Conditional fast expression and function of multimeric TRPV5 channels using Shield-1.
until further notice
SourceAmerican Journal of Physiology : Renal Physiology, 296, 1, (2009), pp. F204-11
Article / Letter to editor
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American Journal of Physiology : Renal Physiology
SubjectIGMD 9: Renal disorder; NCMLS 5: Membrane transport and intracellular motility
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.
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