Exploring the evolution of the SARS-CoV-2 spike in relation to susceptibility to antiviral restriction factors
In a recent study published on bioRxiv* preprint server, researchers investigated the effects of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) spike (S) mutations on interferon-induced transmembrane protein-mediated antiviral restriction ( IFITM) and guanylate-binding proteins (GBP).
Interferon-stimulated genes (ISGs), components of innate immunity, encode antiviral restriction factors that act directly as the first line of defense against viruses, including SARS-CoV-2. GBP2/5 detects an activated interferon response due to SARS-CoV-2 attack in human airway epithelial cells and collectively targets key steps of viral replication to induce an antiviral state; likewise, the IFITM proteins largely act to block the entry of SARS-CoV-2.
Curiously, mutations within the furin cleavage site (FCS) of protein S affect these antiviral restriction factors. Therefore, studying the effect of GBP- and IFITM-mediated restriction is crucial to better understand how dominant SARS-CoV-2 variants of concern (VOCs) evolve their FCS by acquiring mutations to improve their infectivity and transmission potential.
About the study
In the current study, researchers investigated whether antiviral restriction factors, such as GBP1/5 and IFITM, could inhibit S-cleavage resulting in reduced SARS-CoV-2 infectivity. They also tested whether S mutations provided SARS-CoV-2 COVs with the ability to escape this restriction; in other words, they assessed the effect of S mutations on sensitivity to endosomal restriction factors, including IFITMs.
Analysis of the study showed that interferon-inducible restriction factors GBP2 and GBP5 interfered with furin cleavage of Wuhan-Hu-1, Alpha, Delta and Omicron S proteins, similar to inhibition furin by GBP. Additionally, exposure to GBP shifted the viral entry pathway to endosomal entry. On the other hand, IFITM1, but not IFITM 2 or 3 (mainly located in endosomes) inhibited infection by early lineage Wuhan-Hu-1 of SARS-CoV-2, as well as COV Alpha and Delta.
In addition, they observed differential sensitivity of COV S proteins to GBP and IFITM restriction, as restriction by GBP2/5 correlated with differential furin-mediated S processing requirement. Therefore, Middle East respiratory syndrome coronavirus (MERS-CoV) S was sensitive, but SARS-CoV-1 was resistant to inhibition by GBP.
Omicron emerged as a unique COV in that it is sensitive to inhibition by GBP2/5 and IFITM1, 2 and 3 and evolves into the transmembrane protease-independent entry pathway 2 (TMPRSS2). It showed less sensitivity to the TMPRSS2 inhibitor Camostat and more sensitivity to the cathepsin inhibitor E64d. Moreover, it was significantly less infectious than other SARS-CoV-2 isolates on Caco2 cells expressing TMPRSS2.
Additionally, Omicron contains the same P681H FCS-optimizing mutation as Alpha; furthermore, it has an increased affinity for ACE2, just like Alpha and Delta. Thus, the researchers hypothesized that Omicron would be resistant to GBP in pseudovirus (PV) testing, like other VOCs. However, Omicron was sensitive to GBP2/5, behaving like Wuhan-Hu-1. Moreover, increasing Omicron S incorporation into PVs did not rescue them from GBP or IFITM restriction.
The study data revealed that the evolution of Alpha and Delta S also contributed to conferring resistance to GBP restriction, and this was not solely due to the acquisition of enhanced FCS.
Discussion and conclusions
Taken together, the study data revealed that SARS-CoV-2 evolves to balance efficient host cell entry with evasion of compartmentalized restriction factors. Thus, Omicron remarkably adapted its S activity, became less fusogenic and is sensitive to restriction by GBP2/5 and IFITM1/2/3, which interfere with viral fusion and cell entry.
Additionally, compared to Wuhan-Hu-1, Omicron contains three unique mutations (Q954H, N969K, and L981F) in the heptad repeat domain 1 (HR1) of its S that mediates viral fusion. As these substitutions are absent in COV Alpha and Delta, they may also contribute significantly to the differences between Omicron and the other COVs in the context of S fusion, restriction sensitivity and tropism.
Overall, mutations in and around the receptor binding domain (RBD), N-terminal domain (NTD), and S1/S2 cleavage boundary of Omicron S could contribute to the distinctiveness of the Omicron phenotype.
According to the authors, similar processes occurred during the evolution of SARS-CoV-2 in hosts and therefore the need to evade neutralizing antibodies became the dominant selective pressure on Omicron. This interplay between innate and adaptive immunity evasion, and consequences for transmission and tropism, will continue to influence the future evolution of SARS-CoV-2. Therefore, linking this evolution to the phenotype could be crucial for developing insights into understanding the biology and pathogenesis of SARS-CoV-2.
bioRxiv publishes preliminary scientific reports that are not peer-reviewed and, therefore, should not be considered conclusive, guide clinical practice/health-related behaviors, or treated as established information.