Structural determinants of Acid-sensing ion channel potentiation by single chain lipids

Abstract

Acid-sensing ion channels (ASICs) are thought to be endogenous sensors of acidic pain in inflammatory pathways. It has previously been demonstrated that arachidonic acid (AA), a pain and inflammation promoting molecule, potentiates ASICs. However, a mechanistic understanding of how AA regulates ASICs is lacking. Furthermore, little is known regarding modulation by other polyunsaturated fatty acids (PUFAs). Here we show that PUFAs stabilize the open state of the channel by shifting the pH dependence of activation to more alkaline values, increasing max conductance, and slowing channel desensitization. We examine the effects of 35 PUFAs/PUFA derivatives and show that ASICs can be more strongly potentiated by these lipids than was originally seen for AA. In fact, arachidonoyl glycine (AG) can act as a ligand and activate the channel in the absence of acidic pH. We find that the strength of potentiation is critically dependent upon a negatively charged PUFA head group as well as both the length and the number of doubles bonds in the acyl tail. PUFA-induced shifts in the pH dependence of activation could be eliminated upon mutation of a highly conserved, positively charged arginine in the outer segment of TM1 (R64). Combined our results suggest a hypothesis whereby an electrostatic interaction between the charged PUFA head group and the positively charged arginine side chain potentiates ASIC currents by stabilizing the open state of the channel. This work uncovers a novel putative lipid binding site on ASICs and provides the structural basis for future development of compounds targeting ASICs.