Complete genomic RNA sequence of western equine encephalitis virus and expression of the structural genes.
Abstract: The complete nucleotide sequence of the 71V-1658 strain of western equine encephalitis virus (WEE) was determined (minus 25 nucleotides from the 5' end). A 5' RACE reaction was used to sequence the 5' terminus from WEE strain CBA87. The deduced WEE genome was 11508 nucleotides in length, excluding the 5' cap nucleotide and 3' poly(A) tail. The nucleotide composition was 28% A, 25% C, 25% G and 22% U. Comparison with partial WEE sequences of strain 5614 (nsP2-nsP3 of the nonstructural region) and strain BFS1703 (26S structural region) revealed comparatively little variation; a total of 149 nucleotide differences in 8624 bases (1.7% divergence), of which only 28% (42 nucleotides) altered the encoded amino acids. Comparison of deduced nsP1 and nsP4 amino acid sequences from WEE with the corresponding proteins from eastern equine encephalitis virus (EEE) yielded identities of 84.9 and 83.8%, respectively. Previously uncharacterized stem-loop structures were identified in the nontranslated terminal regions. A cDNA clone of the 26S region encoding the structural polyprotein of WEE strain 71V-1658 was placed under the control of a cytomegalovirus promoter and transfected into tissue culture cells. The viral envelope proteins were functionally expressed in tissue culture, as determined by histochemical staining with monoclonal antibodies that recognize WEE antigens, thus, forming the initial step in the investigation of subunit vaccines to WEE.
Publication Date: 2000-01-21 PubMed ID: 10640553DOI: 10.1099/0022-1317-81-1-151Google Scholar: Lookup
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- Journal Article
- Research Support
- Non-U.S. Gov't
Summary
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This research focused on the full study and identification of the genomic sequence of western equine encephalitis virus, a neurotropic virus. They also managed to synthetically express the virus’s structural genes, a crucial first step towards developing vaccines for the disease.
Sequence Determination & Comparison
- The researchers successfully determined the nucleotide sequence of the western equine encephalitis virus (WEE), except for 25 nucleotides from one end. The full sequence was found to be 11508 nucleotides long and had an equal ratio of each nucleotide type (A, C, G, U).
- Additionally, they compared this sequencing with the partial sequences of two other WEE strains, which revealed little variation. Out of 8624 bases, only 149 nucleotide differences were found, and just 42 of these differences altered the encoded amino acids.
- Comparison between WEE and eastern equine encephalitis virus (EEE) sequences also showed high identical rates (84.9% and 83.8%, respectively) in certain parts called nsP1 and nsP4. This similarity could suggest commonalities in how the viruses function or developed.
Identification of Stem-loop Structures
- They also found previously unidentified stem-loop structures in the genomic sequence of WEE. These structures, present in the nontranslated terminal regions of the sequence, could have implications in the replication and translation of the virus, and thus may be instrumental in designing treatments or vaccines.
Successful Expression of Structural Genes
- The researchers managed to express the structural genes of WEE strain within laboratory cell cultures. They accomplished this by creating a cDNA clone of the portion of the WEE genome that codes for the structural polyprotein.
- The resulting expression of the WEE’s structural proteins was verified by staining the cell cultures with antibodies specific to the virus. The successful expression in cells portrays the potential for vaccine production, as these proteins could stimulate an immune response. This finding represents an important starting point in the development of possible vaccines for WEE.
Cite This Article
APA
Netolitzky DJ, Schmaltz FL, Parker MD, Rayner GA, Fisher GR, Trent DW, Bader DE, Nagata LP.
(2000).
Complete genomic RNA sequence of western equine encephalitis virus and expression of the structural genes.
J Gen Virol, 81(Pt 1), 151-159.
https://doi.org/10.1099/0022-1317-81-1-151 Publication
Researcher Affiliations
- Defence Research Establishment Suffield, Medical Countermeasures Section, PO Box 4000 Station Main, Medicine Hat, Alberta, Canada.
MeSH Terms
- 5' Untranslated Regions / genetics
- Base Sequence
- Cell Line
- Encephalitis Virus, Western Equine / genetics
- Encephalitis Virus, Western Equine / growth & development
- Gene Expression Profiling
- Genes, Viral
- Genome, Viral
- Molecular Sequence Data
- Phylogeny
- RNA, Ribosomal / genetics
- RNA, Viral / genetics
- Reverse Transcriptase Polymerase Chain Reaction
- Sequence Analysis, DNA
- Viral Nonstructural Proteins / genetics
- Viral Structural Proteins / genetics
Citations
This article has been cited 9 times.- Abdoulaye AH, Foda MF, Kotta-Loizou I. Viruses Infecting the Plant Pathogenic Fungus Rhizoctonia solani.. Viruses 2019 Nov 30;11(12).
- Kumar B, Manuja A, Gulati BR, Virmani N, Tripathi BN. Zoonotic Viral Diseases of Equines and Their Impact on Human and Animal Health.. Open Virol J 2018;12:80-98.
- Allison AB, Stallknecht DE, Holmes EC. Evolutionary genetics and vector adaptation of recombinant viruses of the western equine encephalitis antigenic complex provides new insights into alphavirus diversity and host switching.. Virology 2015 Jan 1;474:154-62.
- Bergren NA, Auguste AJ, Forrester NL, Negi SS, Braun WA, Weaver SC. Western equine encephalitis virus: evolutionary analysis of a declining alphavirus based on complete genome sequences.. J Virol 2014 Aug;88(16):9260-7.
- Phillips AT, Schountz T, Toth AM, Rico AB, Jarvis DL, Powers AM, Olson KE. Liposome-antigen-nucleic acid complexes protect mice from lethal challenge with western and eastern equine encephalitis viruses.. J Virol 2014 Feb;88(3):1771-80.
- Gauci PJ, Wu JQ, Rayner GA, Barabé ND, Nagata LP, Proll DF. Identification of Western equine encephalitis virus structural proteins that confer protection after DNA vaccination.. Clin Vaccine Immunol 2010 Jan;17(1):176-9.
- Atasheva S, Wang E, Adams AP, Plante KS, Ni S, Taylor K, Miller ME, Frolov I, Weaver SC. Chimeric alphavirus vaccine candidates protect mice from intranasal challenge with western equine encephalitis virus.. Vaccine 2009 Jul 9;27(32):4309-19.
- Logue CH, Bosio CF, Welte T, Keene KM, Ledermann JP, Phillips A, Sheahan BJ, Pierro DJ, Marlenee N, Brault AC, Bosio CM, Singh AJ, Powers AM, Olson KE. Virulence variation among isolates of western equine encephalitis virus in an outbred mouse model.. J Gen Virol 2009 Aug;90(Pt 8):1848-1858.
- Wielgosz MM, Raju R, Huang HV. Sequence requirements for Sindbis virus subgenomic mRNA promoter function in cultured cells.. J Virol 2001 Apr;75(8):3509-19.
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