Molecular determinants of cross-strain influenza A virus recognition by αβ T cell receptors

Cross-reactive αβ T cell receptors (TCRs) recognizing multiple peptide variants can provide effective control of rapidly evolving viruses yet remain understudied. By screening 12 naturally occurring influenza-derived HLA-B*35:01-restricted nucleoprotein (NP)_418-426 epitopes (B*35:01-NP₄₁₈) that emerged since 1918 within influenza A viruses, including 2024 A/H5N1 viruses, we identified functional broadly cross-reactive T cells universally recognizing NP₄₁₈ variants. Binding studies demonstrated that TCR cross-reactivity was concomitant with diminished antigen sensitivity. Primary human B*35:01/NP₄₁₈⁺CD8⁺ T cell lines displayed reduced cross-reactivity in the absence of CD8 coreceptor binding, validating the low avidity of cross-reactive B*35:01-NP₄₁₈⁺CD8⁺ T cell responses. Six TCR-HLA-B*35:01/NP₄₁₈ crystal structures showed how cross-reactive TCRs recognized multiple B*35:01/NP₄₁₈ epitope variants. Specific TCR interactions were formed with invariant and conserved peptide-HLA features, thus remaining distal from highly varied positions of the NP₄₁₈ epitope. Our study defines molecular mechanisms associated with extensive TCR cross-reactivity toward naturally occurring viral variants highly relevant to universal protective immunity against influenza.