Mammalian two-hybrid system: A complementary approach to the yeast two-hybrid system
Mammalian two-hybrid system: A complementary approach to the yeast two-hybrid system
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DOI：
10.2144/97222pf02
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发表时间：
1997-02-01
期刊：
影响因子：
2.7
通讯作者：
Zhu, L
中科院分区：
文献类型：
--
作者：
Luo, Y;Batalao, A;Zhu, L
Here we demonstrate the use of a mammalian two-hybrid system to study protein-protein interactions. Like the yeast two-hybrid system, this is a genetic, in vivo assay based on the reconstitution of the function of a transcriptional activator. In this system, one protein of interest is expressed as a fusion to the Ga14 DNA-binding domain and another protein is expressed as a fusion to the activation domain of the VP16 protein of the herpes simplex virus. The vectors that express these fusion proteins are cotransfected with a reporter chloramphenicol acetyltransferase (CAT) vector into a mammalian cell line. The reporter plasmid contains a cat gene under the control of five consensus Ga14 binding sites. If the two fusion proteins interact there will be a significant increase in expression of the cat reporter gene. Previously, it was reported that mouse p53 antitumor protein and simian virus 40 large T antigen interact in a yeast two-hybrid system. Using a mammalian two-hybrid system, we were able to independently confirm this interaction. The mammalian two-hybrid system can be used as a complementary approach to verify protein-protein interactions detected by a yeast two-hybrid system screening. In addition, the mammalian two-hybrid system has two main advantages: (i) Assay results can be obtained within 48 h of transfection, and (ii) protein interactions in mammalian cells may better mimic actual in vivo interactions.
