The M1 and pre-M1 segments contribute differently to ion selectivity in ASICs and ENaCs

Abstract

The ability to discriminate between different ionic species, termed ion selectivity, is a key feature of ion channels and forms the basis for their physiological function. Members of the degenerin/epithelial sodium channel (DEG/ENaC) superfamily of trimeric ion channels are typically sodium selective, but to a surprisingly variable degree. While acid-sensing ion channels (ASICs) are weakly sodium selective (sodium:potassium ratio ∼10:1), ENaCs show a remarkably high preference for sodium over potassium (>500:1). This discrepancy may be expected to originate from differences in the pore-lining second transmembrane segment (M2). However, these show a relatively high degree of sequence conservation between ASICs and ENaCs, and previous functional and structural studies could not unequivocally establish that differences in M2 alone can account for the disparate degrees of ion selectivity. By contrast, surprisingly little is known about the contributions of the first transmembrane segment (M1) and the preceding pre-M1 region. In this study, we used conventional and noncanonical amino acid–based mutagenesis in combination with a variety of electrophysiological approaches to show that the pre-M1 and M1 regions of mASIC1a channels are major determinants of ion selectivity. Mutational investigations of the corresponding regions in hENaC show that these regions contribute less to ion selectivity, despite affecting ion conductance. In conclusion, our work suggests that the remarkably different degrees of sodium selectivity in ASICs and ENaCs are achieved through different mechanisms. These results further highlight how M1 and pre-M1 are likely to differentially affect pore structure in these related channels.

Structural basis for excitatory neuropeptide signaling

“…Furthermore, we found that larger nitrogen-based cations, ammonium and methylammonium, were much less permeant than Na + in FaNaC1 (Fig. 4f), as observed for ENaC 33 but different from ASIC1, which conducts substantial ammonium and methtylammonium current 34,35 . This suggests that FaNaC1 adopts a narrower open-channel pore than ASIC1, as represented by our FMRFa-gated, diminazene-blocked structure, and that FaNaC1 presumably passes partly dehydrated Na + ions.…”

“…Nonetheless, a better resolved picture of ion conduction throughout the superfamily emerges from the comparison of FaNaC1 and distantly related ASIC1 pores. FaNaC1 is more closely related to ENaC 8,47,48 , both of these channels are poorly permeable to nitrogen-based cations larger than Na + 33 , and our FaNaC1/FMRFa/diminazene structure reveals an ion pathway slightly narrower than both ASIC1 and voltage-gated sodium channels 11,49 , both of which pass ammonium, methylammonium, and hydroxylamine relatively well 34,35,50 . Taken together, our capture of FaNaC1 in different functional states describes the mechanism by which FMRFa elicits excitatory neuronal signals and offers a template for future studies dissecting channel gating and ion conduction in DEG/ENaCs.…”

Single ion free energy calculation in ASIC1: the importance of the HG loop

“…2 The role of pre-TM1 and TM1 residues in ion selectivity have recently been shown, supporting the structural observations. 13…”