Structural basis for cell-type specific evolution of viral fitness by SARS-CoV-2

Gupta, Kapil; Toelzer, Christine; Williamson, Maia Kavanagh; Shoemark, Deborah K.; Oliveira, A. Sofia F.; Matthews, David A.; Almuqrin, Abdulaziz; Staufer, Oskar; Yadav, Sathish K. N.; Borucu, Ufuk; Garzoni, Frédéric; Fitzgerald, Daniel J.; Spatz, Joachim P.; Mulholland, Anthony J.; Davidson, Andrew D.; Schaffitzel, Christiane; Berger, Imre

doi:10.1101/2021.05.11.443384

Cited by 5 publications

(8 citation statements)

References 79 publications

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“…Here, three equilibrium 200 ns MD simulations were performed for the locked form of the unglycosylated and uncleaved ectodomain region of the D614G spike modelled from a wild-type model (17,20) based on the cryo-EM structure 6ZB5 (17) with and without LA bound (see Supplementary Material). In the original wild-type spike, D614 is located at the interface between monomers, with its sidechain directly interacting with residues across the subunit interface (55). RMSF profiles for the wild-type and D614G apo spikes are similar (Figure S15).…”

Section: Dynamic Response Of the D614g Spikementioning

confidence: 97%

“…This site contains a polybasic PRRA insertion not found in other SARS-CoV-related coronaviruses (52). Cellbased assays show that deletion of the PRRA motif affects virus infectivity (19,(52)(53)(54)(55)(56). Note that in the simulations presented here, the furin site (located between R685 and S686) is not cleaved (see Supplementary Material).…”

Section: Fa Sites Modulate Key Motifs For Membrane Fusion or Antigenic Epitopes In The Spikementioning

confidence: 99%

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The fatty acid site is coupled to functional motifs in the SARS-CoV-2 spike protein and modulates spike allosteric behaviour

Oliveira

Shoemark

Ibarra

et al. 2021

Preprint

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The SARS-CoV-2 spike protein is the first contact point between the SARS-CoV-2 virus and host cells and mediates membrane fusion. Recently, a fatty acid binding site was identified in the spike (Toelzer et al. Science 2020). The presence of linoleic acid at this site modulates binding of the spike to the human ACE2 receptor, stabilizing a locked conformation of the protein. Here, dynamical-nonequilibrium molecular dynamics simulations reveal that this fatty acid site is coupled to functionally relevant regions of the spike, some of them far from the fatty acid binding pocket. Removal of a ligand from the fatty acid binding site significantly affects the dynamics of distant, functionally important regions of the spike, including the receptor-binding motif, furin cleavage site and fusion-peptide-adjacent regions. The results also show significant differences in behaviour between clinical variants of the spike: e.g. the D614G mutation shows a significantly different conformational response for some structural motifs relevant for binding and fusion. The simulations identify structural networks through which changes at the fatty acid binding site are transmitted within the protein. These communication networks significantly involve positions that are prone to mutation, indicating that observed genetic variation in the spike may alter its response to linoleate binding and associated allosteric communication.

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Section: Dynamic Response Of the D614g Spikementioning

confidence: 97%

Section: Fa Sites Modulate Key Motifs For Membrane Fusion or Antigenic Epitopes In The Spikementioning

confidence: 99%

The fatty acid site is coupled to functional motifs in the SARS-CoV-2 spike protein and modulates spike allosteric behaviour

Oliveira

Shoemark

Ibarra

et al. 2021

Preprint

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“…Comparison with published fatty acid-protein structures suggested that the length and shape of the density agreed with linoleic acid (LA), an unsaturated C 18 fatty acid with two double bonds. Using hydrophobic interaction liquid chromatography-electrospray ionization mass spectrometry (HILIC-MS) and LC-MS-MS, the ligand in the RBD was confirmed to be LA (Toelzer et al, 2020;Gupta et al, 2022). Notably, only LA and no other fatty acids were identified by mass spectrometry (Toelzer et al, 2020).…”

Section: Cryo-em Of Sars-cov-2 Spike Glycoproteinmentioning

confidence: 99%

“…Deletion or mutation of the furin-cleavage site interferes with viral infection of lung cells and primary human airway epithelial cells (Peacock et al, 2021;Johnson et al, 2021). However, in patients infected with SARS-CoV-2, spikeprotein variants with a deleted furin-cleavage site have been identified as a subpopulation allowing the virus to infect other cell types, including epithelial cells (Peacock et al, 2021;Gupta et al, 2022;Pohl et al, 2021;Ogando et al, 2020;Sasaki et al, 2021;Klimstra et al, 2020). Interestingly, more recent SARS-CoV-2 variants of concern (VOCs) have acquired even more basic cleavage sites (HRRAR in Alpha and Omicron spike proteins and RRRAR in Delta spike proteins), resulting in a more complete cleavage at the furin site, contributing to the increased transmissibility of these VOCs (Wrobel et al, 2022).…”

