Mito-docking: A Novel In Vivo Method to Detect Protein–Protein Interactions

Many methods have been developed to detect protein–protein interactions (PPIs) and explore cellular processes. However, effective methods for detecting complicated PPIs under physical conditions are still in demand. Here, a simple and efficient mitochondria-docking (Mito-docking) method for PPI detection in vivo is developed. The strategy is to anchor a “bait” protein to mitochondrial outer membrane (MOM), and then trap the “prey” protein onto MOM. In this way, interacting signals are enriched to allow easy detection. This method efficiently detects the well-known interaction between two G protein subunits (G_γ2 with G_β1) and is successfully applied to investigate the recognition of importin α superfamily members for the classical nuclear localization signal (NLS) of simian virus 40 (SV40) large T antigen, which are highly dynamic and not easily visualized by conventional methods. As far as is known, this is the first time that the interaction between human importin α receptors with NLSs has been visualized. The results prove that Mito-docking can be used as a simple, straightforward, and intuitive method to study PPIs qualitatively and quantitatively in vivo.