A General Method for Conditional Regulation of Protein Stability in Living Animals

INTRODUCTION The ability to rapidly and reversibly perturb protein levels in living animals is a powerful tool for researchers to determine protein function in complex systems. We recently designed a small protein domain based on the 12-kDa FKBP (FK506 binding protein) that can be fused at either the carboxyl or amino terminus of a protein of interest. This destabilization domain (DD) confers instability to fusion protein partners, allowing targeted degradation of the protein of interest. A small molecule called Shield-1 binds to the DD and protects the fusion protein from degradation. Small-molecule-mediated post-translational regulation of protein stability affords this system rapid, reversible, and tunable control of protein levels and functions in a variety of model systems. Theoretically, a number of transgene delivery methods (e.g., viral, liposomal, or stem cell) can be used for the analysis of a DD fusion protein in an animal model. This protocol uses tumor xenografts in mice as one such mechanism for delivering the fusion protein and presents a method for delivering Shield-1 to regulate the fusion proteins in vivo.