Section: Introductionmentioning

confidence: 99%

Cryo-EM reveals binding of linoleic acid to SARS-CoV-2 spike glycoprotein, suggesting an antiviral treatment strategy

Toelzer¹,

Gupta²,

Berger³

et al. 2023

Acta Cryst Sect D Struct Biol

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The COVID-19 pandemic and concomitant lockdowns presented a global health challenge and triggered unprecedented research efforts to elucidate the molecular mechanisms and pathogenicity of SARS-CoV-2. The spike glycoprotein decorating the surface of SARS-CoV-2 virions is a prime target for vaccine development, antibody therapy and serology as it binds the host cell receptor and is central for viral cell entry. The electron cryo-microscopy structure of the spike protein revealed a hydrophobic pocket in the receptor-binding domain that is occupied by an essential fatty acid, linoleic acid (LA). The LA-bound spike protein adopts a non-infectious locked conformation which is more stable than the infectious form and shields important immunogenic epitopes. Here, the impact of LA binding on viral infectivity and replication, and the evolutionary conservation of the pocket in other highly pathogenic coronaviruses, including SARS-CoV-2 variants of concern (VOCs), are reviewed. The importance of LA metabolic products, the eicosanoids, in regulating the human immune response and inflammation is highlighted. Lipid and fatty-acid binding to a hydrophobic pocket in proteins on the virion surface appears to be a broader strategy employed by viruses, including picornaviruses and Zika virus. Ligand binding stabilizes their protein structure and assembly, and downregulates infectivity. In the case of rhinoviruses, this has been exploited to develop small-molecule antiviral drugs that bind to the hydrophobic pocket. The results suggest a COVID-19 antiviral treatment based on the LA-binding pocket.

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“…Structural studies have revealed that cleavage at the polybasic furin-cleavage site decreases the overall stability of SARS-CoV-2 Spike and facilitates the adoption of the open conformation that is required for Spike to bind the ACE2 receptor [27]. Consistent with this, deletion of the furin cleavage site in SARS-CoV-2 Spike promotes the adoption of closed form of the Spike and prevents cell-cell as well as virus-cell fusion in some cell types [16,26,28,29].…”

Section: Introductionmentioning

confidence: 88%

TMPRSS2 promotes SARS-CoV-2 evasion from NCOA7-mediated restriction

et al. 2021

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Interferons play a critical role in regulating host immune responses to SARS-CoV-2, but the interferon (IFN)-stimulated gene (ISG) effectors that inhibit SARS-CoV-2 are not well characterized. The IFN-inducible short isoform of human nuclear receptor coactivator 7 (NCOA7) inhibits endocytic virus entry, interacts with the vacuolar ATPase, and promotes endo-lysosomal vesicle acidification and lysosomal protease activity. Here, we used ectopic expression and gene knockout to demonstrate that NCOA7 inhibits infection by SARS-CoV-2 as well as by lentivirus particles pseudotyped with SARS-CoV-2 Spike in lung epithelial cells. Infection with the highly pathogenic, SARS-CoV-1 and MERS-CoV, or seasonal, HCoV-229E and HCoV-NL63, coronavirus Spike-pseudotyped viruses was also inhibited by NCOA7. Importantly, either overexpression of TMPRSS2, which promotes plasma membrane fusion versus endosomal fusion of SARS-CoV-2, or removal of Spike’s polybasic furin cleavage site rendered SARS-CoV-2 less sensitive to NCOA7 restriction. Collectively, our data indicate that furin cleavage sensitizes SARS-CoV-2 Spike to the antiviral consequences of endosomal acidification by NCOA7, and suggest that the acquisition of furin cleavage may have favoured the co-option of cell surface TMPRSS proteases as a strategy to evade the suppressive effects of IFN-induced endo-lysosomal dysregulation on virus infection.

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Structural basis for cell-type specific evolution of viral fitness by SARS-CoV-2

Cited by 5 publications

References 79 publications

The fatty acid site is coupled to functional motifs in the SARS-CoV-2 spike protein and modulates spike allosteric behaviour

The fatty acid site is coupled to functional motifs in the SARS-CoV-2 spike protein and modulates spike allosteric behaviour

Cryo-EM reveals binding of linoleic acid to SARS-CoV-2 spike glycoprotein, suggesting an antiviral treatment strategy

TMPRSS2 promotes SARS-CoV-2 evasion from NCOA7-mediated restriction

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