The increased expression of capsid protein in trans enhances production of single-round infectious particles by West Nile virus DNA vaccine candidate

Roby, Justin A., Bielefeldt-Ohmann, Helle, Prow, Natalie A., Chang, David C., Hall, Roy A. and Khromykh, Alexander A. (2014) The increased expression of capsid protein in trans enhances production of single-round infectious particles by West Nile virus DNA vaccine candidate. Journal of General Virology, 95 10: 2176-2191. doi:10.1099/vir.0.064121-0

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Author Roby, Justin A.
Bielefeldt-Ohmann, Helle
Prow, Natalie A.
Chang, David C.
Hall, Roy A.
Khromykh, Alexander A.
Title The increased expression of capsid protein in trans enhances production of single-round infectious particles by West Nile virus DNA vaccine candidate
Formatted title
The increased expression of capsid protein in trans enhances production of single-round infectious particles by West Nile virus DNA vaccine candidate
Journal name Journal of General Virology   Check publisher's open access policy
ISSN 0022-1317
1465-2099
Publication date 2014-10-01
Year available 2014
Sub-type Article (original research)
DOI 10.1099/vir.0.064121-0
Open Access Status Not yet assessed
Volume 95
Issue 10
Start page 2176
End page 2191
Total pages 16
Place of publication Spencers Wood, Reading, Berks, United Kingdom
Publisher Society for General Microbiology
Language eng
Abstract West Nile virus (WNV; genus Flavivirus, family Flaviviridae) is an emerging pathogenic arbovirus responsible for outbreaks of encephalitis around the world. Whilst no vaccines are currently available to prevent WNV infection of humans, the use of cDNA copies of flavivirus RNA genomes with large internal deletions within the capsid (C) appears promising. C-deleted vaccines are able to replicate and secrete large amounts of non-infectious immunogenic subviral particles (SVPs) from transfected cells. We have previously generated a WNV DNA vaccine candidate pKUNdC/C where C-deleted WNV cDNA was placed under the control of one copy of the cytomegalovirus (CMV) promoter and the C gene was placed under the control of a second copy of the CMV promoter in the same plasmid DNA. This DNA was shown to generate single-round infectious particles (SRIPs) capable of delivering self-replicating C-deleted RNA producing SVPs to surrounding cells, thus enhancing the vaccine potential. However, the amounts of both SRIPs and SVPs produced from pKUNdC/C DNA were relatively low. In this investigation, we aimed at increasing SRIP production by optimizing trans-C expression via incorporating different forms of C and the use of a more powerful promoter. The construct containing an elongation factor EF1 alpha promoter encoding an extended form of C was demonstrated to produce the highest titres of SRIPs and was immunogenic in mice. Additionally, SRIP and SVP titres were further improved via incorporation of a glycosylation motif in the envelope protein. The optimized DNA yielded similar to 100-fold greater titres of SRIPs than the original construct, thus providing a promising candidate for further vaccine evaluation.
Formatted abstract
The flavivirus West Nile virus (WNV) is an emerging pathogenic arbovirus responsible for outbreaks of encephalitis around the world. Whilst no vaccines are currently available to prevent WNV infection of humans, the use of flavivirus cDNA genomes with large internal deletions within capsid (C) appears promising. C-deleted vaccines are able to replicate and secrete large amounts of non-infectious immunogenic subviral particles (SVPs) from transfected cells. We have previously generated a WNV DNA vaccine candidate pKUNdC/C where C-deleted WNV cDNA was placed under the control of one copy of the cytomegalovirus promoter (CMV) and C gene was placed under the control of a second copy of CMV promoter in the same plasmid DNA. This DNA was shown to generate single-round infectious particles (SRIPs) capable of delivering self-replicating C-deleted RNA producing SVPs to surrounding cells thus enhancing the vaccine potential. However, the amounts of both SRIPs and SVPs produced from pKUNdC/C DNA were relatively low. In this investigation we aimed at increasing SRIP production by optimizing trans-C expression via incorporating different forms of C and the use of a more powerful promoter. The EF1α promoter encoding an extended form of C was demonstrated to produce the highest titers of SRIPs and was immunogenic in mice. Additionally, SRIP and SVP titers were further improved via incorporation of a glycosylation motif in E protein. The optimized DNA yields ~100-fold greater titers of SRIPs than the original construct, thus providing a promising candidate for further vaccine evaluation.
Keyword DNA vaccine
Flaviviruses
West Nile virus
Virus secretion
Q-Index Code C1
Q-Index Status Confirmed Code
Institutional Status UQ

Document type: Journal Article
Sub-type: Article (original research)
Collections: Official 2015 Collection
School of Chemistry and Molecular Biosciences
School of Veterinary Science Publications
 
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Created: Sat, 28 Jun 2014, 00:45:06 EST by Mrs Louise Nimwegen on behalf of School of Chemistry & Molecular Biosciences