Visualization of interactions among bZIP and Rel family proteins in living cells using bimolecular fluorescence complementation.
Networks of protein interactions coordinate cellular functions. We describe a bimolecular fluorescence complementation (BiFC) assay for determination of the locations of protein interactions in living cells. This approach is based on complementation between two nonfluorescent fragments of the yellow fluorescent protein (YFP) when they are brought together by interactions between proteins fused to each fragment. BiFC analysis was used to investigate interactions among bZIP and Rel family transcription factors. Regions outside the bZIP domains determined the locations of bZIP protein interactions. The subcellular sites of protein interactions were regulated by signaling. Cross-family interactions between bZIP and Rel proteins affected their subcellular localization and modulated transcription activation. These results attest to the general applicability of the BiFC assay for studies of protein interactions.
Hu
Chang-Deng
CD
Howard Hughes Medical Institute and Department of Biological Chemistry, University of Michigan Medical School, Ann Arbor, MI 48109, USA.
Chinenov
Yurii
Y
Kerppola
Tom K
TK
D016475:3T3 Cells; D017398:Alternative Splicing; D000818:Animals; D001426:Bacterial Proteins / Q000737:chemistry*; D019556:COS Cells; D002451:Cell Compartmentation; D002522:Chlorocebus aethiops; D004268:DNA-Binding Proteins / Q000737:chemistry*; D019281:Dimerization; D005453:Fluorescence; D006367:HeLa Cells; D006801:Humans; D020672:I-kappa B Proteins; D016350:Leucine Zippers; D008025:Ligases / Q000737:chemistry*; D008164:Luminescent Proteins / Q000737:chemistry*; D046911:Macromolecular Substances; D051379:Mice; D008856:Microscopy, Fluorescence; D000072000:NF-KappaB Inhibitor alpha; D016328:NF-kappa B / Q000737:chemistry*; D010446:Peptide Fragments / Q000737:chemistry; D025941:Protein Interaction Mapping / Q000379:methods*; D017434:Protein Structure, Tertiary; D021381:Protein Transport; D016760:Proto-Oncogene Proteins c-fos / Q000737:chemistry*; D016755:Proto-Oncogene Proteins c-jun / Q000737:chemistry*; D011993:Recombinant Fusion Proteins / Q000737:chemistry; D017384:Sequence Deletion; D013347:Subcellular Fractions / Q000737:chemistry*; D014158:Transcription, Genetic; D014162:Transfection
D001426:Bacterial Proteins; D004268:DNA-Binding Proteins; D020672:I-kappa B Proteins; D008164:Luminescent Proteins; D046911:Macromolecular Substances; D016328:NF-kappa B; C000606204:NFKBIA protein, human; C000606205:Nfkbia protein, mouse; D010446:Peptide Fragments; D016760:Proto-Oncogene Proteins c-fos; D016755:Proto-Oncogene Proteins c-jun; D011993:Recombinant Fusion Proteins; C054562:yellow fluorescent protein, Bacteria; D000072000:NF-KappaB Inhibitor alpha; D008025:Ligases; C028812:guanosine 3',5'-polyphosphate synthetases