Hannah has a new paper in mBio! Check it out HERE.
Coronaviruses encode 4 structural proteins: Spike (S), Envelope (E), Membrane (M), and Nucleocapsid (N), which are all assembled into new virions in the endoplasmic reticulum/Golgi intermediate compartment (ERGIC) during infection of cells.
Envelope (E) is the smallest structural protein of coronaviruses (75 amino acids for SARS-CoV-2). The effect of E gene deletion varies greatly between different coronaviruses. Hannah showed in this paper that SARS-CoV-2 is only attenuated when E is deleted.
Hannah could have stopped there, but she wanted to understand how SARS-CoV-2 could still replicate without the structural protein E!
Hannah deleted the E gene in SARS-CoV-2 without disrupting expression of the other viral genes. ΔE SARS-CoV-2 could replicate 100 - 10,000-fold lower than WT virus and its virus production was dramatically reduced. She could rescue ΔE SARS-CoV-2 infection by infecting cells stably expressing inducible, codon-optimized E without causing recombination in trans-complemented SARS-CoV-2 (ΔE + E).
ΔE SARS-CoV-2 had tiny plaques and lacked Spike trimers compared to WT virus, leading Hannah to investigate this further. Using several assays, she showed that ΔE SARS-CoV-2 could only replicate by cell-to-cell transmission and not by cell-free transmission. ΔE SARS-CoV-2 wasn’t better at cell fusion compared to WT virus, but that was the only way it could infect other cells.
Hannah then examined Spike expression inside infected cells vs. on the cell surface. ΔE SARS-CoV-2 had less retention of S in the ERGIC compared to WT SARS-CoV-2, which is where new viruses are assembled. Without E, more S was localized on the cell surface, allowing cell fusion to occur.
Unlike S, M protein localization was not changed by deletion of E. But surprisingly, ΔE SARS-CoV-2 infection led to more dramatic reorganization of the ERGIC in cells compared to WT SARS-CoV-2 infection. Priya confirmed this by electron microscopy.
Future work will need to examine how E affects Golgi-mediated trafficking during coronavirus infection and how a lack of E expression during SARS-CoV-2 infection affects the Golgi so dramatically.
We are excited that the story is finally out and even more excited for Hannah's second SARS-CoV-2 project. Stay tuned!
Thanks to Priya and Chris in the lab for their experimental contributions and thank you to our collaborators Drs. James Conway, Simon Watkins, Paul Duprex, and Kevin McCarthey for additional experiments, expertise/advice, and reagents!
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