title: Protein interactions during coronavirus assembly.
abstract: Coronaviruses assemble and obtain their envelope at membranes of the intermediate compartment between the endoplasmic reticulum and Golgi complex. Like other enveloped viruses, coronavirus assembly is presumably dependent on protein localization and protein-protein as well as protein-RNA interactions. We have used the bovine coronavirus (BCV) as a model to study interactions between the viral proteins in virus-infected cells that are important for coronavirus assembly. BCV is a prototype for the coronaviruses that express an additional major structural protein, the hemagglutinin esterase (HE), in addition to the spike (S) glycoprotein, membrane (M) glycoprotein, and nucleocapsid (N) protein. Complexes consisting of the M, S, and HE proteins were detected in virus-infected cells by coimmunoprecipitations. Kinetic analyses demonstrated that S protein and HE each quickly formed a complex with M protein after synthesis, whereas heterocomplexes consisting of all three proteins formed more slowly. The kinetics of HE biosynthesis revealed that the half-life of oligomerization was approximately 30 min, which correlated with the appearance of complexes consisting of M, HE, and S proteins, suggesting that oligomerization and/or conformational changes may be important for the S-M-HE protein complexes to form. Only HE dimers were found associated with the heterocomplexes consisting of all three proteins. S-M-HE protein complexes were detected prior to processing of the oligosaccharide chains on HE, indicating that these protein complexes formed in a premedial Golgi compartment before trimming of sugar chains. Transient coexpressions and double-labeling immunofluorescence demonstrated that HE and S proteins colocalized with M protein. This was further supported by coimmunoprecipitation of specific HE-M and S-M protein complexes from transfected cells, indicating that these proteins can form complexes in the absence of other viral proteins.
authors.affiliation: Division of Molecular Virology, Baylor College of Medicine, Houston, Texas 77030, USA.
authors.forename: V P
authors.initials: VP
authors.lastname: Nguyen
authors.forename: B G
authors.initials: BG
authors.lastname: Hogue
mesh_terms: D000818:Animals; D002417:Cattle; D002451:Cell Compartmentation; D002460:Cell Line; D002522:Chlorocebus aethiops; D000086862:Coronavirus M Proteins; D017938:Coronavirus, Bovine / Q000378:metabolism / Q000502:physiology*; D006224:Cricetinae; D019281:Dimerization; D006023:Glycoproteins / Q000378:metabolism; D006367:HeLa Cells; D006389:Hemagglutinins, Viral / Q000378:metabolism; D006801:Humans; D007700:Kinetics; D008562:Membrane Glycoproteins / Q000378:metabolism; D019251:Nucleocapsid / Q000378:metabolism; D064370:Spike Glycoprotein, Coronavirus; D013347:Subcellular Fractions; D014407:Tumor Cells, Cultured; D014759:Viral Envelope Proteins / Q000378:metabolism; D014760:Viral Fusion Proteins; D014763:Viral Matrix Proteins / Q000378:metabolism; D014764:Viral Proteins / Q000378:metabolism*; D019065:Virus Assembly
chemical_list: D000086862:Coronavirus M Proteins; D006023:Glycoproteins; D006389:Hemagglutinins, Viral; D008562:Membrane Glycoproteins; D064370:Spike Glycoprotein, Coronavirus; D014759:Viral Envelope Proteins; D014760:Viral Fusion Proteins; D014763:Viral Matrix Proteins; D014764:Viral Proteins; C060814:hemagglutinin esterase; C578557:spike glycoprotein, SARS-CoV; C578553:spike protein, mouse hepatitis virus