Phosphatidylserine receptors enhance SARS-CoV-2 infection.

Dana Bohan,Tomasz Stokowy,James B Lorens,Natalie Ruggio,Wendy Maury,Eleni Christakou,Gro Gausdal,John Minna,José A Aguilar Briseño,Jonah M Elliff,Mohammad Badreddine,Hillel Haim,Hanora Van Ert,Roberth Anthony Rojas Chavez,Petri Kursula,David Micklem,Boning Gao,Kai J Rogers,Sean P.J Whelan

doi:10.1371/journal.ppat.1009743

Abstract

Phosphatidylserine (PS) receptors enhance infection of many enveloped viruses through virion-associated PS binding that is termed apoptotic mimicry. Here we show that this broadly shared uptake mechanism is utilized by SARS-CoV-2 in cells that express low surface levels of ACE2. Expression of members of the TIM (TIM-1 and TIM-4) and TAM (AXL) families of PS receptors enhance SARS-CoV-2 binding to cells, facilitate internalization of fluorescently-labeled virions and increase ACE2-dependent infection of SARS-CoV-2; however, PS receptors alone did not mediate infection. We were unable to detect direct interactions of the PS receptor AXL with purified SARS-CoV-2 spike, contrary to a previous report. Instead, our studies indicate that the PS receptors interact with PS on the surface of SARS-CoV-2 virions. In support of this, we demonstrate that: 1) significant quantities of PS are located on the outer leaflet of SARS-CoV-2 virions, 2) PS liposomes, but not phosphatidylcholine liposomes, reduced entry of VSV/Spike pseudovirions and 3) an established mutant of TIM-1 which does not bind to PS is unable to facilitate entry of SARS-CoV-2. As AXL is an abundant PS receptor on a number of airway lines, we evaluated small molecule inhibitors of AXL signaling such as bemcentinib for their ability to inhibit SARS-CoV-2 infection. Bemcentinib robustly inhibited virus infection of Vero E6 cells as well as multiple human lung cell lines that expressed AXL. This inhibition correlated well with inhibitors that block endosomal acidification and cathepsin activity, consistent with AXL-mediated uptake of SARS-CoV-2 into the endosomal compartment. We extended our observations to the related betacoronavirus mouse hepatitis virus (MHV), showing that inhibition or ablation of AXL reduces MHV infection of murine cells. In total, our findings provide evidence that PS receptors facilitate infection of the pandemic coronavirus SARS-CoV-2 and suggest that inhibition of the PS receptor AXL has therapeutic potential against SARS-CoV-2.

Highlights

Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) emerged in late 2019 and quickly spread around the world, resulting in the current public health pandemic
We evaluated if PS receptors serve as cell surface receptors for SARS-CoV-2 and found that the PS receptors, AXL, TIM-1 and TIM-4, facilitated virus infection when the SARS-CoV-2 cognate receptor, Angiotensin Converting Enzyme 2 (ACE2), was present
Consistent with the established mechanism of PS receptor utilization by other viruses, PS liposomes competed with SARS-CoV-2 for binding and entry

Summary

Introduction

Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) emerged in late 2019 and quickly spread around the world, resulting in the current public health pandemic. A herculean scientific effort has resulted in the development of SARS-CoV-2 vaccines which have been shown to be efficacious, potentially stemming the pandemic. In combination with vaccines, continued development of efficacious antivirals is needed, as outbreaks continue in under-vaccinated regions and severe disease caused by SARS-CoV-2 variants of concern is not eradicated following vaccination. Towards this goal, a more comprehensive understanding of SARS-CoV-2 interactions with host cells will be required

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