1Department of Microbiology and Immunobiology, Harvard Medical School, 77 Avenue Louis Pasteur, Boston, MA 02115, USA.
Sci Signal. 2014 Jan 7;7(307):ra3. doi: 10.1126/scisignal.2004577.
Ubiquitylation is an important mechanism for regulating innate immune responses to viral infections. Attachment of lysine 63 (Lys(63))-linked ubiquitin chains to the RNA sensor retinoic acid-inducible gene-I (RIG-I) by the ubiquitin E3 ligase tripartite motif protein 25 (TRIM25) leads to the activation of RIG-I and stimulates production of the antiviral cytokines interferon-α (IFN-α) and IFN-β. Conversely, Lys(48)-linked ubiquitylation of TRIM25 by the linear ubiquitin assembly complex (LUBAC) stimulates the proteasomal degradation of TRIM25, thereby inhibiting the RIG-I signaling pathway. Here, we report that ubiquitin-specific protease 15 (USP15) deubiquitylates TRIM25, preventing the LUBAC-dependent degradation of TRIM25. Through protein purification and mass spectrometry analysis, we identified USP15 as an interaction partner of TRIM25 in human cells. Knockdown of endogenous USP15 by specific small interfering RNA markedly enhanced the ubiquitylation of TRIM25. In contrast, expression of wild-type USP15, but not its catalytically inactive mutant, reduced the Lys(48)-linked ubiquitylation of TRIM25, leading to its stabilization. Furthermore, ectopic expression of USP15 enhanced the TRIM25- and RIG-I-dependent production of type I IFN and suppressed RNA virus replication. In contrast, depletion of USP15 resulted in decreased IFN production and markedly enhanced viral replication. Together, these data identify USP15 as a critical regulator of the TRIM25- and RIG-I-mediated antiviral immune response, thereby highlighting the intricate regulation of innate immune signaling.
泛素化是调节固有免疫反应对抗病毒感染的重要机制。泛素 E3 连接酶三肽基结构域蛋白 25（TRIM25）将赖氨酸 63（Lys(63)）连接的泛素链连接到 RNA 传感器视黄酸诱导基因-I（RIG-I）上，导致 RIG-I 的激活，并刺激抗病毒细胞因子干扰素-α（IFN-α）和 IFN-β的产生。相反，线性泛素组装复合物（LUBAC）对 TRIM25 的 Lys(48)-连接泛素化刺激 TRIM25 的蛋白酶体降解，从而抑制 RIG-I 信号通路。在这里，我们报告泛素特异性蛋白酶 15（USP15）对 TRIM25 进行去泛素化，防止 LUBAC 依赖的 TRIM25 降解。通过蛋白质纯化和质谱分析，我们在人类细胞中鉴定 USP15 为 TRIM25 的相互作用伙伴。特异性小干扰 RNA 敲低内源性 USP15 显着增强了 TRIM25 的泛素化。相比之下，野生型 USP15 的表达，而不是其催化失活突变体，减少了 TRIM25 的 Lys(48)-连接泛素化，导致其稳定。此外，USP15 的异位表达增强了 TRIM25 和 RIG-I 依赖性 I 型 IFN 的产生并抑制了 RNA 病毒复制。相反，USP15 的耗竭导致 IFN 产生减少和病毒复制显着增强。总之，这些数据表明 USP15 是 TRIM25 和 RIG-I 介导的抗病毒免疫反应的关键调节剂，从而突出了先天免疫信号的复杂调节。
