, we screened a library of small interfering RNAs (siRNAs) targeting 299 cellular factors, which principally function in RNA interactions. For the screen, a robust system was established using two cell lines (derived from Huh-7 and U2OS cells) that replicated tricistronic subgenomic replicons (SGRs). We found that the U2OS cell line gave lower levels of intracellular HCV RNA replication compared with Huh-7 cells and was more readily transfected by siRNAs. Consequently, increased gene silencing and greater effects on HCV replication were observed in the U2OS cell line. Thus, U2OS cells provided a suitable and more sensitive alternative to Huh-7 cells for siRNA studies on HCV RNA replication. From the screen, several cellular proteins that enhanced and suppressed HCV RNA replication were identified. One of the genes found to downregulate viral RNA synthesis, ISG15, is expressed in response to alpha interferon and may therefore partly contribute to the clearance of virus from infected individuals. A second gene that inhibited HCV RNA levels was the 5′–3′ exoRNase XRN1, which suggested a role for cellular RNA degradation pathways in modulating the abundance of viral genomes. Therefore, this study provides an important framework for future detailed analyses of these and other cellular proteins.

Item Type:Articles
Status:Published
Refereed:Yes
Glasgow Author(s) Enlighten ID:Domingues, Miss Patricia and Jones, Dr Daniel and Targett-Adams, Dr Paul and McLauchlan, Professor John
Authors: Jones, D.M., Domingues, P., Targett-Adams, P., and McLauchlan, J.
College/School:College of Medical Veterinary and Life Sciences > Institute of Infection Immunity and Inflammation
Journal Name:Journal of General Virology
Journal Abbr.:J. Gen. Virol.
Publisher:Society for General Microbiology
ISSN:0022-1317
ISSN (Online):1465-2099
Published Online:26 May 2010

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Project CodeAward NoProject NamePrincipal InvestigatorFunder's NameFunder RefLead Dept
656341Virus-host interactions in hepatitis C virus infectionJohn MclauchlanMedical Research Council (MRC)MC_UU_12014/1MVLS III - CENTRE FOR VIRUS RESEARCH