Defining the roles of host factors and innate immune signaling in the ocular tropism of enterovirus D70

Project summary Enterovirus D70 (EVD70) is an understudied human enterovirus that causes acute hemorrhagic conjunctivitis, a highly contagious and painful ocular disease and that can, in rare instances, progress to polio-like paralysis. The molecular mechanisms underlying EVD70 infection, including 1) the host factors required for the viral life cycle and the 2) cellular tropism within the eye, remain poorly understood, due in part to a lack of eye-specific models. EVD70 attaches to ocular cells, at least in 2D cell culture, via sialic acids. There is, however, limited evidence suggesting the existence of an as-yet-unidentified proteinaceous receptor for EVD70 that triggers entry and uncoating in ocular cells. Through a genome-wide genetic knockout screen, I have identified host factors necessary for EVD70 infection in vitro, including the orphan receptor Jumping translocation breakpoint (JTB), which has limited demonstrated cellular functions. Notably, genetic knockout of JTB confers resistance to EVD70. I hypothesize that JTB serves as a receptor critical for EVD70 entry and uncoating in ocular cells and that JTB expression contributes to cellular tropism for corneal and conjunctival epithelial cells in the eye. The objectives of this proposal are to 1) determine the mechanism by which JTB is required for EVD70 infection and 2) define the cell-specific tropism and host immune responses to EVD70 in corneal organoids, which recapitulate the cellular diversity and organization of the ocular surface. Aim 1 will systematically evaluate if JTB is an entry and uncoating receptor for EVD70. Aim 2 will define the tropism of EVD70 and investigate host responses, such as interferon signaling, in corneal organoids using unbiased single cell RNA- sequencing and multiplexed cytokine analysis. Together, these aims will provide critical new insights into EVD70 biology and its interaction with ocular cells. Completing the proposed aims will advance our understanding of EVD70 molecular biology and pathogenesis and provide training to better equip me to progress towards independence as a scientist.