Antiviral Activity of Guanylate-Binding Proteins and Viral Countermeasures

鸟苷结合蛋白(GBP)是干扰素诱导的GTP酶，在保护细菌和原生动物的天然免疫中发挥关键作用。人类编码七个英镑对偶数。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(GBP5)具有已知抗逆转录病毒限制因子的特征。功能分析证实，GBP5通过干扰病毒包膜(Env)糖蛋白的加工和病毒粒子的掺入来限制HIV-1和其他逆转录病毒。GBP5的抑制活性需要依赖于异戊二烯基化的高尔基体定位，而不需要它的GTP酶功能。GBP5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了这一抗病毒作用：VPU起始密码子的突变以VPU功能为代价增加了Env的表达。我们正在进行的研究表明，GBP5也被L-27和TCRCD3信号诱导，GBP2也限制慢病毒的复制，并且GBP2和GBP5的抗病毒活性在进化上是保守的。在拟议的项目中，我们希望定义潜在的限制以及慢病毒逃避或对抗的确切机制(S)。首先，我们将定位对GBP5的抗病毒活性至关重要的氨基酸残基，并分析它们对环境病毒的加工、运输和病毒粒子掺入的影响。此外，我们将确定HIV-1和HIV-2对GBP2/5抑制的易感性在从类人猿和煤烟芒果中传播SIV后是如何演变的，以阐明GBP是否构成成功传播人畜共患病毒的障碍。这些研究还将揭示这些病毒如何逃避或抵消GBP2/5的抑制作用。另一个焦点将是不同细胞因子在原代病毒靶细胞中靶向诱导GBP2/5的表达。最后，我们的初步结果表明，GBP2和GBP5不仅能抑制逆转录病毒。因此，我们将分析GBP蛋白对多种病毒病原体的影响，如流感、埃博拉、马尔堡、Lyssa、呼吸道合胞病毒和SARS冠状病毒。这一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.