Diverse Animal Sarbecoviruses Can Infect Human Cells Through ACE2-Dependent and ACE2-Independent Pathways, Impacting Antibody Neutralization

Core Concepts

Animal sarbecoviruses can infect human cells through ACE2-dependent and ACE2-independent pathways, with the latter pathway reducing susceptibility to antibody-mediated neutralization.

Abstract

The study examines the determinants that govern the ability of animal sarbecoviruses to infect human cells and the implications for antibody-mediated neutralization. Key findings include:

Raccoon dog ACE2 exhibits the broadest receptor activity for diverse animal sarbecoviruses, suggesting raccoon dogs could serve as potential intermediate hosts or reservoirs.

The receptor binding domain (RBD) is a key determinant controlling whether sarbecovirus entry into human cells is ACE2-dependent or ACE2-independent and trypsin-dependent.

Several type II transmembrane serine proteases (TTSPs), including TMPRSS11A and TMPRSS11D, can cleave and activate sarbecovirus S proteins for ACE2-independent entry, potentially facilitating viral spread in the respiratory tract.

Insertion of a multibasic cleavage site into the S protein increases lung cell entry for most sarbecoviruses, but does not enable trypsin-independent entry for the Rs4081 and Rs4237 S proteins.

Antibodies induced by quadruple COVID-19 vaccination robustly neutralize entry driven by diverse sarbecovirus S proteins. However, usage of the ACE2-independent, trypsin-dependent entry pathway reduces susceptibility to antibody-mediated neutralization.

Stats

The study reports the following key metrics and figures:

Binding of soluble human ACE2 to cells expressing different sarbecovirus S proteins (Figure 2A)
Receptor activity of ACE2 orthologues from different animal species for entry driven by sarbecovirus S proteins (Figure 2B)
Efficiency of sarbecovirus S protein-driven entry into various human and animal cell lines, with and without trypsin treatment (Figure 3A)
Concentration-dependent cleavage of sarbecovirus S proteins by trypsin (Figure 4A)
Concentration-dependent effects of trypsin on sarbecovirus S protein-driven cell entry (Figure 4B)
Cleavage of sarbecovirus S proteins by different proteases (Figure 5A, 5E)
Effects of protease expression on sarbecovirus S protein-driven cell entry (Figure 5B, 5D, 5F)
Impact of inserting a multibasic cleavage site into sarbecovirus S proteins on entry into 293T-ACE2 and Calu-3-ACE2 cells (Figure 6C)
Cell entry driven by chimeric SARS-1-S and Rs4081 S proteins, with and without trypsin treatment (Figure 7D)
Neutralization of sarbecovirus S protein-driven entry by the pan-sarbecovirus antibody S2H97, with and without trypsin treatment (Figure 8)
Neutralization of sarbecovirus S protein-driven entry by antibodies from COVID-19 convalescent, double-vaccinated, triple-vaccinated, and quadruple-vaccinated individuals (Figure 9A)
Impact of trypsin treatment on neutralization of sarbecovirus S protein-driven entry by antibodies from triple-vaccinated individuals (Figure 9B)

Quotes

"Raccoon dog ACE2 supported entry driven by all tested clade 1 and 3 sarbecovirus S proteins with at least the same or, for several S proteins, even higher efficiency than human ACE2."
"Several TMPRSS2-related cellular proteases but not the insertion of a multibasic cleavage site into the S protein allowed for ACE2-independent entry in the absence of trypsin and may support viral spread in the respiratory tract."
"Antibodies induced upon quadruple COVID-19 vaccination robustly neutralized entry driven by all S proteins studied and might thus install appreciable protection against zoonotic animal sarbecoviruses."

Key Insights Distilled From

ACE2-independent sarbecovirus cell entry is supported by TMPRSS2-related enzymes and reduces sensitivity to antibody-mediated neutralization

by Zhang,L., Ch... at www.biorxiv.org 04-18-2024

https://www.biorxiv.org/content/10.1101/2024.04.18.590061v1

Deeper Inquiries

What other animal species or environmental factors might contribute to the zoonotic potential of diverse sarbecoviruses beyond the determinants examined in this study

In addition to the determinants examined in this study, other animal species and environmental factors could contribute to the zoonotic potential of diverse sarbecoviruses. One key factor to consider is the diversity of ACE2 receptors across different animal species. While this study highlighted the broad receptor activity of raccoon dog ACE2 for sarbecoviruses, other animal species may possess ACE2 orthologues with varying affinities for viral entry. For example, studies have shown that certain bat species harbor ACE2 receptors that can facilitate viral entry, making them potential reservoirs for sarbecoviruses. Additionally, the presence of other cellular proteases, besides TMPRSS2-related enzymes, in different animal species could also influence the ability of sarbecoviruses to infect and spread. Environmental factors such as habitat overlap between different animal species, human encroachment into wildlife habitats, and the presence of intermediate hosts could also play a role in the zoonotic transmission of sarbecoviruses.

How might the ACE2-independent, trypsin-dependent entry pathway impact sarbecovirus pathogenesis and tissue tropism in vivo, and what are the implications for disease severity and transmission

The ACE2-independent, trypsin-dependent entry pathway identified in this study could have significant implications for sarbecovirus pathogenesis, tissue tropism, disease severity, and transmission in vivo. This pathway may allow sarbecoviruses to infect cells in the respiratory tract and other tissues without relying solely on ACE2 receptors, potentially broadening their host range and tissue tropism. The ability to utilize trypsin for entry could enhance viral spread within the respiratory tract, as trypsin is present in the respiratory epithelium and can activate the S protein for membrane fusion. This could lead to increased viral replication, tissue damage, and potentially more severe respiratory disease. The ACE2-independent, trypsin-dependent entry pathway may also impact viral transmission by enabling efficient cell entry in the absence of ACE2, potentially facilitating transmission between hosts and contributing to the zoonotic potential of sarbecoviruses.

Given the potential for antibody evasion through the ACE2-independent entry pathway, what other immune responses or antiviral strategies could be leveraged to provide broad protection against diverse sarbecovirus threats

In light of the potential for antibody evasion through the ACE2-independent, trypsin-dependent entry pathway, it is crucial to explore alternative immune responses and antiviral strategies to provide broad protection against diverse sarbecovirus threats. One approach could involve the development of vaccines that target conserved regions of the viral proteins, such as the S2 subunit, to elicit cross-reactive and neutralizing antibodies against multiple sarbecoviruses. Broad-spectrum antiviral therapies targeting viral entry, replication, and spread mechanisms could also be explored to complement vaccine strategies. Additionally, enhancing innate immune responses, such as interferon signaling pathways, could help bolster the host's ability to combat sarbecovirus infections and reduce the risk of immune evasion. Collaborative efforts between researchers, public health agencies, and pharmaceutical companies will be essential to develop comprehensive strategies for combating the zoonotic and pandemic potential of sarbecoviruses.