Mapping of functional SARS-CoV-2 receptors in human lungs establishes differences in variant binding and SLC1A5 as a viral entry modulator of hACE2

Background The COVID-19 pandemic is an infectious disease caused by SARS-CoV-2. The first step of SARS-CoV-2 infection is the recognition of angiotensin-converting enzyme 2 (ACE2) receptors by the receptor-binding domain (RBD) of the viral Spike (S) glycoprotein. Although the molecular and structural bases of the SARS-CoV-2-RBD/ hACE2 interaction have been thoroughly investigated in vitro, the relationship between hACE2 expression and in vivo infection is less understood. Methods Here, we developed an efficient SARS-CoV-2-RBD binding assay suitable for super resolution microscopy and simultaneous hACE2 immunodetection and mapped the correlation between hACE2 receptor abundance and SARS-CoV-2-RBD binding, both in vitro and in human lung biopsies. Next, we explored the specific proteome of SARS-CoV-2-RBD/hACE2 through a comparative mass spectrometry approach. Findings We found that only a minority of hACE2 positive spots are actually SARS-CoV-2-RBD binding sites, and that the relationship between SARS-CoV-2-RBD binding and hACE2 presence is variable, suggesting the existence of additional factors. Indeed, we found several interactors that are involved in receptor localization and viral entry and characterized one of them: SLC1A5, an amino acid transporter. High-resolution receptor-binding studies showed that co-expression of membrane-bound SLC1A5 with hACE2 predicted SARS-CoV-2 binding and entry better than hACE2 expression alone. SLC1A5 depletion reduces SARS-CoV-2 binding and entry. Notably, the Omicron variant is more efficient in binding hACE2 sites, but equally sensitive to SLC1A5 downregulation. Interpretation We propose a method for mapping functional SARS-CoV-2 receptors in vivo. We confirm the existence of hACE2 co-factors that may contribute to differential sensitivity of cells to infection. Copyright (c) 2022 The Authors. Published by Elsevier B.V. This is an open access article under the CC BY-NC-ND license (http://creativecommons.org/licenses/by-nc-nd/4.0/).

Automated summary

The study found that the binding of the SARS-CoV-2 virus to hACE2 receptors is the first step in infection, but the abundance of receptors is influenced by various factors. Further research revealed SLC1A5 as a factor involved in receptor localization and viral entry, and co-expression with hACE2 improved the prediction of SARS-CoV-2 binding and entry. The Omicron variant showed higher efficiency in binding to hACE2 sites, but was equally sensitive to SLC1A5 downregulation.