Vagal airway sensory nerve activation by beta-coronavirus spike protein

项目总结

PROJECT SUMMARY Activation of sensory nerves, in particular nociceptive C-fibers, is a feature of most respiratory viruses. Evidence of such activation is found in the classical consequences of C-fiber activation including sneezing, sore throat, coughing, and reflex secretions. As well as causing the troubling symptoms of viral infection, the activation of these nerves allows viruses to escape the body and be transmitted to other hosts, i.e. nociceptor activation amplifies viral spread in a community. In addition, activation of airway vagal C-fibers can lead to strong reflex bronchoconstriction and excessive secretions that likely contribute to the exacerbation of asthma particularly in children. Given the relevance to human disease, surprisingly little is known about how virus infection induces C-fiber activation and sensitization. In theory, viral infection leads to C-fiber activation by two general mechanisms. The first is that viral infection of epithelial cells leads to the production of a mediator(s) that stimulates the C-fiber terminals. The second is that the virus itself directly activates the nerves. This second mechanism will likely be dependent on the specific virus type. This proposal focuses on this second (direct) mechanism of activation as it relates to coronaviruses. I hypothesize that the coronavirus spike protein interacts directly with C-fiber terminals in a manner that activates and sensitizes the nociceptive C-fibers. My preliminary data, using three orthogonal approaches, support the conclusion that the spike protein directly activates (evokes action potential discharge) about 40- 50% of vagal C-fibers in mouse airways. My first aim is to characterize the subtype of vagal C-fibers that are activated by spike protein and also to assess whether the spike protein, short of overt activation, leads to the sensitization of C-fiber terminals, i.e. renders them more sensitive to other activating stimuli. My second aim focuses on the mechanism. I hypothesize that this interaction involves the galactin-3 fold in the spike protein, and occurs independently of the spike protein receptor ACE2 or toll-like receptors. Irrespective of the proximal binding target, I will address our hypothesis that activation is secondary to the opening of TRPV1 and or TRPA1 channels. These aims will be addressed using single cell RT-PCR analysis of mRNA expression in airway specific nociceptive C-fibers, extracellular and patch-clamp electrophysiology, and 2-photon live imaging techniques. The results of the studies are expected to provide insights into a novel mechanism of coronavirus induced airway C-fiber activation.