The primary role of CD8+ T cells is to eliminate pathogen infected cells and cancers. Central to CD8+ T cell function is the ability of the T cell receptor (TCR) to recognize foreign peptides (p) or tumour peptides presented on MHC Class I (MHCI) molecules. Despite the extraordinary diversity inherent in all components of TCR-pMHCI recognition, structural analyses from both mice and humans has revealed a highly conserved recognition polarity; namely the TCRa chain reproducibly docks over the MHCI a2 helix while the TCRb chain docks over the MHCI a1 helix. It remains unknown whether this conserved modality of TCR recognition of pMHCI is driven by germline encoded recognition motifs within the TCR and MHCI, or due to constraints imposed by the nature of TCR-CD3 signaling. We recently solved the first ternary structures of epitope-specific TCRs from a naïve mouse repertoire in complex with their pMHCI ligand, revealing a completely reversed docking polarity. By expressing these reversed docking TCRs in vitro and in vivo, and in conjunction with state of the art cellular, biochemical and imaging techniques, we have been able to dissect the critical requirements for effective TCR recognition of pMHC and the mechanisms by which this translates to full T cell activation and effector function.