Noninvasive quantitative imaging of protein–protein interactions in living subjects

doi:10.1073/pnas.052710999

Noninvasive quantitative imaging of protein–protein interactions in living subjects

^*The Crump Institute for Molecular Imaging, ^†Department of Molecular and Medical Pharmacology, ^§University of California at Los Angeles–Jonsson Comprehensive Cancer Center, and ^‡Department of Biomathematics, University of California Los Angeles School of Medicine, Los Angeles, CA 90095

Contributed by M. E. Phelps

Abstract

We are developing methods to image molecular and cellular events in living subjects. In this study, we validate imaging of protein—protein interactions in living mice by using bioluminescent optical imaging. We use the well studied yeast two-hybrid system adapted for mammalian cells and modify it to be inducible. We employ the NF-κB promoter to drive expression of two fusion proteins (VP16-MyoD and GAL4-ID). We modulate the NF-κB promoter through tumor necrosis factor α. Firefly luciferase reporter gene expression is driven by the interaction of MyoD and ID through a transcriptional activation strategy. We demonstrate the ability to detect this induced protein–protein interaction in cell culture and image this induced interaction in living mice by using transiently transfected cells. The current approach will be a valuable and potentially generalizable tool to noninvasively and quantitatively image protein–protein interactions in living subjects. The approaches validated should have important implications for the study of protein–protein interactions in cells maintained in their natural in vivo environment as well as for the in vivo evaluation of new pharmaceuticals targeted to modulate protein–protein interactions.

Footnotes

↵ ¶ To whom reprint requests should be addressed at: Crump Institute for Molecular Imaging, University of California at Los Angeles School of Medicine, B3-399A BRI 700 Westwood Plaza, Los Angeles, CA 90095-1770. E-mail: sgambhir{at}mednet.ucla.edu.
↵ ‖ Tjuvajev, J. G., Avril, N., Safer, M., Joshi, R., Oku, T., Sasjima, T., Miyagawa, T., Beattie, B., Daghighian, F., Augenson, F., et al. (1997) J. Nucl. Med. 38, 239P (abstr.).
Abbreviations:

fl,

firefly luciferase reporter gene;

FL,

firefly luciferase enzyme/protein;

CCD,

charge-coupled device;

PET,

positron-emission tomography;

IY2H,

inducible yeast two-hybrid system;

CMV,

cytomegalovirus;

TNF-α,

tumor necrosis factor α