Project Summary COVID-19 is a dangerous pandemic disease caused by the highly infectious coronavirus SARS-CoV-2, which arose by zoonotic transfer from bats. CD8-positive T cells contribute to viral elimination by killing infected cells, preventing viral expansion in an infected individual and limiting the exposure of others to infection. CD8-positive T cells kill virally infected cells by recognizing Major Histocompatibility Complex class I (MHC-I) molecules on their surface that are associated with peptides derived from viral proteins. Like many viruses, SARS-CoV-2 encodes specific proteins that affect surface expression of MHC-I-peptide complexes, resulting in resistance to T cell-mediated killing. We hypothesize that, since SARS-CoV-2 transferred from bats to humans, its MHC-I inhibitory properties evolved in bats. We further hypothesize that, because evolutionary pressure on the virus was mediated by natural selection in the face of chronic exposure to the bat immune system, inhibition of MHC-I function will be more efficient in bats than in humans. Investigating these hypotheses is limited by the lack of appropriate reagents. This proposal seeks to remedy the situation by developing antibodies in mice and rabbits and nanobodies in yeast that identify bat MHC-I proteins as well as the bat equivalents of the additional human gene products that facilitate the generation and surface expression of MHC-I molecules containing bound antigenic peptides, namely TAP1 and TAP2, tapasin, calreticulin and the thiol oxidoreductase ERp57 (PDIA3). As we produce these reagents we will use them to characterize MHC-I-restricted antigen processing in bats and determine how SARS-CoV-2 infection affects it. We have identified four SARS-CoV-2 proteins that independently cause MHC-I down-regulation in humans, and we suggest that these proteins acting in combination are responsible for the reduction in MHC-I surface expression in infected cells. Our goal is to determine whether these gene products, and potentially others, affect the same process in bat cells and how well they do it. Understanding MHC-I function and adaptive immunity in bats will help us to uncover why they are a such potent reservoir for coronaviruses. Determining how bats survive infection by such viruses may suggest targeted therapeutic approaches to improve the ability of humans to resist them and enhance our ability to control and defeat future coronavirus pandemics.