title: Visualizing infection spread: dual-color fluorescent reporting of virus-host interactions.
abstract: Although the molecular mechanisms by which host cells defend themselves against viral infection have been studied in great depth, and countermeasures viruses employ to suppress such defensive responses have been widely documented, relatively little attention has been devoted toward elucidating how such interactions between virus and host are resolved over multiple rounds of infection. Here, we describe the design, synthesis, and validation of a dual-color fluorescent reporter system to study how viral infections spread through a host cell monolayer and how the cellular innate immune system mounts an antiviral response. We employed recombinant, red fluorescent protein expressing mutants of a prototypical RNA virus, vesicular stomatitis virus to enable identification and tracking of infected cells. Further, we generated stable reporter cells that use green fluorescent protein to report on the expression of IFIT2, an interferon stimulated gene involved in the interference of viral protein translation, and a marker of antiviral defense activation. The presence of the fluorescent protein reporters had minimal effects on the normal behavior of the cells or viruses. Moreover, expression of the virus and cell reporters correlated with the kinetics of viral replication and activation of an anti-viral response, respectively. This two-color system enabled us to track and quantify in live cells how viral replication and activation of host defensive responses play out over multiple rounds of infection. Initial study of propagating infections demonstrated that antiviral activation over multiple rounds was critical for slowing and ultimately halting the spread of infection.
authors: Department of Chemical and Biological Engineering, University of Wisconsin-Madison, 1415 Engineering Drive, Madison, Wisconsin, 53706-1607; Wisconsin Institute for Discovery-Systems Biology Theme, University of Wisconsin-Madison, Madison, Wisconsin.
authors: Adam
authors: A
authors: Swick
authors: Ashley
authors: A
authors: Baltes
authors: John
authors: J
authors: Yin
mesh_terms: D051017:Apoptosis Regulatory Proteins; D003116:Color; D054884:Host-Pathogen Interactions; D008164:Luminescent Proteins / Q000032:analysis / Q000235:genetics; D008856:Microscopy, Fluorescence / Q000379:methods*; D011506:Proteins / Q000032:analysis / Q000235:genetics; D016601:RNA-Binding Proteins; D011994:Recombinant Proteins / Q000032:analysis / Q000235:genetics; D013194:Staining and Labeling / Q000379:methods*; D018116:Vesiculovirus / Q000254:growth & development / Q000502:physiology*; D014773:Virology / Q000379:methods*
chemical_list: D051017:Apoptosis Regulatory Proteins; C484289:IFIT2 protein, human; D008164:Luminescent Proteins; D011506:Proteins; D016601:RNA-Binding Proteins; D011994:Recombinant Proteins
keywords: infection spread; innate immune response; interferon; reporter virus; vesicular stomatitis virus; virus-cell interactions