Visualizing protein–protein interactions in plants by rapamycin-dependent delocalization

• DOI: 10.1093/plcell/koab004
• Publication Date: 2021-01-08T18:50:39Z
• Authors: Joanna Winkler; Evelien Mylle; Andreas De Meyer; Benjamin Pavie; Julie Merchie; Peter Grones; Dani�l Van Damme
• Keywords: Nicotiana; Biochemistry & Molecular Biology; Green Fluorescent Proteins; Plant Science; Tacrolimus Binding Proteins; Plant Cells; REVEALS; Protein Interaction Mapping; LIVING CELLS; Plant Proteins; Sirolimus; Science & Technology; Plant Sciences; Biology and Life Sciences; LOCALIZATION; Cell Biology; ARABIDOPSIS; Recombinant Proteins; TRANSLOCATION; Mitochondria; Plant Leaves; Luminescent Proteins; CYTOKINESIS; ESTABLISHMENT; GROWTH; VECTORS; Protein Multimerization; Life Sciences & Biomedicine; SYSTEM; Red Fluorescent Protein

AbstractIdentifying protein–protein interactions (PPIs) is crucial for understanding biological processes. Many PPI tools are available, yet only some function within the context of a plant cell. Narrowing down even further, only a few tools allow complex multi-protein interactions to be visualized. Here, we present a conditional in vivo PPI tool for plant research that meets these criteria. Knocksideways in plants (KSP) is based on the ability of rapamycin to alter the localization of a bait protein and its interactors via the heterodimerization of FKBP and FRB domains. KSP is inherently free from many limitations of other PPI systems. This in vivo tool does not require spatial proximity of the bait and prey fluorophores and it is compatible with a broad range of fluorophores. KSP is also a conditional tool and therefore the visualization of the proteins in the absence of rapamycin acts as an internal control. We used KSP to confirm previously identified interactions in Nicotiana benthamiana leaf epidermal cells. Furthermore, the scripts that we generated allow the interactions to be quantified at high throughput. Finally, we demonstrate that KSP can easily be used to visualize complex multi-protein interactions. KSP is therefore a versatile tool with unique characteristics and applications that complements other plant PPI methods.