A single amino acid mutation in the spike protein of coronavirus infectious bronchitis virus hampers its maturation and incorporation into virions at the nonpermissive temperature

• Published in: Virology (New York, N.Y.) Vol. 326; no. 2; pp. 288 - 298
• Main Authors: Shen, S, Law, Y.C, Liu, D.X
• Format: Journal Article
• Language: English
• Published: United States Elsevier Inc 01.09.2004
• Subjects: Acclimatization; Amino Acid Substitution; Animals; cell lines; Chlorocebus aethiops; Coronavirus; Cytoplasm - metabolism; Glutamine - chemistry; Glycoprotein; glycoproteins; Glycoside Hydrolases; HeLa Cells; Humans; Infectious bronchitis virus; Infectious bronchitis virus - genetics; Infectious bronchitis virus - physiology; Leucine - chemistry; Membrane Glycoproteins - biosynthesis; Membrane Glycoproteins - genetics; Membrane Glycoproteins - physiology; monkeys; Point Mutation; Spike Glycoprotein, Coronavirus; Temperature; Transfection; Vero Cells; Viral Envelope Proteins - biosynthesis; Viral Envelope Proteins - genetics; Viral Envelope Proteins - physiology; Virion; Virus Assembly; Virion; Coronavirus; Glycoprotein
• ISSN: 0042-6822; 1096-0341
• DOI: 10.1016/j.virol.2004.06.016

The spike (S) glycoprotein of coronavirus is responsible for receptor binding and membrane fusion. A number of variants with deletions and mutations in the S protein have been isolated from naturally and persistently infected animals and tissue cultures. Here, we report the emergence and isolation of two temperature sensitive (ts) mutants and a revertant in the process of cold-adaptation of coronavirus infectious bronchitis virus (IBV) to a monkey kidney cell line. The complete sequences of wild type (wt) virus, two ts mutants, and the revertant were compared and variations linked to phenotypes were mapped. A single amino acid reversion (L 294-to-Q) in the S protein is sufficient to abrogate the ts phenotype. Interestingly, unlike wt virus, the revertant grows well at and below 32 °C, the permissive temperature, as it carries other mutations in multiple genes that might be associated with the cold-adaptation phenotype. If the two ts mutants were allowed to enter cells at 32 °C, the S protein was synthesized, core-glycosylated and at least partially modified at 40 °C. However, compared with wt virus and the revertant, no infectious particles of these ts mutants were assembled and released from the ts mutant-infected cells at 40 °C. Evidence presented demonstrated that the Q 294-to-L 294 mutation, located at a highly conserved domain of the S1 subunit, might hamper processing of the S protein to a matured 180-kDa, endo-glycosidase H-resistant glycoprotein and the translocation of the protein to the cell surface. Consequently, some essential functions of the S protein, including mediation of cell-to-cell fusion and its incorporation into virions, were completely abolished.