With a single exception, all isolates of hepatitis C virus (HCV) require adaptive mutations to replicate efficiently in cell culture. Here, we show that a major class of adaptive mutations regulates the activity of a cellular lipid kinase, phosphatidylinositol 4-kinase IIIα (PI4KA). HCV needs to stimulate PI4KA to create a permissive phosphatidylinositol 4-phosphate-enriched membrane microenvironment in the liver and in primary human hepatocytes (PHHs). In contrast, in Huh7 hepatoma cells, the virus must acquire loss-of-function mutations that prevent PI4KA overactivation. This adaptive mechanism is necessitated by increased PI4KA levels in Huh7 cells compared with PHHs, and is conserved across HCV genotypes. PI4KA-specific inhibitors promote replication of unadapted viral isolates and allow efficient replication of patient-derived virus in cell culture. In summary, this study has uncovered a long-sought mechanism of HCV cell-culture adaptation and demonstrates how a virus can adapt to changes in a cellular environment associated with tumorigenesis.
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The authors would especially like to thank U. Herian, R. Klein, S. Kallis and M. Bartenschlager for excellent technical assistance. We are grateful to T. Wakita for the JFH-1 isolate, to S. Breitfelder for compounds PI4KA-A and PI4KA-B, J. Botyanszki for compounds PI4KA-F1 and PI4KA-G1 and M. Harris for the polyclonal sheep serum recognizing NS5A. We are grateful to the Electron Microscopy Core Facility at the Bioquant (Heidelberg University) for providing access to their equipment and for excellent support. This project was funded by grants from the Deutsche Forschungsgemeinschaft (LO 1556/1-2, LO 1556/4-1 and TRR77, TPA1 to V.L., and TRR83, TP13 and TRR77, TPA1 to R.B.), as well as an HBIGS Postdoc stipend to C.H. This work was supported in part by the National Institutes of Health National Cancer Institute grant R01CA057973 and National Institute of Allergy and Infectious Diseases grants R01AI072613 and R01AI099284 (to C.M.R.) and by a Helmsley Postdoctoral Fellowship for Basic and Translational Research on Disorders of the Digestive System at The Rockefeller University (to M.S.). The Greenberg Medical Research Institute, Starr Foundation, Ronald A. Shellow, M.D. Memorial Fund and anonymous donors provided additional funding (to C.M.R.).
The authors declare no competing financial interests.
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Harak, C., Meyrath, M., Romero-Brey, I. et al. Tuning a cellular lipid kinase activity adapts hepatitis C virus to replication in cell culture. Nat Microbiol 2, 16247 (2017). https://doi.org/10.1038/nmicrobiol.2016.247
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