Center for Inflammation Research, VIB, Unit of Molecular Signal Transduction in Inflammation, Technologiepark 927, Zwijnaarde, Ghent 9052, Belgium; Department of Biomedical Molecular Biology, Ghent University, Technologiepark 927, Zwijnaarde, Ghent 9052, Belgium.
Institute of Cell Biology and Immunology, University of Stuttgart, Allmandring 31, 70569 Stuttgart, Germany; Department of Biology, Drexel University, 3245 Chestnut Street, Philadelphia, PA 19104, USA.
Biochem Pharmacol. 2018 Jul;153:292-298. doi: 10.1016/j.bcp.2018.01.039. Epub 2018 Jan 31.
Tumor Necrosis Factor (TNF) is a proinflammatory cytokine that elicits its action by binding to two cell surface TNF receptors (TNFR), TNFR1 and TNFR2, which are expressed by many different cell types. Stimulation of TNFR1 activates canonical NF-κB signaling, leading to the NF-κB dependent expression of a large number of genes. Canonical NF-κB signaling requires the assembly of a TNFR1 signaling complex at the cell membrane, whose formation is regulated by different protein ubiquitination events. In this context, recruitment of the Linear Ubiquitin Chain Assembly Complex (LUBAC) to TNFR1 plays an important role by mediating M1-linked polyubiquitination of specific NF-κB signaling proteins. In contrast to TNFR1, much less is known about the role of ubiquitination in TNFR2 signaling. Here we demonstrate that specific TNFR2 stimulation rapidly triggers M1- and K63-linked polyubiquitination at the TNFR2 signaling complex. In agreement, TNFR2 stimulation induces the recruitment of HOIP, a LUBAC component and the only known E3 ubiquitin ligase for M1-polyubiquitination, to the TNFR2 signaling complex. Also cIAP1, a E3 ubiquitin ligase able to modify proteins with K63-polyubiquitin chains, was recruited to the TNFR2 signaling complex. Treatment of cells with a cIAP antagonist inhibited the recruitment of HOIP and prevented HOIP-mediated M1-ubiquitination of the TNFR2 signaling complex, indicating that HOIP recruitment to the TNFR2 relies on cIAPs. Finally, we show that both HOIP and cIAP1 are required for TNFR2-induced canonical NF-κB activation. Together, our findings demonstrate an important role for M1- and K63-linked polyubiquitination in TNFR2 signaling.
肿瘤坏死因子 (TNF) 是一种促炎细胞因子，通过与两种细胞表面 TNF 受体 (TNFR)，即 TNFR1 和 TNFR2 结合来发挥作用，TNFR1 和 TNFR2 表达于许多不同类型的细胞。TNFR1 的刺激激活经典 NF-κB 信号通路，导致 NF-κB 依赖性大量基因的表达。经典 NF-κB 信号通路需要在细胞膜上组装 TNFR1 信号复合物，其形成受不同蛋白泛素化事件的调节。在这种情况下，线性泛素链组装复合物 (LUBAC) 招募到 TNFR1 发挥重要作用，通过介导特定 NF-κB 信号蛋白的 M1 连接多泛素化。与 TNFR1 相比，关于泛素化在 TNFR2 信号中的作用知之甚少。在这里，我们证明特定的 TNFR2 刺激迅速触发 TNFR2 信号复合物上的 M1 和 K63 连接多泛素化。一致地，TNFR2 刺激诱导 LUBAC 成分 HOIP 和唯一已知的 M1-多泛素化 E3 泛素连接酶招募到 TNFR2 信号复合物。也招募到 TNFR2 信号复合物的 cIAP1，一种能够修饰具有 K63-多泛素链的蛋白的 E3 泛素连接酶。用 cIAP 拮抗剂处理细胞可抑制 HOIP 的募集并阻止 HOIP 介导的 TNFR2 信号复合物的 M1 泛素化，表明 HOIP 向 TNFR2 的募集依赖于 cIAP。最后，我们表明 HOIP 和 cIAP1 都需要 TNFR2 诱导的经典 NF-κB 激活。总之，我们的研究结果表明 M1 和 K63 连接的多泛素化在 TNFR2 信号中起着重要作用。
