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Monitoring Viral Entry in Real-Time Using a Luciferase Recombinant Vesicular Stomatitis Virus Producing SARS-CoV-2, EBOV, LASV, CHIKV, and VSV Glycoproteins.
Lay Mendoza, Maria Fernanda; Acciani, Marissa Danielle; Levit, Courtney Nina; Santa Maria, Christopher; Brindley, Melinda Ann.
  • Lay Mendoza MF; Department of Infectious Diseases, College of Veterinary Medicine, University of Georgia, Athens, GA 30602, USA.
  • Acciani MD; Department of Infectious Diseases, College of Veterinary Medicine, University of Georgia, Athens, GA 30602, USA.
  • Levit CN; Department of Infectious Diseases, College of Veterinary Medicine, University of Georgia, Athens, GA 30602, USA.
  • Santa Maria C; Department of Infectious Diseases, College of Veterinary Medicine, University of Georgia, Athens, GA 30602, USA.
  • Brindley MA; Department of Infectious Diseases, College of Veterinary Medicine, University of Georgia, Athens, GA 30602, USA.
Viruses ; 12(12)2020 12 17.
Article in English | MEDLINE | ID: covidwho-979668
ABSTRACT
Viral entry is the first stage in the virus replication cycle and, for enveloped viruses, is mediated by virally encoded glycoproteins. Viral glycoproteins have different receptor affinities and triggering mechanisms. We employed vesicular stomatitis virus (VSV), a BSL-2 enveloped virus that can incorporate non-native glycoproteins, to examine the entry efficiencies of diverse viral glycoproteins. To compare the glycoprotein-mediated entry efficiencies of VSV glycoprotein (G), Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) spike (S), Ebola (EBOV) glycoprotein (GP), Lassa (LASV) GP, and Chikungunya (CHIKV) envelope (E) protein, we produced recombinant VSV (rVSV) viruses that produce the five glycoproteins. The rVSV virions encoded a nano luciferase (NLucP) reporter gene fused to a destabilization domain (PEST), which we used in combination with the live-cell substrate EndurazineTM to monitor viral entry kinetics in real time. Our data indicate that rVSV particles with glycoproteins that require more post-internalization priming typically demonstrate delayed entry in comparison to VSV G. In addition to determining the time required for each virus to complete entry, we also used our system to evaluate viral cell surface receptor preferences, monitor fusion, and elucidate endocytosis mechanisms. This system can be rapidly employed to examine diverse viral glycoproteins and their entry requirements.
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Full text: Available Collection: International databases Database: MEDLINE Main subject: Glycoproteins / Gene Expression / Viral Envelope Proteins / Vesicular stomatitis Indiana virus / Virus Internalization / Genetic Vectors Type of study: Experimental Studies Limits: Animals / Humans Language: English Year: 2020 Document Type: Article Affiliation country: V12121457

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Full text: Available Collection: International databases Database: MEDLINE Main subject: Glycoproteins / Gene Expression / Viral Envelope Proteins / Vesicular stomatitis Indiana virus / Virus Internalization / Genetic Vectors Type of study: Experimental Studies Limits: Animals / Humans Language: English Year: 2020 Document Type: Article Affiliation country: V12121457