Imaging protein interactions with bioluminescence resonance energy transfer (BRET) in plant and mammalian cells and tissues.

Xu X, Soutto M, Xie Q, Servick S, Subramanian C, von Arnim AG, Johnson CH
Proc Natl Acad Sci U S A. 2007 104 (24): 10264-9

PMID: 17551013 · PMCID: PMC1891211 · DOI:10.1073/pnas.0701987104

FRET is a well established method for cellular and subcellular imaging of protein interactions. However, FRET obligatorily necessitates fluorescence excitation with its concomitant problems of photobleaching, autofluorescence, phototoxicity, and undesirable stimulation of photobiological processes. A sister technique, bioluminescence resonance energy transfer (BRET), avoids these problems because it uses enzyme-catalyzed luminescence; however, BRET signals usually have been too dim to image effectively in the past. Using a new generation electron bombardment-charge-coupled device camera coupled to an image splitter, we demonstrate that BRET can be used to image protein interactions in plant and animal cells and in tissues; even subcellular imaging is possible. We have applied this technology to image two different protein interactions: (i) dimerization of the developmental regulator, COP1, in plant seedlings; and (ii) CCAAT/enhancer binding protein alpha (C/EBPalpha) in the mammalian nucleus. This advance heralds a host of applications for imaging without fluorescent excitation and its consequent limitations.

MeSH Terms (23)

Animals Arabidopsis Arabidopsis Proteins CCAAT-Enhancer-Binding Proteins Cell Culture Techniques Cell Line Cell Nucleus Cells, Cultured Dimerization Fluorescence Resonance Energy Transfer Humans Luminescent Measurements Luminescent Proteins Mice Microscopy, Fluorescence Pituitary Gland Plant Proteins Protein Binding Proteins Recombinant Fusion Proteins Seedlings Spectrometry, Fluorescence Tobacco

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