Abstract
Coronaviruses are characterized by their progeny assembly and budding in the endoplasmic reticulum-Golgi intermediate compartment (ERGIC). Our previous studies demonstrated that truncation of 9 amino acids in the cytoplasmic tail (CT) of the infectious bronchitis virus (IBV) spike (S) protein impairs its localization to the ERGIC, resulting in increased expression at the plasma membrane. However, the precise mechanism underlying this phenomenon remained elusive. In this study, we provide evidence that the IBV S protein could utilize coatomer protein-I (COPI)-coated vesicles for retrograde transport from the Golgi to the endoplasmic reticulum (ER). We identified the KKSV motif as the critical binding site within the CT domain of IBV S protein for COPI interaction. Further analysis reveals that IBV infection does not modulate host COPI expression. However, when COPI expression is disrupted, a higher proportion of S protein escapes to the plasma membrane. Moreover, inhibition of COPI-mediated transport during viral infection severely impairs progeny virion production and leads to increased S protein accumulation at the plasma membrane, inducing cell-cell fusion and syncytia formation. Our findings contribute to a deeper understanding of S protein intracellular trafficking during coronavirus infection, and offer valuable insights into the molecular mechanisms of viral replication and host cell biology.IMPORTANCEViruses hijack or modify host cellular machinery and associated pathways to facilitate their own replication. Here, we demonstrate that the infectious bronchitis virus (IBV) S protein directly interacts with coatomer protein-I (COPI)-coated vesicles through the KKSV motif in its cytoplasmic tail. COPI-coated vesicles mediate the retrograde transport of S protein from the Golgi apparatus to the endoplasmic reticulum-Golgi intermediate compartment, where viral particle assembly occurs. Our findings not only advance our understanding of IBV S protein trafficking mechanisms but also provide valuable insights for developing more effective vaccine strategies.