Exploitation of the Secretory Pathway by SARS-CoV-2 Envelope

Pearson, Guy J; Broncel, Malgorzata; Snijders, Ambrosius P; Carlton, Jeremy G

The beta-coronavirus SARS-CoV-2 is the causative agent of the current global COVID-19 pandemic. Coronaviruses are enveloped RNA viruses. Assembly and budding of coronavirus particles occur at the Endoplasmic Reticulum-Golgi Intermediate Compartment (ERGIC), with the structural proteins Nucleocapsid, Spike, Membrane and Envelope facilitating budding and release of virions into the secretory pathway lumen. This allows viral release which can occur through delivery of virus particles to deacidified lysosomes and subsequent lysosomal secretion. Coronaviral Envelope proteins are necessary for coronavirus assembly, play important roles in replication and can form oligomeric cation channels. Whilst synthesised in the ER, the mechanism by which Envelope achieves its steady state localisation to the ERGIC remains unclear. Here, we used fluorescent reporters to illuminate the Envelope protein from SARS-CoV-2. We discovered that internal tagging of this protein is necessary to preserve the functionality of a C-terminal ER-export motif and to allow localisation of Envelope to the ERGIC. Using this non-disruptive form of tagging, we used proximity biotinylation to define the vicinal proteome of wild type and ER-restricted versions of Envelope. We show that both Envelope and the presence of its ER-export motif contribute to the packaging of nucleocapsid into virus like particles. Finally, using our labelled versions of Envelope, we discovered that a minor pool of this protein is delivered to lysosomes. We show that lysosomal Envelope is oligomeric and can contribute to pH neutralisation in these organelles.