Unconventional CD8+ T cell surveillance of cytomegalovirus via Qa-1/HLA-E-restricted epitope recognition.

Nonclassical CD8+ T cells can compensate for classical CD8+ T cell effector responses during murine cytomegalovirus (MCMV) infection. Through a combination of motif-based discovery, predictive algorithms, AlphaFold3 structural modeling, and biological assays, we identified multiple MCMV and human cytomegalovirus (HCMV) peptides that bind to Qa-1 and HLA-E, respectively. In the mouse system, we demonstrated that these virally encoded antigens stimulate Qa-1-restricted CD8+ T cells ex vivo, which can be tracked using MCMV peptide-loaded Qa-1 tetramers. Adoptive transfer of predominantly Qa-1 tetramer+ CD8+ T cells into RAG-1-deficient mice protects them from mortality, underscoring the critical role of these cells in host defense. Single-cell RNA (scRNA)/TotalSeq and single-cell T cell receptor sequencing (scTCR-seq) reveal the expansion of unique TCR αβ clonotypes, indicating convergent antigen specificity. Together, our findings uncover a conserved and functionally important nonclassical CD8+ T cell axis mediated by Qa-1/HLA-E modulating adaptive immunity independent of classical major histocompatibility complex class I (MHC-I) pathways and present previously unidentified opportunities for vaccine development.