Abstract
Here, we report a method to regulate cellular protein levels by introducing a ubiquitin variant between a destabilizing domain (DD) and the regulated protein. When produced in the absence of a stabilizing ligand the DD dominates and the entire fusion protein is processively degraded by the proteasome. In the presence of the stabilizing ligand the fusion protein is metabolically stable and becomes a substrate for abundant ubiquitin-specific proteases, liberating a native, or a near-native protein-of-interest. This technique is thus particularly useful for the study of proteins whose free N terminus is required for proper function. In addition, removal of the DD in the presence of stabilizing ligand leads to higher expression levels of regulated protein when cells experience transient exposure to a stabilizing ligand, such as in a living animal receiving a single dose of a pharmacological agent as the stabilizing ligand.