T cell receptor (TCR) recognition of peptide–major histocompatibility complexes (pMHCs) is characterized by a highly conserved docking polarity. Whether this polarity is driven by recognition or signaling constraints remains unclear. Using “reversed-docking” TCRβ-variable (TRBV) 17⁺ TCRs from the naïve mouse CD8⁺ T cell repertoire that recognizes the H-2D^b–NP₃₆₆ epitope, we demonstrate that their inability to support T cell activation and in vivo recruitment is a direct consequence of reversed docking polarity and not TCR–pMHCI binding or clustering characteristics. Canonical TCR–pMHCI docking optimally localizes CD8/Lck to the CD3 complex, which is prevented by reversed TCR–pMHCI polarity. The requirement for canonical docking was circumvented by dissociating Lck from CD8. Thus, the consensus TCR–pMHC docking topology is mandated by T cell signaling constraints.

T 细胞受体 (TCR) 识别肽-主要组织相容性复合物 (pMHC) 的特征在于高度保守的对接极性。这种极性是否由识别或信号限制驱动仍不清楚。使用来自可识别 H-2D ^b –NP ₃₆₆的幼稚小鼠 CD8 ⁺ T 细胞库的“反向对接”TCRβ 变量 (TRBV) 17 ⁺ TCR表位，我们证明它们无法支持 T 细胞激活和体内募集是对接极性反转的直接结果，而不是 TCR-pMHCI 结合或聚类特征。典型的 TCR-pMHCI 对接将 CD8/Lck 最佳地定位到 CD3 复合物，这被反转的 TCR-pMHCI 极性所阻止。通过将 Lck 与 CD8 分离，规避了对规范对接的要求。因此，一致的 TCR-pMHC 对接拓扑结构受 T 细胞信号传导限制的约束。