Organization of Flaviviral Replicase Proteins in Virus-Induced Membranes: a Role for NS1′ in Japanese Encephalitis Virus RNA Synthesis

  1. Gregory Bock Organizer and
  2. Jamie Goode
  1. Vijaya Satchidanandam,
  2. Pradeep Devappa Uchil and
  3. Priti Kumar

Published Online: 7 OCT 2008

DOI: 10.1002/0470058005.ch10

New Treatment Strategies for Dengue and Other Flaviviral Diseases: Novartis Foundation Symposium 277

New Treatment Strategies for Dengue and Other Flaviviral Diseases: Novartis Foundation Symposium 277

How to Cite

Satchidanandam, V., Uchil, P. D. and Kumar, P. (2006) Organization of Flaviviral Replicase Proteins in Virus-Induced Membranes: a Role for NS1′ in Japanese Encephalitis Virus RNA Synthesis, in New Treatment Strategies for Dengue and Other Flaviviral Diseases: Novartis Foundation Symposium 277 (eds G. Bock and J. Goode), John Wiley & Sons, Ltd, Chichester, UK. doi: 10.1002/0470058005.ch10

Author Information

  1. Department of Microbiology and Cell Biology, Room 254A, Sir C.V. Raman Avenue, Indian Institute of Science, Bangalore 560012, India

Publication History

  1. Published Online: 7 OCT 2008
  2. Published Print: 25 AUG 2006

Book Series:

  1. Novartis Foundation Symposia

Book Series Editors:

  1. Novartis Foundation

ISBN Information

Print ISBN: 9780470016435

Online ISBN: 9780470058008

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Keywords:

  • flaviviral replicase protein organization;
  • ‘emerging infectious agents’- West Nile (WNV), (DENV) and Japanese encephalitis (JEV);
  • flavivirus replicase proteins in catalytic amounts;
  • flaviviral replication complexes and in vitro RdRp assay;
  • replicase proteins

Summary

The organization of flaviviral replicase proteins within the membrane-bound replication complexes of West Nile (WNV), dengue (DENV) and Japanese encephalitis viruses (JEV) was probed by investigating the combined effect of detergents and trypsin on both viral replicase activity and profile of metabolically labelled viral proteins. While trypsin treatment of virus-induced membrane fractions degraded the vast majority of replicase proteins, viral RNA-dependent RNA polymerase (RdRp) activity remained completely unaffected. Solubilization of the membranes with deoxycholate (DOC) however rendered the replicase accessible to trypsin. Triton X-100 (TX100) treatment reduced RdRp activity by half in WNV but totally destroyed RdRp activity in JEV. TX100 also dissociated NS1′ in addition to NS1 from NS5 and NS3 in JEV. Antibodies to NS3 coprecipitated NS1′ along with NS5 only from DOC-solubilized but not from TX100-treated extracts, the former of which alone retained RdRp activity. Exogenous addition of recombinant NS1′ to TX100 treated JEV-induced membranes restored the defect in the release step of RNA synthesis. Our results suggest for the first time a direct role for JEV NS1′ in viral RNA synthesis in vitro.