Antiviral Activity of Guanylate-Binding Proteins and Viral Countermeasures

鸟苷酸结合蛋白（GBP）是干扰素诱导的GTP酶，已知在针对细菌和原生动物的保护性先天免疫中起关键作用。人类编码七种GBP旁系同源物。利用进化基因组学和蛋白质相互作用数据，我们发现其中一个（GBP 5）具有已知抗逆转录病毒限制因子的特征。功能分析证实，GBP 5通过干扰病毒包膜（Env）糖蛋白的加工和病毒体掺入来限制HIV-1和其他逆转录病毒。GBP 5的抑制活性需要异戊烯化依赖的高尔基体定位，但不需要其GT3功能。GBP 5有效地限制了人类巨噬细胞中的HIV-1，一些脑源性HIV-1毒株通过一种不寻常的权衡机制逃避这种抗病毒作用：Vpu起始密码子的突变以Vpu功能为代价增加了Env表达。我们正在进行的研究表明，GBP 5也可通过L-27和TCR-CD 3信号传导诱导，GBP 2也限制慢病毒复制，并且GBP 2和GBP 5的抗病毒活性在进化上是保守的。在拟议的项目中，我们希望定义限制以及慢病毒逃避或抵消的确切机制。首先，我们将绘制对GBP 5的抗病毒活性至关重要的氨基酸残基，并分析它们对Env加工、运输和病毒体掺入的影响。此外，我们将确定HIV-1和HIV-2对GBP 2/5抑制的易感性在从类人猿和白眉猴传播SIV后是如何演变的，以阐明GBPs是否构成成功的人畜共患病病毒传播的障碍。这些研究还将揭示这些病毒如何逃避或抵消GBP 2/5的抑制作用。另一个焦点将是通过不同的细胞因子在原代病毒靶细胞中靶向诱导GBP 2/5表达。最后，我们的初步结果表明，GBP 2和GBP 5不仅抑制逆转录病毒。因此，我们将分析GBP蛋白对各种病毒病原体的影响，如流感病毒、埃博拉病毒、马尔堡病毒、莱莎病毒、呼吸道合胞病毒和SARS冠状病毒。这些结果将对抗病毒防御机制和病毒对策产生重要的见解。

Guanylate-binding proteins (GBPs) are interferon-inducible GTPases known to play key roles in protective innate immunity against bacteria and protozoa. Humans encode seven GBP paralogs. Using evolutionary genomics and protein interaction data, we found that one of them (GBP5) shares characteristics of known antiretroviral restriction factors. Functional analyses confirmed that GBP5 restricts HIV-1 and other retroviruses by interfering with processing and virion incorporation of the viral envelope (Env) glycoprotein. The inhibitory activity of GBP5 requires isoprenylation-dependent Golgi-localization but not its GTPase function. GBP5 efficiently restricts HIV-1 in human macrophages, and some brain-derived HIV-1 strains evade this antiviral effect by an unusual tradeoff mechanism: mutations in the vpu initiation codon increase Env expression at the cost of Vpu function. Our ongoing studies demonstrate that GBP5 is also inducible by L-27 and TCR-CD3 signaling, that GBP2 also restricts lentiviral replication, and that the antiviral activity of GBP2 and GBP5 is evolutionarily conserved. In the proposed project, we want to define the exact mechanism(s) underlying restriction as well as lentiviral evasion or counteraction. Initially, we will map the amino acid residues that are critical for the antiviral activity of GBP5 and analyze their effect on Env processing, trafficking and virion incorporation. Furthermore, we will determine how the susceptibility of HIV-1 and HIV-2 to GBP2/5 inhibition evolved after transmission of SIVs from great apes and sooty mangabeys to elucidate whether GBPs constitute a barrier to successful zoonotic viral transmission. These studies will also reveal how these viruses evade or counteract the inhibitory effect of GBP2/5. Another focus will be the targeted induction of GBP2/5 expression in in primary viral target cells by different cytokines. Finally, our preliminary results show that GBP2 and GBP5 do not only inhibit retroviruses. Thus, we will analyse the effects of GBP proteins on a variety of viral pathogens, such as Influenza, Ebola, Marburg, Lyssa, Respiratory Syncytial and SARS Coronavirus. The results will yield important insights into antiviral defence mechanisms and viral countermeasures.