Identification of RNA-protein interaction networks involved in the norovirus life cycle

J Virol. 2012 Nov;86(22):11977-90. doi: 10.1128/JVI.00432-12. Epub 2012 Aug 29.

Abstract

Human noroviruses are one of the major causes of acute gastroenteritis in the developed world, yet our understanding of their molecular mechanisms of genome translation and replication lags behind that for many RNA viruses. Due to the nonculturable nature of human noroviruses, many related members of the Caliciviridae family of small RNA viruses are often used as model systems to dissect the finer details of the norovirus life cycle. Murine norovirus (MNV) has provided one such system with which to study the basic mechanisms of norovirus translation and replication in cell culture. In this report we describe the use of riboproteomics to identify host factors that interact with the extremities of the MNV genome. This network of RNA-protein interactions contains many well-characterized host factors, including PTB, La, and DDX3, which have been shown to play a role in the life cycle of other RNA viruses. By using RNA coimmunoprecipitation, we confirmed that a number of the factors identified using riboproteomics are associated with the viral RNA during virus replication in cell culture. We further demonstrated that RNA inhibition-mediated knockdown of the intracellular levels of a number of these factors inhibits or slows norovirus replication in cell culture, allowing identification of new intracellular targets for this important group of pathogens.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Cell Line
  • DEAD-box RNA Helicases
  • Humans
  • Immunoprecipitation
  • Macrophages / metabolism
  • Macrophages / virology
  • Mass Spectrometry / methods
  • Mice
  • Microscopy, Fluorescence / methods
  • Norovirus / genetics*
  • Norovirus / physiology*
  • Nucleic Acid Conformation
  • Polypyrimidine Tract-Binding Protein / chemistry
  • Protein Interaction Mapping
  • Proteomics / methods
  • RNA Helicases / chemistry
  • RNA, Viral / genetics*
  • RNA-Binding Proteins / chemistry
  • Rabbits
  • Reticulocytes / cytology

Substances

  • Larp1 protein, mouse
  • RNA, Viral
  • RNA-Binding Proteins
  • Polypyrimidine Tract-Binding Protein
  • DEAD-box RNA Helicases
  • Ddx3x protein, mouse
  • RNA Helicases