Conditional fast expression and function of multimeric TRPV5 channels using Shield-1

Schoeber JP, van de Graaf SF, Lee KP, Wittgen HG, Hoenderop JG, Bindels RJ. Conditional fast expression and function of multimeric TRPV5 channels using Shield-1. Am J Physiol Renal Physiol 296: F204-F211, 2009. First published October 8, 2008; doi: 10.1152/ajprenal.90473.2008.-A recently described novel controllable method to regulate protein expression is based on a mutated FK506-binding protein-12 (mtFKBP) that is unstable and rapidly degraded in mammalian cells. This instability can be conferred to other proteins directly fused to mtFKBP. Binding of a synthetic cell-permeant ligand (Shield-1) to mtFKBP reverses the instability, allowing conditional expression of mtFKBP-fused proteins. We adapted this strategy to study multimeric plasma membrane proteins using the ion channel TRPV5 as model protein. mtFKBP-TRPV5 forms functional ion channels and its expression can be controlled in a time- and dose-dependent fashion using Shield-1. Moreover, in the presence of Shield-1, mtFKBP-TRPV5 formed heteromultimeric channels with untagged TRPV5, which were codegraded upon washout of Shield-1, providing a strategy to study multimeric plasma membrane protein complexes without the need to destabilize all individual subunits.