Functional and Pharmacological Characterization of C. elegans DEG/ENaC/ASIC Channels

Abstract

Activating or blocking ion channels is a central goal of drug discovery. However, ion channel families have multiple variants which differ in function. Additionally, channel subunits assemble to form multimeric complexes with varying biophysical properties and drug sensitivities. Degenerin (DEG)/epithelial sodium channel (ENaC)/acid-sensing ion channel (ASIC) proteins arebelieved to assemble as homomeric or heteromeric trimers into non-voltage gated, sodium-selective, and amiloride sensitive ion channels. Here, we analyzed a subset of the 30 DEG/ENaC/ASIC proteins encoded by the C. elegans genome in order to test the hypothesis that individual family members have diverse properties. Specifically, we expressed five C. elegans isoforms in Xenopus laevis oocytes (DEGT-1d, DEL-1d, UNC-8d, MEC-10d and MEC-4d) and measured current amplitude, selectivity among monovalent cations, sensitivity to amiloride and its analogs, and sensitivity to nonsteroidal anti-inflammatory drugs (NSAIDs). Overall, we find substantial variations in the pharmacological footprints of the two drug families we studied (amiloride, NSAIDs) and among the DEG/ENaC/ASIC proteins. DEGT-1d is the first channel in this family demonstrated to be equally permeable to sodium and potassium, and has no response to amiloride and its analogs. We identified at least one NSAID that blocked DEGT-1d and UNC-8d currents. Surprisingly, MEC4d currents were potentiated by five NSAIDs. A mutation homologous to ASIC1a[K422], MEC4[E704K], decreased the sensitivity to NSAIDs. The molecular origin of this potentiation will be the focus of future studies, along with identifying which structural elements are responsible for inhibition and potentiation. This work will help to advance drug design for this family and enhance our understanding of how NSAIDs exert their function. Collectively, these data establish that DEG/ENaC/ASIC ion channel proteins vary in their ion permeability and drug sensitivity. Work supported by DFG (SF), SSRP (ID), R01NS047715, R35NS129879 (MBG)