New Paper (Other): Structure, Function, and Antigenicity of the SARS-CoV-2 Spike Glycoprotein

[juliettemarie0405]

Title: Structure, Function, and Antigenicity of the SARS-CoV-2 Spike Glycoprotein

General Information

Please paste a link to the paper or a citation here:

Link: https://www.ncbi.nlm.nih.gov/pubmed/32155444

What is the paper's Manubot-style citation?

Citation: @doi:10.1016/j.cell.2020.02.058

Is this paper primarily relevant to Background or Pathogesis?

• [x] Background
• [x] Pathogenesis
• [ ] Methods

Please list some keywords (3-10) that help identify the relevance of this paper to COVID-19

• cryo-EM
• neutralizing antibodies
• ACE2
• spike glycoprotein
• furin cleavage site

Which areas of expertise are particularly relevant to the paper?

• [x] virology
• [ ] epidemiology
• [ ] biostatistics
• [x] immunology
• [ ] pharmacology
• [ ] other:

[juliettemarie0405]

Summary

Suggested questions to answer about each paper:

What did they analyze?

They compared structural and sequence conservation between SARS-CoV and SARS-CoV-2 spike glycoproteins (S), examined the functional effects of identified sequence variation (i.e. a furin cleavage site at the S1/S2 boundary identified in SARS-CoV-2 S but absent in SARS-CoV), and tested the ability of polyclonal sera from SARS-CoV-infected mice to cross-neutralize SARS-CoV-2 S-MLV pseudovirus. They also tested the ability of SARS-CoV-2 S-MLV pseudovirus to use the ACE2 receptor for cell entry and the affinity of human ACE2 for SARS-CoV-2 S.

What methods did they use?

• Pseudovirus entry assay (to determine ACE2 is functional receptor; to test impact of furin cleavage site on cell entry; to test ability of SARS-CoV mouse polyclonal sera to inhibit SARS-CoV-2 S-MLV pseudovirus cell entry)

• Western blot (to determine impact of furin cleavage site on spike glycoprotein cleavage)

• Biolayer interferometry (to study binding kinetics and affinity of human ACE2 to SARS-CoV-2 S)

• Cryo-EM (to examine the structure of SARS-CoV-2 S ectodomain trimer and its open and closed conformations)

Does this paper study COVID-19, SARS-CoV-2, or a related disease and/or virus?

SARS-CoV-2 in relation to SARS-CoV and other SARS-related CoVs

What is the main finding (or a few main takeaways)?

• SARS-CoV-2 uses human ACE2 receptors to enter target cells, with the receptor-binding domain demonstrating similar binding affinity to that of SARS-CoV

• SARS-CoV-2 contains a furin cleavage site at the S1/S2 boundary (which is not found in SARS-CoV and SARS-related CoVs). Mutation of the furin cleavage site in SARS-CoV-2 caused S to remain uncleaved upon budding (like SARS-Cov), but did not abolish S-mediated cell entry into VeroE6 and hACE2-expressing BHK cells.

• SARS-CoV murine polyclonal plasma potently inhibited SARS-CoV-2 S-MLV pseudovirus entry into target cells up to 80-90%.

• Based on cryo-EM structures of SARS-CoV-2 S ectodomain trimer, they identified a closed conformation (where all three SB domains of the S1 apex are buried) and an open conformation (where one of three SB domains of the S1 apex is open). Previous SARS-CoV and MERS-CoV data have demonstrated similar conformational variability of the SB domains. From comparing their cryo-EM structures, they concluded that the SARS-CoV-2 and SARS-CoV S2 subunits are structurally conserved.

What does this paper tell us about the background and/or diagnostics/therapeutics for COVID-19 / SARS-CoV-2?

• The authors report that the polybasic cleavage site (which is present in SARS-CoV-2 but not SARS-CoV or other SARS-related CoVs) is a signature of highly pathogenic avian influenza viruses and pathogenic Newcastle disease virus. They suggest that because furin-like proteases are ubiquitously expressed, the presence of this cleavage site could expand cell and tissue tropism of SARS-CoV-2 beyond that of SARS-CoV (as well as contribute to increased transmissibility and pathogenicity).

• The fact that the S2 subunit is more conserved than the S1 subunit across SARS-related CoVs suggests that vaccines eliciting neutralizing antibody responses against the S2 subunit could not only protect against SARS-CoV-2, but also other SARS-related coronaviruses that currently infect humans or may one day jump to humans from animal reservoirs.

• Polyclonal sera from SARS-CoV-infected mice was able to potently neutralize SARS-CoV-2 S-MLV pseudovirus, suggesting that previously isolated anti-SARS-CoV antibodies could be used in passive immunization. However, the authors note that previous research shows that several anti-SARS-CoV antibodies targeting the SB domain of the S1 subunit do not cross-neutralize SARS-CoV-2, and that most SARS-CoV-directed antibodies isolated to date target the SB domain…

Do you have any concerns about methodology or the interpretation of these results beyond this analysis?

Overall no, though I have no experience with cryo-EM or biolayer interferometry.

I did notice that they simply added 20 ul of pseudovirus stock to their cells in the pseudovirus entry assays instead of titering the stocks to standardize viral input. I'm wondering how we can compare the RLU values of the different pseudovirus stocks in figure 1 if the viral input was not standardized?

Any comments or notes?

I don't have a PhD, so I would appreciate someone else reviewing this paper too!