Generating reagents to explore the anitviral potential of TRIM family proteins

描述（由申请人提供）：TRIM蛋白家族介导了许多最近报道的针对不同病毒的抗病毒活性。这些活性中研究得最好的是TRIM51蛋白在感染早期通过结合病毒衣壳核心上的决定因素抑制某些逆转录病毒（包括HIV-1）复制的能力。最近的数据表明，其他TRIM家族蛋白可以在病毒复制周期的其他步骤影响HIV-1的复制。大量研究表明，TRIM家族蛋白也可以负性调节多种病毒的复制，包括腺病毒、疱疹病毒和流感病毒，尽管这些活动的特征尚不清楚。最近的研究发现，许多TRIM家族蛋白的表达在对干扰素治疗的反应中上调，这也表明TRIM家族蛋白是先天免疫系统的细胞内元件，可抑制细胞内病原体的复制。由于TRIM51倾向于在细胞中多聚成高阶结构，因此对该蛋白的生化分析异常困难。其他TRIM家族成员可能也是如此。我以前的研究利用定量荧光显微镜来可视化和量化TRIM51与细胞质中HIV-1病毒复合物之间发生的相互作用。该项目将开发逆转录病毒载体，诱导表位标记的TRIM家族成员的表达，并利用这些载体产生稳定表达这些蛋白质的细胞系。这些载体和细胞系将有助于鉴定具有抗病毒活性的TRIM蛋白，并揭示病毒与TRIM家族蛋白之间在细胞中发生的新的相互作用。一旦确定，这些相同的载体和细胞系将允许可视化TRIM蛋白与病毒蛋白和/或受TRIM活性影响的遗传元件之间发生的相互作用。这将为我的实验室催化新的研究领域，并增加我们对TRIM家族蛋白抗病毒活性的理解。

DESCRIPTION (provided by applicant):The TRIM family of proteins mediates a number of recently reported antiviral activities mediated against diverse viruses. The best studied of these activities is the ability of the TRIM51 protein to inhibit the replication of certain retroviruses, including HIV-1, by binding determinants present on the viral capsid core during early stages of infection. Recent data suggests that other TRIM family proteins can affect HIV-1 replication at other steps in the viral replication cycle. Numerous studies suggest that TRIM family proteins can also negatively regulate the replication of a diverse range of viruses, including adenoviruses, herpesviruses and influenza viruses, although these activities remain poorly characterized. The recent finding that the expression of many TRIM family proteins are upregulated in response to interferon treatment also indicates that the TRIM family proteins represent an intracellular element of the innate immune system, acting to inhibit the replication of intracellular pathogens. Because of the tendency of TRIM51 to multimerize into high-order structures in cells, biochemical analysis of this protein has been unusually difficult. This will likely be true of other TRIM family members. My previous studies have utilized quantitative fluorescent microscopy to visualize and quantify the interactions occurring between TRIM51 and HIV-1 viral complexes in the cytoplasm. This project would develop retroviral vectors that induce the expression of epitope tagged TRIM family members and use these vectors to generate cell lines stably expressing these proteins. These vectors and cell lines will be useful in identifying TRIM proteins with antiviral activity against diverse viruses and expose novel interactions occurring in cells between viruses and TRIM family proteins. Once identified, these same vectors and cell lines will allow for the visualization of interactions occurring between TRIM proteins and viral proteins and/or genetic elements affected by TRIM activity. This will catalyze new areas of research for my lab and increase our understanding of the antiviral activity of TRIM family proteins.