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Alterations of pr-M cleavage and virus export in pr-M junction chimeric dengue viruses

Poonsook Keelapang1, Roongtawan Sriburi, Sanpaechuda Supasa

  • 1Medical Biotechnology Unit, National Center for Genetic Engineering and Biotechnology, National Science and Technology Development Agency, Bangkok 10400, USA.

Journal of Virology
|February 14, 2004
PubMed

Insights

Flavivirus prM protein cleavage by furin is crucial for infectivity. Altering dengue virus prM sequences with those from other flaviviruses showed enhanced cleavage can unexpectedly hinder virus export, suggesting sequence conservation is key for natural dengue virus strains.

Area of Science:

  • Virology
  • Molecular Biology
  • Protein Biochemistry

Background:

  • Flavivirus particle maturation involves secretory pathway processing, including cleavage of the prM glycoprotein by furin.
  • This prM cleavage is essential for E glycoprotein rearrangement and subsequent virus infectivity.
  • Conserved charged residues in the prM cleavage-proximal region are characteristic of flavivirus antigenic complexes.

Purpose of the Study:

  • To investigate the impact of diverse flavivirus prM junction-proximal sequences on prM cleavage and virus replication.
  • To explore how altered charged residue composition affects dengue virus (DENV) processing and infectivity.
  • To understand the mechanism behind sequence conservation at the DENV prM junction.

Main Methods:

  • Construction of chimeric viruses by replacing the DENV-16681 prM cleavage-proximal region with sequences from tick-borne encephalitis virus (TBEV), yellow fever virus (YFV), and Japanese encephalitis virus (JEV).
  • Generation of chimeric viruses via RNA transfection of mosquito cells.
  • Analysis of prM cleavage efficiency, virion export, replication kinetics, and cell binding in various cell lines (C6/36, PS, Vero).

Main Results:

  • Chimeric viruses showed differential prM cleavage: enhanced in JEVpr/16681, moderately enhanced in YFVpr/16681, and slightly reduced in TBEVpr/16681.
  • JEVpr/16681 exhibited significantly reduced focus size, peak titer, and replication, correlated with delayed virion export but not altered specific infectivity or heparin binding.
  • Greatly enhanced prM cleavability in JEVpr/16681 adversely affected DENV export with minimal impact on infectivity.

Conclusions:

  • Flavivirus prM junction-proximal sequences differentially influence prM cleavage within a DENV background.
  • Excessive prM cleavability can negatively impact DENV particle export, even with minimal effects on infectivity.
  • The observed adverse effects of highly altered prM sequences support a mechanism for maintaining sequence conservation in natural DENV isolates.

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