HIV-1跨种传播限制性和易感非人灵长类动物模型初步研究

发布时间：2018-12-26 16:13

【摘要】： 艾滋病研究仍缺乏适合的动物模型。非人灵长类动物模型被认为是研究艾滋病毒(HIV-1)感染和机体反应的理想动物模型,但是这些动物中存在多种遗传因子限制了HIV-1的跨种传播感染,使该模型应用受到限制。近年来的研究发现了非人灵长类动物体内的两个HIV-1关键性限制因子,即TRIM5α和APOBEC3G。猴TRIM5α能够结合HIV-1衣壳蛋白(CA)而限制其复制,但不能限制猴艾滋病毒(SIV)的复制。HIV-1Vif蛋白能够有效地对抗人APOBEC3G的限制活性,而对猴APOBEC3G无效,但SIV Vif能有效地对抗猴APOBEC3G的限制功能。因此,用SIV CA和Vif基因替换HIV-1的相应基因后,可能实现HIV-1对猴体的跨种传播感染。在另一方面,在部分旧大陆猴的猕猴中发现了TRIM5α的一种CypA基因插入剪接突变体TRIMCyp。与TRIM5α的限制活性不同,猕猴TRIMCyp融合蛋白能够抑制某些逆转录病毒如HIV-2的复制,但不能限制HIV-1的复制。因此TRIMCyp基因型猕猴对HIV-1的易感性可能明显高于TRIM5α个体,这为HIV-1的跨种传播感染感染提供了有利条件。本研究分病毒改造、易感宿主筛选和病毒功能研究3部分进行。为去除猕猴TRIM5α和APOBEC3G的限制性作用靶点,对HIV-1进行了分子改造,分别或同时用SIV CA和Vif替换了HIV-1的相应基因后,构建了一系列重组HIV-1,命名为HSIV(human-simian immunodeficiency virus)。该系列病毒经一系列检测,表明具有生物学活性,证明重组病毒拯救成功。同时,对中国猕猴的TRIMCyp基因型存在和分布状况进行了调查。首次在中国品系的恒河猴中发现了TRIMCyp基因型的存在,并从食蟹猴中筛选出了TRIMCyp/TRIMCyp纯合子个体,以用于HSIV的跨种传播感染研究。体外的细胞感染实验结果表明,所构建的重组HSIV能够成功克服猕猴TRIMCyp蛋白和APOBEC3G蛋白的限制,进行有效的复制,为进一步的细胞感染实验以及猴体跨种传播感染实验提供了基础。此外,本论文亦为研究逆转录病毒跨种间感染和宿主的抗病毒固有免疫提供了重要的实验论据。
[Abstract]:AIDS research still lacks suitable animal models. Non-human primate models are thought to be ideal models for studying HIV-1 infection and body response, but the presence of multiple genetic factors in these animals limits the interspecies transmission of HIV-1. The application of the model is limited. In recent years, two key HIV-1 limiting factors, TRIM5 伪 and APOBEC3G., have been found in non-human primates. Monkey TRIM5 伪 can restrict the replication of HIV-1 capsid protein (CA), but can not limit the replication of (SIV). HIV-1Vif protein can effectively resist the restricted activity of human APOBEC3G, but not in monkey APOBEC3G. But SIV Vif can effectively resist the restricted function of monkey APOBEC3G. Therefore, by replacing the corresponding genes of HIV-1 with SIV CA and Vif genes, the interspecies transmission of HIV-1 to monkey may be realized. On the other hand, a CypA gene of TRIM5 伪 was found in rhesus monkeys of some Old World monkeys and inserted into splicing mutant TRIMCyp.. In contrast to the limited activity of TRIM5 伪, the TRIMCyp fusion protein of rhesus monkeys can inhibit the replication of some retroviruses such as HIV-2, but cannot restrict the replication of HIV-1. Therefore, the susceptibility of rhesus monkeys with TRIMCyp genotype to HIV-1 may be significantly higher than that of TRIM5 伪 individuals, which provides favorable conditions for the interspecies transmission of HIV-1. This study is divided into three parts: virus modification, susceptible host screening and virus function study. In order to remove the restrictive targets of TRIM5 伪 and APOBEC3G in rhesus monkeys, HIV-1 was modified by molecular modification. After the corresponding genes of HIV-1 were replaced with SIV CA and Vif respectively, a series of recombinant HIV-1, named HSIV (human-simian immunodeficiency virus).) were constructed. After a series of tests, this series of viruses showed biological activity and proved that the recombinant virus was successfully saved. At the same time, the existence and distribution of TRIMCyp genotypes in Chinese rhesus monkeys were investigated. TRIMCyp genotypes were first found in Chinese rhesus monkeys, and TRIMCyp/TRIMCyp homozygous individuals were screened from crab-eating monkeys for cross-species transmission of HSIV. The results of cell infection in vitro showed that the constructed recombinant HSIV could successfully overcome the limitations of TRIMCyp and APOBEC3G proteins in rhesus monkeys and carry out effective replication, which provided a basis for further experiments of cell infection and transmissible infection in monkeys. In addition, this paper also provides important experimental evidence for the study of retrovirus interspecific infection and host innate immunity.
【学位授予单位】：中国农业科学院
【学位级别】：博士
【学位授予年份】：2010
【分类号】：R-332

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