The CTLA-4 and PD-1 checkpoints control immune responses to self-antigens and are key targets in cancer immunotherapy. Both pathways are connected via a cis interaction between CD80 and PD-L1, the ligands for CTLA-4 and PD-1 respectively. This cis interaction prevents PD-1 binding to PD-L1 but is reversed by CTLA-4 trans-endocytosis of CD80. However, the mechanism by which CTLA-4 selectively removes CD80 but not PD-L1 is unclear. Here we show that CTLA-4 - CD80 interactions are unimpeded by PD-L1 and that CTLA-4 binding with CD80 does not displace PD-L1 per se. Rather, both the rigidity and bivalency of the WT CTLA-4 molecule is required to orientate CD80 such that PD-L1 interactions are no longer permissible. Moreover, soluble CTLA-4 released PD-L1 only at specific expression levels of CD80 and PD-L1, whereas CTLA-4 trans-endocytosis released PD-L1 in all conditions. These data show that PD-L1 release from CD80 is driven by biophysical factors associated with orientation and bivalent cross-linking of proteins in the cell membrane and that trans-endocytosis of CD80 efficiently promotes PD-L1 availability.

中文翻译：

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需要与 CD80 结合的刚性二价 CTLA-4 才能破坏顺式 CD80 / PD-L1 相互作用

CTLA-4 和 PD-1 检查点控制对自身抗原的免疫反应，是癌症免疫治疗的关键靶点。两条通路通过 CD80 和 PD-L1（分别是 CTLA-4 和 PD-1 的配体）之间的顺式相互作用连接。这种顺式相互作用阻止 PD-1 与 PD-L1 结合，但可通过 CD80 的 CTLA-4 反式内吞作用逆转。然而，CTLA-4 选择性去除 CD80 而不是 PD-L1 的机制尚不清楚。在这里，我们表明 CTLA-4 - CD80 相互作用不受 PD-L1 阻碍，并且 CTLA-4 与 CD80 的结合不会取代 PD-L1 本身。相反，WT CTLA-4 分子的刚性和二价性都需要定向 CD80，从而不再允许 PD-L1 相互作用。此外，可溶性CTLA-4仅在CD80和PD-L1的特定表达水平下释放PD-L1，而CTLA-4反式内吞作用在所有条件下都释放PD-L1。这些数据表明，CD80 释放 PD-L1 是由与细胞膜中蛋白质的方向和二价交联相关的生物物理因素驱动的，并且 CD80 的反式内吞作用可有效促进 PD-L1 的可用性。