Mutations in the palm domain disrupt modulation of acid-sensing ion channel 1a currents by neuropeptides

Benoîte Bargeton,Stephan Kellenberger,Justyna Iwaszkiewicz,Sophie Roy,Gaetano Bonifacio,Vincent Zoete

doi:10.1038/s41598-018-37426-5

Benoîte Bargeton, Stephan Kellenberger + Show 4 more

Open Access

https://doi.org/10.1038/s41598-018-37426-5

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Abstract

Modulation by neuropeptides enhances several functions of acid-sensing ion channels (ASICs), such as pain sensation and acid-induced neuronal injury. The acid-induced opening of ASICs is transient, because of a rapid desensitization. Neuropeptides containing an Arg-Phe-amide motif affect ASIC desensitization and allow continuous activity of ASICs. In spite of the importance of the sustained ASIC activity during prolonged acidification, the molecular mechanisms of ASIC modulation by neuropeptides is only poorly understood. To identify the FRRFa (Phe-Arg-Arg-Phe-amide) binding site on ASIC1a, we carried out an in silico docking analysis and verified functionally the docking predictions. The docking experiments indicated three possible binding pockets, located (1) in the acidic pocket between the thumb, finger, β-ball and palm domains, (2) in a pocket at the bottom of the thumb domain, and (3) in the central vestibule along with the connected side cavities. Functional measurements of mutant ASIC1a confirmed the importance of residues of the lower palm, which encloses the central vestibule and its side cavities, for the FRRFa effects. The combined docking and functional experiments strongly suggest that FRRFa binds to the central vestibule and its side cavities to change ASIC desensitization.

Highlights

Acid-sensing ion channels (ASICs) are voltage-independent Na+ channels that are activated by extracellular acidification[1,2,3]
When discussing here the docking results, we present briefly the docking results on the desensitized or open ASIC1a model to illustrate the basis of the mutagenesis strategy, and in more detail the results obtained with the acid-sensing ion channels (ASICs) model in the closed conformation
Previous observations resulted in hypotheses about the sites of action of RFa peptides in ASIC1a

Summary

Introduction

Acid-sensing ion channels (ASICs) are voltage-independent Na+ channels that are activated by extracellular acidification[1,2,3]. Extracellular pH changes affect the protonation status of several amino acid residues located in different domains, such as the acidic pocket, the palm and the wrist, contributing thereby to activation and desensitization of the channel[18,21,22,23,24,25]. The palm domain undergoes large conformational changes during desensitization; its integrity is required for a normal, complete desensitization[16,22,25,26,27] It is not known where in the ASICs the RFa peptides bind. Structure-function studies showed that residues in the thumb and the palm co-determine the effect of RFa peptides[16,28], it is not clear whether these domains participate in the formation of the binding site or whether they are important for transducing binding into functional effects. Mutagenesis of residues being part of predicted binding sites, followed by functional analysis, demonstrated impairment of FRRFa effects in ASICs carrying mutations in the central vestibule or its side cavities, strongly suggesting this as the binding site for FRRFa

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Results

Conclusion