CX3CR1 Is a Receptor for Human Respiratory Syncytial Virus in Cotton Rats

Abstract

Respiratory syncytial virus (RSV) has been reported to use CX3CR1 in vitro as a receptor on cultured primary human airway epithelial cultures. To evaluate CX3CR1 as the receptor for RSV in vivo , we used the cotton rat animal model because of its high permissiveness for RSV infection. Sequencing the cotton rat CX3CR1 gene revealed 91% amino acid similarity to human CX3CR1. Previous work found that RSV binds to CX3CR1 via its attachment glycoprotein (G protein) to infect primary human airway cultures. To determine whether CX3CR1-G protein interaction is necessary for RSV infection, recombinant RSVs containing mutations in the CX3CR1 binding site of the G protein were tested in cotton rats. In contrast to wildtype virus, viral mutants did not grow in the lungs of cotton rats. When RSV was incubated with an antibody blocking the CX3CR1 binding site of G protein and subsequently inoculated intranasally into cotton rats, no virus was found in the lungs four days post-infection. In contrast, growth of RSV was not affected after pre-incubation with heparan sulfate (the receptor for RSV on immortalized cell lines). A reduction in CX3CR1 expression in the cotton rat lung through the use of peptide-conjugated morpholino oligomers led to a 10-fold reduction in RSV titers at day four post-infection. In summary, these results indicate that CX3CR1 functions as a receptor for RSV in cotton rats, and in combination with data from human airway epithelial cell cultures, strongly suggest that CX3CR1 is a primary receptor for naturally acquired RSV infection. Importance The knowledge about a virus receptor is useful to better understand the uptake of a virus into a cell and potentially develop antivirals either directed against the receptor molecule on the cell or the receptor-binding protein of the virus. Amongst a number of potential receptor proteins, human CX3CR1 has been demonstrated to act as receptor for respiratory syncytial virus (RSV) on human epithelial cells in tissue culture. Here we report that the cotton rat CX3CR1 which is similar to the human molecule acts as receptor in vivo. This study strengthens the argument that CX3CR1 is a receptor molecule for RSV.

The Respiratory Syncytial Virus G Protein Enhances the Immune Responses to the RSV F Protein in an Enveloped Virus-like Particle Vaccine Candidate

“…Furthermore, it has been recently shown that cotton rats express CX3CR1 in their lungs that is very similar to the human equivalent (34). Importantly, it has also been shown that RSV infection of cotton rat lungs is dependent upon CX3CR1 further validating these animals as a model of human infection and for vaccine candidate testing (34).…”

Immunopathology of RSV: An Updated Review

“…Moreover, CRs can be used in maternal antibody studies, as RSV antibodies transfer from dames to pups via the placenta and breast milk [297][298][299][300]. Further, CX3CR1 is a receptor used by RSV for infection of CRs [301]. While the cotton rat model has been used to evaluate drugs and vaccines, limitations of this model include cost, procurement (i.e., limited suppliers), and lack of robust tools to evaluate immune responses.…”

Receptors for Respiratory Syncytial Virus Infection and Host Factors Regulating the Life Cycle of Respiratory Syncytial Virus

“…G protein mediating virus attachment; inhibiting IFN type I production; promoting Th2-polarized response. (Imai et al, 1997;Tripp et al, 2001;Harcourt et al, 2006;Chirkova et al, 2013;Chirkova et al, 2015;Jeong et al, 2015;Johnson et al, 2015;Zhivaki et al, 2017;Anderson et al, 2020;Green et al, 2021) Nucleolin F protein mediating virus internalization (Tayyari et al, 2011;Hegele, 2012;Griffiths et al, 2020;Mastrangelo et al, 2021) EGFR F protein inducing macropinocytosis of RSV; promoting virus fusion; increasing airway mucus secretion. (Currier et al, 2016;Lingemann et al, 2019) IGF1R F protein facilitating the translocation of nucleolin to cell membrane (Griffiths et al, 2020) HSPGs G protein virus attachment.…”

“…The CX3CR1-CX3CL1 axis has been shown to play an important role in the pathogenesis of brain and neurodegenerative diseases, such as Alzheimer's disease (Lee et al, 2018), but its role in lung diseases, especially in infectious respiratory diseases, is poorly characterized. In RSV infection, the conserved CXC3 motif of its G glycoprotein was able to bind to CX3CR1 in vitro and in vivo to initiate infection (Jeong et al, 2015;Anderson et al, 2020;Green et al, 2021). Given its expression pattern in the lung, which matched the tropism of RSV infection, studies have shown that CX3CR1 is a more attractive candidate than HSPGs in vivo, since HSPGs are a primary receptor on immortalized cell lines but are expressed at much lower levels in the human airway epithelium, the primary site of RSV infection (Anderson et al, 1988;Johnson et al, 2015;Anderson et al, 2020;Green et al, 2021).…”

“…Adding heparin-subserted cell lines or reducing the heparan sulfate expression on the cellular surface could significantly reduce RSV infection. However, a lack of HSPGs expression on the apical surface of human bronchial epithelium or basal only expression indicated that HSPG might not be a candidate receptor for RSV to initiate infection in vivo (Anderson et al, 2020;Green et al, 2021).…”