Label-free cell phenotypic profiling decodes the composition and signaling of an endogenous ATP-sensitive potassium channel

• DOI: 10.1038/srep04934
• Publication Date: 2014-05-12T09:06:57Z
• Authors: Haiyan Sun; Ying Wei; Huayun Deng; Qiaojie Xiong; Min Li; Joydeep Lahiri; Ye Fang
• Keywords: 0301 basic medicine; 572; Drug Evaluation, Preclinical; Gene Expression; Ligands; Article; Cell Line; Membrane Potentials; 03 medical and health sciences; KATP Channels; cell signaling; Cluster Analysis; Humans; RNA, Messenger; Protein Kinase Inhibitors; rho-Associated Kinases; 0303 health sciences; Pinacidil; Janus Kinase 3; Janus Kinase 2; Actins; Phenotype; Gene Knockdown Techniques; ion channel; RNA Interference; Protein Kinases; signal transduction; Signal Transduction

AbstractCurrent technologies for studying ion channels are fundamentally limited because of their inability to functionally link ion channel activity to cellular pathways. Herein, we report the use of label-free cell phenotypic profiling to decode the composition and signaling of an endogenous ATP-sensitive potassium ion channel (KATP) in HepG2C3A, a hepatocellular carcinoma cell line. Label-free cell phenotypic agonist profiling showed that pinacidil triggered characteristically similar dynamic mass redistribution (DMR) signals in A431, A549, HT29 and HepG2C3A, but not in HepG2 cells. Reverse transcriptase PCR, RNAi knockdown and KATPblocker profiling showed that the pinacidil DMR is due to the activation of SUR2/Kir6.2 KATPchannels in HepG2C3A cells. Kinase inhibition and RNAi knockdown showed that the pinacidil activated KATPchannels trigger signaling through Rho kinase and Janus kinase-3 and cause actin remodeling. The results are the first demonstration of a label-free methodology to characterize the composition and signaling of an endogenous ATP-sensitive potassium ion channel.