Complete in vitro assembly of the reovirus outer capsid produces highly infectious particles suitable for genetic studies of the receptor-binding protein

Kartik Chandran, Xing Zhang, Norman H. Olson, Stephen B. Walker, James D. Chappell, Terence S. Dermody, Timothy S. Baker, Max L. Nibert

Research output: Contribution to journalArticle

44 Citations (Scopus)

Abstract

Mammalian reoviruses, prototype members of the Reoviridae family of nonenveloped double-stranded RNA viruses, use at least three proteins - σ1, μ1, and σ3 - to enter host cells, σ1, a major determinant of cell tropism, mediates viral attachment to cellular receptors. Studies of σ1 functions in reovirus entry have been restricted by the lack of methodologies to produce infectious virions containing engineered mutations in viral proteins. To mitigate this problem, we produced virion-like particles by "recoating" genome-containing core particles that lacked σ1, μ1, and σ3 with recombinant forms of these proteins in vitro. Image reconstructions from cryoelectron micrographs of the recoated particles revealed that they closely resembled native virions in three-dimensional structure, including features attributable to σ1. The recoated particles bound to and infected cultured cells in a σ1-dependent manner and were approximately 1 million times as infectious as cores and 0.5 times as infectious as native virions. Experiments with recoated particles containing recombinant σ1 from either of two different reovirus strains confirmed that differences in cell attachment and infectivity previously observed between those strains are determined by the σ1 protein. Additional experiments showed that recoated particles containing σ1 proteins with engineered mutations can be used to analyze the effects of such mutations on the roles of particle-bound σ1 in infection. The results demonstrate a powerful new system for molecular genetic dissections of σ1 with respect to its structure, assembly into particles, and roles in entry.

Original languageEnglish (US)
Pages (from-to)5335-5342
Number of pages8
JournalJournal of Virology
Volume75
Issue number11
DOIs
StatePublished - 2001
Externally publishedYes

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Reoviridae
capsid
Capsid
Virion
binding proteins
Carrier Proteins
receptors
virion
Mutation
Viral Tropism
Proteins
Computer-Assisted Image Processing
Double-Stranded RNA
RNA Viruses
Viral Proteins
mutation
Recombinant Proteins
Dissection
Molecular Biology
Cultured Cells

ASJC Scopus subject areas

  • Immunology

Cite this

Complete in vitro assembly of the reovirus outer capsid produces highly infectious particles suitable for genetic studies of the receptor-binding protein. / Chandran, Kartik; Zhang, Xing; Olson, Norman H.; Walker, Stephen B.; Chappell, James D.; Dermody, Terence S.; Baker, Timothy S.; Nibert, Max L.

In: Journal of Virology, Vol. 75, No. 11, 2001, p. 5335-5342.

Research output: Contribution to journalArticle

Chandran, Kartik ; Zhang, Xing ; Olson, Norman H. ; Walker, Stephen B. ; Chappell, James D. ; Dermody, Terence S. ; Baker, Timothy S. ; Nibert, Max L. / Complete in vitro assembly of the reovirus outer capsid produces highly infectious particles suitable for genetic studies of the receptor-binding protein. In: Journal of Virology. 2001 ; Vol. 75, No. 11. pp. 5335-5342.
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