Luker Gary D, Sharma Vijay, Pica Christina M, Dahlheimer Julie L, Li Wei, Ochesky Joseph, Ryan Christine E, Piwnica-Worms Helen, Piwnica-Worms David
Molecular Imaging Center, Mallinckrodt Institute of Radiology and Department of Molecular Biology, Washington University School of Medicine, St. Louis, MO 63110, USA.
Proc Natl Acad Sci U S A. 2002 May 14;99(10):6961-6. doi: 10.1073/pnas.092022399. Epub 2002 May 7.
Protein-protein interactions control transcription, cell division, and cell proliferation as well as mediate signal transduction, oncogenic transformation, and regulation of cell death. Although a variety of methods have been used to investigate protein interactions in vitro and in cultured cells, none can analyze these interactions in intact, living animals. To enable noninvasive molecular imaging of protein-protein interactions in vivo by positron-emission tomography and fluorescence imaging, we engineered a fusion reporter gene comprising a mutant herpes simplex virus 1 thymidine kinase and green fluorescent protein for readout of a tetracycline-inducible, two-hybrid system in vivo. By using micro-positron-emission tomography, interactions between p53 tumor suppressor and the large T antigen of simian virus 40 were visualized in tumor xenografts of HeLa cells stably transfected with the imaging constructs. Imaging protein-binding partners in vivo will enable functional proteomics in whole animals and provide a tool for screening compounds targeted to specific protein-protein interactions in living animals.
蛋白质-蛋白质相互作用控制着转录、细胞分裂和细胞增殖，同时介导信号转导、致癌转化以及细胞死亡的调控。尽管已经使用了多种方法来体外和培养细胞中研究蛋白质相互作用，但没有一种方法能够在完整的活体动物中分析这些相互作用。为了通过正电子发射断层扫描和荧光成像在体内对蛋白质-蛋白质相互作用进行无创分子成像，我们设计了一个融合报告基因，其包含突变的单纯疱疹病毒1胸苷激酶和绿色荧光蛋白，用于在体内读出四环素诱导的双杂交系统。通过使用微型正电子发射断层扫描，在稳定转染了成像构建体的HeLa细胞的肿瘤异种移植中可视化了p53肿瘤抑制因子与猿猴病毒40大T抗原之间的相互作用。在体内对蛋白质结合伴侣进行成像将实现全动物的功能蛋白质组学，并为筛选针对活体动物中特定蛋白质-蛋白质相互作用的化合物提供一种工具。
