RT Journal Article
SR Electronic
T1 Discovery of Regulators of Receptor Internalization with High-Throughput Flow Cytometry
JF Molecular Pharmacology
JO Mol Pharmacol
FD American Society for Pharmacology and Experimental Therapeutics
SP 645
OP 657
DO 10.1124/mol.112.079897
VO 82
IS 4
A1 Yang Wu
A1 Phillip H. Tapia
A1 Gregory W. Fisher
A1 Peter C. Simons
A1 J. Jacob Strouse
A1 Terry Foutz
A1 Alan S. Waggoner
A1 Jonathan Jarvik
A1 Larry A. Sklar
YR 2012
UL http://molpharm.aspetjournals.org/content/82/4/645.abstract
AB We developed a platform combining fluorogen-activating protein (FAP) technology with high-throughput flow cytometry to detect real-time protein trafficking to and from the plasma membrane in living cells. The hybrid platform facilitates drug discovery for trafficking receptors such as G protein-coupled receptors and was validated with the β2-adrenergic receptor (β2AR) system. When a chemical library containing ∼1200 off-patent drugs was screened against cells expressing FAP-tagged β2ARs, all 33 known β2AR-active ligands in the library were successfully identified, together with a number of compounds that might regulate receptor internalization in a nontraditional manner. Results indicated that the platform identified ligands of target proteins regardless of the associated signaling pathway; therefore, this approach presents opportunities to search for biased receptor modulators and is suitable for screening of multiplexed targets for improved efficiency. The results revealed that ligands may be biased with respect to the rate or duration of receptor internalization and that receptor internalization may be independent of activation of the mitogen-activated protein kinase pathway.