Supplementary Material for: Inducible GBP5 Mediates the Antiviral Response via Interferon-Related Pathways during Influenza A Virus Infection

Guanylate binding protein (GBP) 5 belongs to the GBP family, which is involved in important cellular processes, including signal transduction, translation, vesicle trafficking, and exocytosis. Structurally, GBPs display a high degree of homology and share highly conserved GTP-binding or hydrolysis domains. GBP5 was reported to be a critical cellular factor in inflammasome assembly. However, little is known about its role in the host antiviral innate immune response. In this study, we found that GBP5 expression was significantly elevated in influenza patients and influenza A virus-infected A549 human lung epithelial cells. The overexpression of GBP5 inhibited virus replication by enhancing the expression of virus-induced interferon (IFN) and IFN-related effectors. Knockdown of GBP5 had the opposite effect. Moreover, GBP5 enhanced endogenous IFN expression by interacting with the NF-κB-essential modulator complex and stimulating NF-κB signaling. Additionally, the expression of proinflammatory factors, such as IL-6, IL-8, tumor necrosis factor-α, cyclooxygenase-2, and inducible nitric oxide synthase, was also activated by GBP5. Taken together, our results reveal that GBP5 inhibited virus replication through the activation of IFN signaling and proinflammatory factors.

鸟苷酸结合蛋白5（Guanylate binding protein 5, GBP5）属于GBP家族，该家族参与包括信号转导、翻译、囊泡运输及胞吐作用在内的诸多关键细胞进程。从结构层面而言，GBP家族成员具有高度的序列同源性，且共享高度保守的GTP结合或水解结构域。已有研究证实，GBP5是介导炎性小体组装的关键细胞因子。然而，目前学界对于其在宿主抗病毒天然免疫应答中的具体功能仍知之甚少。本研究中，我们发现流感患者体内以及甲型流感病毒感染的人肺上皮A549细胞中，GBP5的表达水平均显著升高。过表达GBP5可通过上调病毒诱导的干扰素（interferon, IFN）及其相关效应分子的表达，进而抑制病毒复制；而敲低GBP5则会产生完全相反的效应。此外，GBP5可通过与NF-κB必需调节蛋白复合物（NF-κB-essential modulator complex）相互作用，激活NF-κB信号通路，促进内源性IFN的表达。同时，白细胞介素6（interleukin 6, IL-6）、白细胞介素8（interleukin 8, IL-8）、肿瘤坏死因子α（tumor necrosis factor-α, TNF-α）、环氧合酶2（cyclooxygenase-2, COX-2）及诱导型一氧化氮合酶（inducible nitric oxide synthase, iNOS）等促炎因子的表达，亦可被GBP5激活。综上，本研究结果揭示，GBP5可通过激活IFN信号通路及促炎因子的表达，抑制病毒复制。