Two Hosts, One Receptor: Comparing TLR4 Responses to Viral Glycoproteins in Humans and Bats
Abstract
Innate immunity represents the first line of defence against invading pathogens. A key component of the innate immune system is Toll-like receptor 4 (TLR4), which is classically activated by bacterial lipopolysaccharide (LPS) leading to the release of pro-inflammatory cytokines. Interestingly, recent literature has shown that viral glycoproteins, including SARS-CoV-2 spike and Ebola virus glycoprotein (EBOV GP) can also activate TLR4. Bats harbour many pathogenic viruses while exhibiting minimal signs of disease, suggesting differences in immune responses between bats and humans. Our lab has shown that bat TLR4 is less responsive to LPS stimulation, while being hyperresponsive to viral glycoproteins, suggesting differences in TLR4 signalling between bats and humans. Using AlphaFold and HDOCK, we performed in silico structural analyses to identify potential interactions between the TLR4 signaling complex and viral glycoproteins. We observed direct interactions between viral glycoproteins and the TLR4 complex. To assess downstream signalling of TLR4, we utilized a luciferase-based reporter system, in which luciferase expression is indicative of TLR4 activation. Since TLR4 signaling classically requires accessory proteins MD2 and CD14, we characterized their roles in viral glycoprotein-mediated TLR4 activation. We found that viral activation of TLR4 is abolished in the absence of MD2 and dampened without CD14. Ongoing cloning of EBOV GP into the vesicular stomatitis virus genome will enable further characterization of EBOV GP immune activation in a surrogate biosafety level 2 model that more closely mimics viral infection compared to our overexpression model. Future studies will investigate TLR4 dimerization in the presence of viral glycoproteins using co-immunoprecipitation. Collectively, this work contributes to a greater understanding of the molecular mechanisms underlying how viral glycoproteins activate TLR4 and differences in innate immune signalling between humans and bats. Further investigation may provide insight into the regulation of immune activation during viral infection and inform future therapeutic strategies. Faculty supervisor: Che C. Colpitts supporting lab instructor: Trinity H. Tooley