In vivo analysis of infectivity, fusogenicity, and incorporation of a mutagenic viral glycoprotein library reveals determinants for virus incorporation

Daniel J. Salamango, Khalid K. Alam, Donald H. Burke, Marc C. Johnson

Research output: Contribution to journalArticlepeer-review

4 Scopus citations

Abstract

Enveloped viruses utilize transmembrane surface glycoproteins to gain entry into target cells. Glycoproteins from diverse viral families can be incorporated into nonnative viral particles in a process termed pseudotyping; however, the molecular mechanisms governing acquisition of these glycoproteins are poorly understood. For murine leukemia virus envelope (MLV Env) glycoprotein, incorporation into foreign viral particles has been shown to be an active process, but it does not appear to be caused by direct interactions among viral proteins. In this study, we coupled in vivo selection systems with Illumina next-generation sequencing (NGS) to test hundreds of thousands of MLV Env mutants for the ability to be enriched in viral particles and to perform other glycoprotein functions. NGS analyses on a subset of these mutants predicted that the residues important for incorporation are in the membrane-proximal external region (MPER), particularly W127 and W137, and the residues in the membranespanning domain (MSD) and also immediately flanking it (T140 to L163). These predictions were validated by directly measuring the impact of mutations in these regions on fusogenicity, infectivity, and incorporation. We suggest that these two regions dictate pseudotyping through interactions with specific lipid environments formed during viral assembly.

Original languageEnglish (US)
Pages (from-to)6502-6514
Number of pages13
JournalJournal of virology
Volume90
Issue number14
DOIs
StatePublished - 2016

Bibliographical note

Funding Information:
This work, including the efforts of Donald H Burke, was funded by HHS | NIH | National Institute of Allergy and Infectious Diseases (NIAID) (AI074389). This work, including the efforts of Marc C Johnson, was funded by HHS | NIH | National Institute of General Medical Sciences (NIGMS) (GM 110776).

Publisher Copyright:
© 2016, American Society for Microbiology.

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