Similar activation-driven transcriptional programs in lung CMV- and flu- specific memory CD8+ T cells despite differences in tissue residency profiles

To investigate whether virus-specific human lung memory CD8? T cells exhibit distinct transcriptional responses based on viral tropism, we stimulated total human lung cells ex vivo with peptide pools from either influenza virus (epithelial-restricted) or cytomegalovirus (CMV, broader tropism). Antigen-specific CD8? T cells were identified using activation-induced markers (CD25 and CD137) and sorted for bulk RNA sequencing. Despite differences in tissue residency phenotypes, influenza- and CMV-specific CD8? T cells displayed highly similar activation-induced transcriptional programs across nine donors, with shared enrichment of inflammatory and T cell signaling pathways. These findings suggest that lung memory CD8? T cells mount comparable responses to antigen stimulation, regardless of viral specificity or residency profile. Overall design: Total human lung cells from n=9 donors were stimulated in vitro for 18 hours with peptide pools specific for either influenza virus or cytomegalovirus (CMV), or left unstimulated. Antigen-specific CD8? T cells were identified using activation-induced markers (CD25 and CD137) and isolated by FACS for bulk RNA sequencing. A PMA/ionomycin-stimulated sample was included as a positive control to set sorting gates. Unstimulated memory CD8?CD69? T cells were also sorted as a comparison population.