Generation of synthetic severe acute respiratory syndrome coronavirus pseudoparticles: implications for assembly and vaccine production.
The recently emerged severe acute respiratory syndrome coronavirus (SARS-CoV) contains four structural genes, two replicase-transcriptase open reading frames, and more than five potential genes of unknown function. Despite this relative simplicity, the molecular regulation of SARS-CoV replication and assembly is not understood. Here, we report that two viral genes, encoding the SARS-CoV membrane (M) and nucleocapsid (N) proteins, are necessary and sufficient for formation of virus-like particles. Expression vectors encoding these two proteins were synthesized by using preferred human codons. When M and N expression plasmids were cotransfected into human 293 renal epithelial cells, pseudoparticles formed readily. The addition of a third gene, encoding the spike (S) glycoprotein, facilitated budding of particles that contained a corona-like halo resembling SARS-CoV when examined by transmission electron microscopy, with a buoyant density characteristic of coronaviruses. Specific biochemical interactions of these proteins were also shown in vitro. The S, M, and N proteins of the SARS-CoV are, therefore, necessary and sufficient for pseudovirus assembly. These findings advance the understanding of the morphogenesis of SARS-CoV and enable the generation of safe, conformational mimetics of the SARS virus that may facilitate the development of vaccines and antiviral drugs.
Huang
Yue
Y
Vaccine Research Center, Natiuonal Institute of Allergy and Infectious Diseases, National Institutes of Health, Bldg. 40, Room 4502, MSC 3005, 40 Convent Drive, Bethesda, MD 20982-3005, USA.
Yang
Zhi-yong
ZY
Kong
Wing-pui
WP
Nabel
Gary J
GJ
D001483:Base Sequence; D005814:Genes, Viral / Q000502:physiology*; D006801:Humans; D008969:Molecular Sequence Data; D019251:Nucleocapsid / Q000502:physiology; D019590:Nucleocapsid Proteins / Q000235:genetics; D045473:Severe acute respiratory syndrome-related coronavirus / Q000235:genetics* / Q000276:immunology / Q000502:physiology*; D014763:Viral Matrix Proteins / Q000235:genetics; D014765:Viral Vaccines / Q000276:immunology*; D014771:Virion / Q000502:physiology*; D019065:Virus Assembly
D019590:Nucleocapsid Proteins; D014763:Viral Matrix Proteins; D014765:Viral Vaccines