NDLI: Visualization of AP-1–NF-κB ternary complexes in living cells by using a BiFC-based FRET

Content Provider	PubMed Central
Author	Shyu, Y. John Suarez, Christopher D. Hu, Chang-deng
Copyright Year	2008
Abstract	Protein–protein interactions are essential for maintaining cell structure and for executing almost all cellular processes. Determination of where and how each protein interacts with its partners provides significant insight into proteins' cellular roles. Although several assays, such as FRET and bimolecular fluorescence complementation (BiFC), have been developed and widely used for visualization and identification of protein interactions in living cells, there is no simple and convenient assay to visualize and identify multiple protein complexes in living cells. Because many signaling molecules often function as ternary complexes, availability of an assay for visualization and identification of ternary complexes will significantly expand the repertoire of protein interaction studies in living cells. By using the Fos–Jun–nuclear factor of activated T cells (NFAT) ternary complex as a model and the fluorescent proteins Cerulean and Venus, two mutant proteins of CFP and YFP with better folding and less environment sensitivity, as a donor and acceptor, respectively, we have combined a Venus-based BiFC system with Cerulean to develop a BiFC-based FRET (BiFC-FRET) assay for visualization of ternary complexes in living cells with a conventional three-filter FRET setup. We also have applied the BiFC-FRET to identify a ternary complex formed between Fos–Jun heterodimers and the NF-κB subunit, p65. This finding reveals a cross-talk between AP-1 and NF-κB. Thus, the BiFC-FRET represents a convenient assay for identification and visualization of ternary complexes in living cells.
Related Links	http://dx.doi.org/10.1073/pnas.0705181105
Ending Page	156
Page Count	6
Starting Page	151
File Format	PDF
ISSN	00278424
e-ISSN	10916490
Journal	Proceedings of the National Academy of Sciences of the United States of America
Issue Number	1
Volume Number	105
Language	English
Publisher	National Academy of Sciences
Publisher Date	2008-01-08
Access Restriction	Open
Rights Holder	National Academy of Sciences
Subject Keyword	General Research in Higher Education
Content Type	Text
Resource Type	Article
Subject	Multidisciplinary

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