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.