Modulation of Human Acid-Sensing Ion Channel 1A Open Channel Inactivation by FRRFamide

Abstract

The ASICs (Acid-Sensing Ion Channels) are involved in neuronal signaling in the central and peripheral nervous system. These non-voltage-gated channels are involved in learning, the expression of fear, neurodegeneration after ischemia, and pain sensation. The molecular bases underlying their activity are not yet fully understood. During an extracellular acidification, ASICs open transiently before inactivating in the continued presence of the low extracellular pH. Modulators of ASIC inactivation may contribute to the physiological and pathological functions of ASICs. FRRFamide (FRRFa) and related peptides have been shown to slow the ASIC inactivation time course and to induce a small sustained current. In the present study we have carried out in silico docking of FRRFa to human ASIC1a, which predicted two cavities as FRRFa binding site. It has previously been shown that a part of the thumb region differs between ASIC1 orthologs, and that FRRFa induces greater sustained currents in human than in mouse ASIC1a, suggesting that this region may be involved in the effect of FRRFa.The role of the residues predicted to be part of the FRRFa binding site (one of the top docking poses) has been tested by site-directed mutagenesis and functional studies. While mutation of only one residue of the predicted binding site decreased the effect of FRRFa, several point mutations in the β9-α4 region increased the FRRFa-induced sustained current. Our results indicate that the β9-α4 region is likely not the FRRFa binding site, however that it is involved in the effect of FRRFa on ASIC inactivation.