BREAKING! King’s College London Discover That P681H Mutation In Spike Protein Confers Type I Interferon Resistance In SARS-CoV-2 Alpha Variant

Scientists from Department of Infectious Diseases, King’s College London along with other experts from the MRC-University of Glasgow Centre for Virus Research and the Guy’s and St Thomas’ NHS Foundation Trust-UK have in a new study discovered that the P681H mutation in spike protein confers type 1 interferon resistance in SARS-CoV-2 Alpha variant.

 
VOCs or variants of concern (VOCs) of the SARS-CoV-2 coronavirus continue threaten the global response to the COVID-19 pandemic with many developing a variety of immune evasive modes.
 
The alpha (B.1.1.7) variant appeared in the UK became dominant in Europe and North America in early 2021. The Spike glycoprotein of alpha has acquired a number of mutations including the P681H mutation in the polybasic cleavage site that has been suggested to enhance Spike cleavage.
 
The study findings however the alpha Spike protein also confers a level of resistance to the effects of interferon-β (IFNβ) in lung epithelial cells. This correlates with resistance to restriction mediated by interferon-induced transmembrane protein-2 (IFITM2) and a pronounced infection enhancement by IFITM3.
 
Furthermore, the P681H mutation is necessary for comparative resistance to IFNβ in a molecularly cloned SARS-CoV-2 encoding alpha Spike.
 
The study findings suggest that in addition to adaptive immune escape, mutations associated with VOCs also confer replication advantage through adaptation to resist innate immunity.
 
The study findings were published on a preprint server and are currently being peer reviewed. https://www.biorxiv.org/content/10.1101/2021.11.09.467693v1
 
Past studies have already suggested that variants of SARS-CoV-2 have begun to evolve further resistance to interferon-induced innate immunity
https://pubmed.ncbi.nlm.nih.gov/33758840/
 
https://pubmed.ncbi.nlm.nih.gov/34127972/
 
In the first linked study above, viral isolates over the pandemic show a reduced sensitivity to type I interferons in culture; in a second the alpha variant has a significantly reduced propensity to trigger pattern recognition in epithelial cells.
 
However, in contrast, another study shows no difference in IFN sensitivity of the new variants in African green monkey Vero-E6 cells, although species-specificity in viral sensitivity to ISGs (Interferon-stimulated gene) is a well characterized trait. https://pubmed.ncbi.nlm.nih.gov/34601723/
 
The SARS-CoV-2 genome contains multiple mechanisms to counteract host innate immune responses, and much remains to be learned about the mechanisms deployed by this virus and its relatives. While many reports on SARS-CoV-2 evolution have naturally focused on the pressing concern of potential for vaccine escape, it is very unlikely that all selective adaptations that we see arising in VOCs can be solely due to escape from adaptive immunity.
 
The alpha varia nt Spike, for example, only displays a minor reduction in sensitivity to neutralizing antibodies (NAbs)
https://pubmed.ncbi.nlm.nih.gov/33526412/
 
https://pubmed.ncbi.nlm.nih.gov/34237773/
 
However, this VOC had a considerable transmission advantage, with suspicions that it may have arisen in an immunocompromised individual with a persistent infection giving ample time for changes to be selected that further evade innate immunity, including those that target viral entry.
 
In terms of IFITM (Interferon-induced transmembrane) resistance of VOC Spike proteins, so far we have only seen marked change in phenotype for the alpha variant. This is despite the fact that delta, which has superseded alpha in many places around the world in 2021, also shows an adaptation for enhanced S1/S2 cleavage with an P681R change.
https://pubmed.ncbi.nlm.nih.gov/34462752/
 
https://www.biorxiv.org/content/10.1101/2021.05.28.446163v1
 
This would suggest that efficient cleavage of S1/S2 is necessary but not sufficient for IFITM resistance, and indeed the studt data implicate a context dependency of the NTD deletion at 69/70. While unique to the alpha VOC, the 69/70 deletion has been observed in persistent infection of immunosuppressed individuals and is thought to enhance viral fitness and Spike stability. https://pubmed.ncbi.nlm.nih.gov/34166617/
 
Although deletions in the NTD do affect NAb binding, it is primarily the 144 deletion in the alpha Spike that escapes neutralization by NTD-directed Nabs and the study findings show that this has no impact on IFITM sensitivity. https://pubmed.ncbi.nlm.nih.gov/32571838/
 
However, by contrast, the more pronounced antibody evasion by the beta, gamma and delta variants is related to mutations in the major neutralizing epitopes of the RBD, suggesting that they may well have been driven by antibody escape. https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7901269/
 
The viral glycoproteins are dynamic structures that shift through large-scale conformational changes while interacting with their cognate receptors mediating viral membrane fusion. Such context dependency is therefore likely to be complex and will arise under competing selective pressures.
 
Already it has been previously shown that the HIV-1 envelope glycoprotein of transmitted viruses is IFITM insensitive and this contributes to their overall type I IFN resistance. https://pubmed.ncbi.nlm.nih.gov/27640936/
 
Interestingly as HIV-1 infection progresses over the first 6 months in an infected person, the circulating variants increase in IFN/IFITM sensitivity and this is determined by adaptive changes in Env that resist the early neutralizing antibody response. https://retrovirology.biomedcentral.com/articles/10.1186/1742-4690-10-146
 
Such escape has structural and functional implications for such dynamic proteins that may impact upon receptor interactions and route of entry into the target cell.
 
While furin cleavage of the SARS-CoV-2 Spike reduces its IFITM sensitivity, other interferon-induced proteins may contribute to this phenotype. The guanylate binding protein family, and particularly GBP2 and GBP5, have been shown to have a general antiviral activity against enveloped viruses by dysregulating furin processing of diverse viral and cellular proteins.
https://pubmed.ncbi.nlm.nih.gov/31091448/
 
In the same way, IFITM overexpression in HIV-infected cells can lead to their incorporation into virions and in some cases promote defects in glycoprotein incorporation.
https://retrovirology.biomedcentral.com/articles/10.1186/s12977-014-0103-y
 
Future studies will confirm whether either of these mechanisms are involved in the IFN-resistance associated with the P681H mutation in alpha.
 
In summary, the spike protein of SARS-CoV-2 alpha increases resistance to IFN-I and this correlates with the P681H mutation.
 
Furthermore, this correlates with resistance to IFITM-mediated entry restriction. This suggests that in addition to adaptive immune escape, fixed mutations associated with VOCs may well also confer replication and/or transmission advantage through adaptation to resist innate immune mechanisms.
 
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