CD8⁺ T cells play an important role in anti-tumor immunity. Better understanding of their regulation could advance cancer immunotherapies. Here we identify, via stepwise CRISPR-based screening, that CUL5 is a negative regulator of the core signaling pathways of CD8⁺ T cells. Knocking out CUL5 in mouse CD8⁺ T cells significantly improves their tumor growth inhibiting ability, with significant proteomic alterations that broadly enhance TCR and cytokine signaling and their effector functions. Chemical inhibition of neddylation required by CUL5 activation, also enhances CD8 effector activities with CUL5 validated as a major target. Mechanistically, CUL5, which is upregulated by TCR stimulation, interacts with the SOCS-box-containing protein PCMTD2 and inhibits TCR and IL2 signaling. Additionally, CTLA4 is markedly upregulated by CUL5 knockout, and its inactivation further enhances the anti-tumor effect of CUL5 KO. These results together reveal a negative regulatory mechanism for CD8⁺ T cells and have strong translational implications in cancer immunotherapy.

CD8 ⁺ T细胞在抗肿瘤免疫中发挥重要作用。更好地了解它们的调节可以促进癌症免疫疗法。在这里，我们通过基于 CRISPR 的逐步筛选，确定 CUL5 是 CD8 ⁺ T 细胞核心信号通路的负调节因子。敲除小鼠 CD8 ⁺ T 细胞中的 CUL5 可以显着提高其肿瘤生长抑制能力，并具有显着的蛋白质组学改变，广泛增强 TCR 和细胞因子信号传导及其效应器功能。 CUL5 激活所需的 neddylation 的化学抑制也增强了 CD8 效应子活性，其中 CUL5 被验证为主要靶点。从机制上讲，TCR 刺激上调的 CUL5 与含有 SOCS-box 的蛋白 PCMTD2 相互作用，并抑制 TCR 和 IL2 信号传导。此外，CUL5敲除后CTLA4显着上调，其失活进一步增强了CUL5 KO的抗肿瘤作用。这些结果共同揭示了 CD8 ⁺ T 细胞的负调控机制，并对癌症免疫治疗具有很强的转化意义。