The altered entry pathway and antigenic distance of the SARS-CoV-2 Omicron variant map to separate domains of spike protein
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Abstract
The SARS-CoV-2 Omicron/BA.1 lineage emerged in late 2021 and rapidly displaced the Delta variant before being overtaken itself globally by, the Omicron/BA.2 lineage in early 2022. Here, we describe how Omicron BA.1 and BA.2 show a lower severity phenotype in a hamster model of pathogenicity which maps specifically to the spike gene. We further show that Omicron is attenuated in a lung cell line but replicates more rapidly, albeit to lower peak titres, in human primary nasal cells. This replication phenotype also maps to the spike gene. Omicron spike (including the emerging Omicron lineage BA.4) shows attenuated fusogenicity and a preference for cell entry via the endosomal route. We map the altered Omicron spike entry route and partially map the lower fusogenicity to the S2 domain, particularly the substitution N969K. Finally, we show that pseudovirus with Omicron spike, engineered in the S2 domain to confer a more Delta-like cell entry route retains the antigenic properties of Omicron. This shows a distinct separation between the genetic determinants of these two key Omicron phenotypes, raising the concerning possibility that future variants with large antigenic distance from currently circulating and vaccine strains will not necessarily display the lower intrinsic severity seen during Omicron infection.
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SciScore for 10.1101/2021.12.31.474653: (What is this?)
Please note, not all rigor criteria are appropriate for all manuscripts.
Table 1: Rigor
Ethics not detected. Sex as a biological variable not detected. Randomization not detected. Blinding not detected. Power Analysis not detected. Cell Line Authentication not detected. Table 2: Resources
Antibodies Sentences Resources Near infra-red (NIR) secondary antibodies, IRDye® 680RD Goat anti-mouse (abcam; ab216776) and IRDye® 800CW Goat anti-rabbit (abcam; ab216773) were subsequently used to probe membranes. anti-mousesuggested: Noneanti-rabbitsuggested: NoneExperimental Models: Cell Lines Sentences Resources Delta and Omicron virus were isolated by inoculating 100 µL of neat swab material onto Vero E6-ACE2-TMPRSS2 cells, incubating at 37°C for 1 h before replacing with growth media supplemented with 1x … SciScore for 10.1101/2021.12.31.474653: (What is this?)
Please note, not all rigor criteria are appropriate for all manuscripts.
Table 1: Rigor
Ethics not detected. Sex as a biological variable not detected. Randomization not detected. Blinding not detected. Power Analysis not detected. Cell Line Authentication not detected. Table 2: Resources
Antibodies Sentences Resources Near infra-red (NIR) secondary antibodies, IRDye® 680RD Goat anti-mouse (abcam; ab216776) and IRDye® 800CW Goat anti-rabbit (abcam; ab216773) were subsequently used to probe membranes. anti-mousesuggested: Noneanti-rabbitsuggested: NoneExperimental Models: Cell Lines Sentences Resources Delta and Omicron virus were isolated by inoculating 100 µL of neat swab material onto Vero E6-ACE2-TMPRSS2 cells, incubating at 37°C for 1 h before replacing with growth media supplemented with 1x penicillin/streptomycin and 1x amphotericin B. E6-ACE2-TMPRSS2suggested: NoneIsolates were passaged a further two times in Vero E6-ACE2-TMPRSS2 cells12. Vero E6-ACE2-TMPRSS2suggested: NoneOverexpression experiments in 293T cells were performed by co-transfecting either pCAGGS-ACE2-FLAG or non-cleavable pCAGGs-ACE2-C4-FLAG, with or without TMPRSS2 into 100 mm dishes of 293T cells. 293Tsuggested: KCB Cat# KCB 200744YJ, RRID:CVCL_0063)Briefly, BHK cells were transfected with 500 ng of ACE2 or empty vector (pDISPLAY) using TransIT-X2 BHKsuggested: None293T, BHK-21, or Calu-3 target cells stably expressing rLuC-GFP-8-11 (target cells) were co-transfected with ACE2 expression constructs. BHK-21suggested: NoneFlow cytometry-based Spike-ACE2 affinity assay: HEK 293T cells were transfected with mammalian expression plasmids encoding the relevant SARS-CoV2 spike protein. HEK 293Tsuggested: KCB Cat# KCB 200744YJ, RRID:CVCL_0063)Recombinant DNA Sentences Resources Plasmids, pseudovirus and pseudovirus entry assays: Spike-encoding pcDNA3.1 plasmids were generated by mutagenesis or by gene synthesis as described elsewhere4,37. pcDNA3.1suggested: RRID:Addgene_79663)pCAGGs-ACE2-FLAG and non-cleavable pCAGGs-ACE2C4-FLAG were used a previously described13,38. pCAGGs-ACE2-FLAGsuggested: NonepCAGGs-ACE2C4-FLAGsuggested: NoneOverexpression experiments in 293T cells were performed by co-transfecting either pCAGGS-ACE2-FLAG or non-cleavable pCAGGs-ACE2-C4-FLAG, with or without TMPRSS2 into 100 mm dishes of 293T cells. pCAGGs-ACE2-C4-FLAGsuggested: NoneBriefly, BHK cells were transfected with 500 ng of ACE2 or empty vector (pDISPLAY) using TransIT-X2 pDISPLAYsuggested: RRID:Addgene_51053)Results from OddPub: We did not detect open data. We also did not detect open code. Researchers are encouraged to share open data when possible (see Nature blog).
Results from LimitationRecognizer: An explicit section about the limitations of the techniques employed in this study was not found. We encourage authors to address study limitations.Results from TrialIdentifier: No clinical trial numbers were referenced.
Results from Barzooka: We did not find any issues relating to the usage of bar graphs.
Results from JetFighter: We did not find any issues relating to colormaps.
Results from rtransparent:- Thank you for including a conflict of interest statement. Authors are encouraged to include this statement when submitting to a journal.
- Thank you for including a funding statement. Authors are encouraged to include this statement when submitting to a journal.
- No protocol registration statement was detected.
Results from scite Reference Check: We found no unreliable references.
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