Visualization of Positive and Negative Sense Viral RNA for Probing the Mechanism of Direct-Acting Antivirals against Hepatitis C Virus
Dandan Liu,
Philip R. Tedbury,
Shuiyun Lan,
Andrew D. Huber,
Maritza N. Puray-Chavez,
Juan Ji,
Eleftherios Michailidis,
Mohsan Saeed,
Tanyaradzwa P. Ndongwe,
Leda C. Bassit,
Raymond F. Schinazi,
Robert Ralston,
Charles M. Rice,
Stefan G. Sarafianos
Affiliations
Dandan Liu
CS Bond Life Sciences Center, University of Missouri, Columbia, MO 65201, USA
Philip R. Tedbury
CS Bond Life Sciences Center, University of Missouri, Columbia, MO 65201, USA
Shuiyun Lan
Center for AIDS Research, Laboratory of Biochemical Pharmacology, Department of Pediatrics, Emory University School of Medicine, Atlanta, GA 30322, USA
Andrew D. Huber
CS Bond Life Sciences Center, University of Missouri, Columbia, MO 65201, USA
Maritza N. Puray-Chavez
CS Bond Life Sciences Center, University of Missouri, Columbia, MO 65201, USA
Juan Ji
CS Bond Life Sciences Center, University of Missouri, Columbia, MO 65201, USA
Eleftherios Michailidis
Laboratory of Virology and Infectious Disease, The Rockefeller University, New York, NY 10065, USA
Mohsan Saeed
Laboratory of Virology and Infectious Disease, The Rockefeller University, New York, NY 10065, USA
Tanyaradzwa P. Ndongwe
CS Bond Life Sciences Center, University of Missouri, Columbia, MO 65201, USA
Leda C. Bassit
Center for AIDS Research, Laboratory of Biochemical Pharmacology, Department of Pediatrics, Emory University School of Medicine, Atlanta, GA 30322, USA
Raymond F. Schinazi
Center for AIDS Research, Laboratory of Biochemical Pharmacology, Department of Pediatrics, Emory University School of Medicine, Atlanta, GA 30322, USA
Robert Ralston
CS Bond Life Sciences Center, University of Missouri, Columbia, MO 65201, USA
Charles M. Rice
Laboratory of Virology and Infectious Disease, The Rockefeller University, New York, NY 10065, USA
Stefan G. Sarafianos
CS Bond Life Sciences Center, University of Missouri, Columbia, MO 65201, USA
RNA viruses are highly successful pathogens and are the causative agents for many important diseases. To fully understand the replication of these viruses it is necessary to address the roles of both positive-strand RNA ((+)RNA) and negative-strand RNA ((−)RNA), and their interplay with viral and host proteins. Here we used branched DNA (bDNA) fluorescence in situ hybridization (FISH) to stain both the abundant (+)RNA and the far less abundant (−)RNA in both hepatitis C virus (HCV)- and Zika virus-infected cells, and combined these analyses with visualization of viral proteins through confocal imaging. We were able to phenotypically examine HCV-infected cells in the presence of uninfected cells and revealed the effect of direct-acting antivirals on HCV (+)RNA, (−)RNA, and protein, within hours of commencing treatment. Herein, we demonstrate that bDNA FISH is a powerful tool for the study of RNA viruses that can provide insights into drug efficacy and mechanism of action.
