Heterologous viral RNA export elements improve expression of severe acute respiratory syndrome (SARS) coronavirus spike protein and protective efficacy of DNA vaccines against SARS.
The SARS-CoV spike glycoprotein (S) is the main target of the protective immune response in humans and animal models of SARS. Here, we demonstrated that efficient expression of S from the wild-type spike gene in cultured cells required the use of improved plasmid vectors containing donor and acceptor splice sites, as well as heterologous viral RNA export elements, such as the CTE of Mazon-Pfizer monkey virus or the PRE of Woodchuck hepatitis virus (WPRE). The presence of both splice sites and WPRE markedly improved the immunogenicity of S-based DNA vaccines against SARS. Upon immunization of mice with low doses (2 microg) of naked DNA, only intron and WPRE-containing vectors could induce neutralizing anti-S antibodies and provide protection against challenge with SARS-CoV. Our observations are likely to be useful for the construction of plasmid and viral vectors designed for optimal expression of intronless genes derived from cytoplasmic RNA viruses.
Callendret
Benoît
B
Unité de Génétique Moléculaire des Virus Respiratoires, URA CNRS 1966, EA 302 Université Paris 7, France.
Lorin
Valérie
V
Charneau
Pierre
P
Marianneau
Philippe
P
Contamin
Hugues
H
Betton
Jean-Michel
JM
van der Werf
Sylvie
S
Escriou
Nicolas
N
D000818:Animals; D000914:Antibodies, Viral / Q000097:blood / Q000276:immunology; D002460:Cell Line; D004742:Enhancer Elements, Genetic / Q000502:physiology*; D005260:Female; D005786:Gene Expression Regulation; D005822:Genetic Vectors / Q000235:genetics / Q000378:metabolism; D017919:Hepatitis B Virus, Woodchuck / Q000235:genetics*; D006801:Humans; D007114:Immunization; D007273:Injections, Intramuscular; D016093:Mason-Pfizer monkey virus / Q000235:genetics*; D008562:Membrane Glycoproteins / Q000235:genetics / Q000276:immunology* / Q000378:metabolism*; D051379:Mice; D008807:Mice, Inbred BALB C; D009500:Neutralization Tests; D010957:Plasmids / Q000235:genetics / Q000378:metabolism; D045473:Severe acute respiratory syndrome-related coronavirus / Q000235:genetics / Q000276:immunology* / Q000378:metabolism*; D045169:Severe Acute Respiratory Syndrome / Q000097:blood / Q000276:immunology* / Q000517:prevention & control*; D064370:Spike Glycoprotein, Coronavirus; D014162:Transfection; D019444:Vaccines, DNA / Q000008:administration & dosage / Q000276:immunology; D014759:Viral Envelope Proteins / Q000235:genetics / Q000276:immunology* / Q000378:metabolism*; D014765:Viral Vaccines / Q000008:administration & dosage / Q000276:immunology*
D000914:Antibodies, Viral; D008562:Membrane Glycoproteins; D064370:Spike Glycoprotein, Coronavirus; D019444:Vaccines, DNA; D014759:Viral Envelope Proteins; D014765:Viral Vaccines; C578557:spike glycoprotein, SARS-CoV