The coronavirus envelope protein has multiple roles in virus assembly and trafficking

Coronaviruses (CoVs) are enveloped viruses with large positive-sense single stranded genomes. CoVs infect a variety of mammalian and avian species, and can cause serious disease in humans, as exemplified during the 2002-2003 outbreak of severe acute respiratory syndrome. Unlike many other enveloped viruses, CoVs assemble intracellularly by budding into intermediate compartment between the endoplasmic reticulum and the Golgi complex. Virions must then traverse the host secretory pathway for egress. The advantage of intracellular assembly is not clear, nor is it known if CoVs need to alter the host secretory pathway for efficient exocytosis of virions. The CoV E protein is a small membrane protein that is a minor component of the virion envelope. CoV E is important for the assembly of virions, but may have other roles in infection. CoV E has ion channel activity in vitro, although a role for this in infection has not been demonstrated. The role for the ion channel activity was investigated by developing a recombinant version of the model CoV infectious bronchitis virus (IBV) carrying a mutant version of E where the hydrophobic domain was replaced with a heterologous sequence (IBV-EG3). A defect in the release of infectious IBV-EG3 particles was observed. This defect is likely due to the improper trafficking of virions. When the E and EG3 proteins were transiently expressed, the wild-type protein caused a dramatic alteration of the secretory pathway, including the disassembly of the Golgi complex and reduction in protein cargo trafficking, whereas EG3 had no effect. Using site directed mutagenesis, it was determined that a single polar uncharged residue within the hydrophobic domain of IBV E correlated with the disruption of the secretory pathway. We speculate that the ion channel activity of IBV E may disrupt the secretory pathway, while at the same time creating an environment that facilitates the release of infectious particles.;CoV E may adopt a transmembrane or membrane hairpin topology, each with distinct functions. To investigate how multiple topologies contributed to the function of IBV E, versions of the protein were developed that adopted either a transmembrane or membrane hairpin conformation. A transmembrane topology was necessary for disrupting the secretory pathway, but was not as efficient at assembly compared to the wild-type protein in a virus-like particle assay. The membrane hairpin was unable to disrupt the secretory pathway, or support virus like particle release. We speculate that a small portion of the wild-type protein may exist in the hairpin conformation to promote assembly, while the transmembrane conformation promotes release.;The results presented here expand the role of IBV E beyond assembly, and demonstrate that IBV E is necessary for the efficient trafficking of virions. Furthermore, this activity is correlated with a single polar residue in the hydrophobic domain of the protein, and requires a transmembrane topology.