Expression of L protein of Hantaan virus 84FLi strain and its application for recovery of minigenomes

APMIS. 2008 Dec;116(12):1089-96. doi: 10.1111/j.1600-0463.2008.01011.x.

Abstract

Hantaan virus (HTNV) is an Old World hantavirus associated with hemorrhagic fever with renal syndrome (HFRS). To visualize the localization of the L protein of HTNV strain 84FLi within cells, a fusion protein composed of enhanced green fluorescent protein and L protein, EGFP-L, was expressed in Vero cells. The 273 KDa expressed fusion protein of EGFP-L localized in the perinuclear region. We also described the development of a reverse genetics system for HTNV strain 84FLi. The RNA polymerase I (pol I)-mediated transcription system was used to generate artificial viral RNA genome segments (minigenomes), which contained the chloramphenicol acetyltransferase (CAT) reporter gene in antisense (virus RNA) or sense (virus-complementary RNA) orientation flanked by the noncoding regions of HTNV 84FLi L segment. CAT could be detected in cells after transfection, indicating the successful encapsidation, transcription and replication of the pol I-derived minigenomes. The passaged transfer of CAT demonstrates that recombinant virus containing packaged pol I-derived minigenomes has been produced. This system may be helpful in studying the gene function and pathogenesis of HTNV.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Chloramphenicol O-Acetyltransferase / genetics
  • Chlorocebus aethiops
  • Cytoplasm / metabolism
  • Gene Expression
  • Genes, Reporter
  • Genome, Viral
  • Green Fluorescent Proteins / analysis
  • Green Fluorescent Proteins / biosynthesis
  • Hantaan virus / genetics
  • Hantaan virus / physiology*
  • Hantavirus Infections / virology*
  • Humans
  • RNA Polymerase I / metabolism
  • RNA, Viral / genetics
  • RNA-Dependent RNA Polymerase / analysis*
  • RNA-Dependent RNA Polymerase / biosynthesis
  • Recombinant Fusion Proteins / antagonists & inhibitors*
  • Recombinant Fusion Proteins / biosynthesis
  • Reverse Transcription
  • Vero Cells / metabolism
  • Viral Proteins / analysis*
  • Viral Proteins / biosynthesis

Substances

  • RNA, Viral
  • Recombinant Fusion Proteins
  • Viral Proteins
  • enhanced green fluorescent protein
  • Green Fluorescent Proteins
  • Chloramphenicol O-Acetyltransferase
  • L protein, hantavirus
  • RNA-Dependent RNA Polymerase
  • RNA Polymerase I