The hydrophilic amino-terminal arm of reovirus core shell protein lambda1 is dispensable for particle assembly

J Virol. 2002 Dec;76(23):12211-22. doi: 10.1128/jvi.76.23.12211-12222.2002.

Abstract

The reovirus core particle is a molecular machine that mediates synthesis, capping, and export of the viral plus strand RNA transcripts. Its assembly and structure-function relationships remain to be well understood. Following the lead of previous studies with other Reoviridae family members, most notably orbiviruses and rotaviruses, we used recombinant baculoviruses to coexpress reovirus core proteins lambda1, lambda2, and sigma2 in insect cells. The resulting core-like particles (CLPs) were purified and characterized. They were found to be similar to cores with regard to their sizes, morphologies, and protein compositions. Like cores, they could also be coated in vitro with the two major outer-capsid proteins, micro 1 and sigma3, to produce virion-like particles. Coexpression of core shell protein lambda1 and core nodule protein sigma2 was sufficient to yield CLPs that could withstand purification, whereas expression of lambda1 alone was not, indicating a required role for sigma2 as a previous study also suggested. In addition, CLPs that lacked lambda2 (formed from lambda1 and sigma2 only) could not be coated with micro 1 and sigma3, indicating a required role for lambda2 in the assembly of these outer-capsid proteins into particles. To extend the use of this system for understanding the core and its assembly, we addressed the hypothesis that the hydrophilic amino-terminal region of lambda1, which adopts an extended arm-like conformation around each threefold axis in the reovirus core crystal structure, plays an important role in assembling the core shell. Using a series of lambda1 deletion mutants, we showed that the amino-terminal 230 residues of lambda1, including its zinc finger, are dispensable for CLP assembly. Residues in the 231-to-259 region of lambda1, however, were required. The core crystal structure suggests that residues in the 231-to-259 region are necessary because they affect the interaction of lambda1 with the threefold and/or fivefold copies of sigma2. An effective system for studies of reovirus core structure, assembly, and functions is hereby established.

Publication types

  • Research Support, Non-U.S. Gov't
  • Research Support, U.S. Gov't, Non-P.H.S.
  • Research Support, U.S. Gov't, P.H.S.

MeSH terms

  • Capsid / chemistry*
  • Capsid / physiology*
  • Capsid Proteins*
  • DNA-Binding Proteins / chemistry*
  • DNA-Binding Proteins / genetics
  • DNA-Binding Proteins / physiology*
  • Genes, Viral
  • Genetic Complementation Test
  • Image Processing, Computer-Assisted
  • Microscopy, Electron
  • Models, Molecular
  • Orthoreovirus, Mammalian / genetics
  • Orthoreovirus, Mammalian / physiology*
  • Orthoreovirus, Mammalian / ultrastructure
  • Protein Conformation
  • RNA-Binding Proteins / chemistry*
  • RNA-Binding Proteins / genetics
  • RNA-Binding Proteins / physiology*
  • Recombinant Proteins / chemistry
  • Recombinant Proteins / genetics
  • Recombinant Proteins / metabolism
  • Sequence Deletion
  • Viral Core Proteins / genetics
  • Viral Core Proteins / physiology
  • Virus Assembly / genetics
  • Virus Assembly / physiology*

Substances

  • Capsid Proteins
  • DNA-Binding Proteins
  • RNA-Binding Proteins
  • Recombinant Proteins
  • S2 core protein, Reovirus
  • Viral Core Proteins
  • lambda 1 protein, reovirus
  • mu1 protein, Reovirus