A rapidly reversible chemical dimerizer system to study lipid signaling in living cells.
Chemical dimerizers are powerful tools for non-invasive manipulation of enzyme activities in intact cells. Here we introduce the first rapidly reversible small-molecule-based dimerization system and demonstrate a sufficiently fast switch-off to determine kinetics of lipid metabolizing enzymes in living cells. We applied this new method to induce and stop phosphatidylinositol 3-kinase (PI3K) activity, allowing us to quantitatively measure the turnover of phosphatidylinositol 3,4,5-trisphosphate (PIP3) and its downstream effectors by confocal fluorescence microscopy as well as standard biochemical methods.
Cell Biology & Biophysics Unit, European Molecular Biology Laboratory, Meyerhofstrasse 1, 69117 Heidelberg (Germany).
Suihan
S
Feng
Vibor
V
Laketa
Frank
F
Stein
Anna
A
Rutkowska
Aidan
A
MacNamara
Sofia
S
Depner
Ursula
U
Klingmüller
Julio
J
Saez-Rodriguez
Carsten
C
Schultz
D019281:Dimerization; D006367:HeLa Cells; D006801:Humans; D007700:Kinetics; D019869:Phosphatidylinositol 3-Kinases / Q000378:metabolism*; D018129:Phosphatidylinositol Phosphates / Q000378:metabolism*; D010766:Phosphorylation; D051057:Proto-Oncogene Proteins c-akt / Q000378:metabolism; D015398:Signal Transduction / Q000187:drug effects; D016559:Tacrolimus / Q000494:pharmacology
D018129:Phosphatidylinositol Phosphates; C060974:phosphatidylinositol 3,4,5-triphosphate; D019869:Phosphatidylinositol 3-Kinases; D051057:Proto-Oncogene Proteins c-akt; D016559:Tacrolimus
FKBP; PI3K; PIP3; SNAP; reversible chemical dimerizer