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Exploration of the Peptide Recognition of an Amiloride-sensitive FMRFamide Peptide-gated Sodium Channel.

ABSTRACT: FMRFamide (Phe-Met-Arg-Phe-NH2)-activated sodium channel (FaNaC) is an amiloride-sensitive sodium channel activated by endogenous tetrapeptide in invertebrates, and belongs to the epithelial sodium channel/degenerin (ENaC/DEG) superfamily. The ENaC/DEG superfamily differs markedly in its means of activation, such as spontaneously opening or gating by mechanical stimuli or tissue acidosis. Recently, it has been observed that a number of ENaC/DEG channels can be activated by small molecules or peptides, indicating that the ligand-gating may be an important feature of this superfamily. The peptide ligand control of the channel gating might be an ancient ligand-gating feature in this superfamily. Therefore, studying the peptide recognition of FaNaC channels would advance our understanding of the ligand-gating properties of this superfamily of ion channels. Here we demonstrate that Tyr-131, Asn-134, Asp-154, and Ile-160, located in the putative upper finger domain ofHelix aspersaFaNaC (HaFaNaC) channels, are key residues for peptide recognition of this ion channel. Two HaFaNaC specific-insertion motifs among the ENaC/DEG superfamily, residing at the putative ?4-?5 linker of the upper thumb domain and the ?6-?7 linker of the upper knuckle domain, are also essential for the peptide recognition of HaFaNaC channels. Chemical modifications and double mutant cycle analysis further indicated that those two specific inserts and key residues in the upper finger domain together participate in peptide recognition of HaFaNaC channels. This ligand recognition site is distinct from that of acid-sensing ion channels (ASICs) by a longer distance between the recognition site and the channel gate, carrying useful information about the ligand gating and the evolution of the trimeric ENaC/DEG superfamily of ion channels.

SUBMITTER: Niu YY

PROVIDER: S-EPMC4817185 | biostudies-literature | 2016 Apr

REPOSITORIES: biostudies-literature

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Exploration of the Peptide Recognition of an Amiloride-sensitive FMRFamide Peptide-gated Sodium Channel.

Niu You-Ya YY Yang Yang Y Liu Yan Y Huang Li-Dong LD Yang Xiao-Na XN Fan Ying-Zhe YZ Cheng Xiao-Yang XY Cao Peng P Hu You-Min YM Li Lingyong L Lu Xiang-Yang XY Tian Yun Y Yu Ye Y

The Journal of biological chemistry 20160211 14

FMRFamide (Phe-Met-Arg-Phe-NH2)-activated sodium channel (FaNaC) is an amiloride-sensitive sodium channel activated by endogenous tetrapeptide in invertebrates, and belongs to the epithelial sodium channel/degenerin (ENaC/DEG) superfamily. The ENaC/DEG superfamily differs markedly in its means of activation, such as spontaneously opening or gating by mechanical stimuli or tissue acidosis. Recently, it has been observed that a number of ENaC/DEG channels can be activated by small molecules or pep ...[more]

PMID: 26867576

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Project description:Considerable effort has been directed toward the development of methods to selectively activate specific subtypes of neurons. Focus has been placed on the heterologous expression of proteins that are capable of exciting neurons in which they are expressed. Here we describe the heterologous expression of the invertebrate FMRFamide (H-phenylalanine-methionine-arginine-phenylalanine-NH2) -gated sodium channel from Helix aspersa (HaFaNaC) in hippocampal slice cultures. HaFaNaC was co-expressed with a fluorescent protein (green fluorescent protein (GFP), red fluorescent protein from Discosoma sp (dsRed) or mutated form of red fluorescent protein from Discosoma sp (tdTomato)) in CA3 pyramidal neurons of rat hippocampal slice cultures using single cell electroporation. Pressure application of the agonist FMRFamide to HaFaNaC-expressing neuronal somata produced large prolonged depolarizations and bursts of action potentials (APs). FMRFamide responses were inhibited by amiloride (100 microM). In contrast, pressure application of FMRFamide to the axons of neurons expressing HaFaNaC produced no response. Fusion of GFP to the N-terminus of HaFaNaC showed that GFP-HaFaNaC was absent from axons. Bath application of FMRFamide produced persistent AP firing in HaFaNaC-expressing neurons. This FMRFamide-induced increase in the frequency of APs was dose-dependent. The concentrations of FMRFamide required to activate HaFaNaC-expressing neurons were below that required to activate the homologous acid sensing ion channel normally found in mammalian neurons. Furthermore, the mammalian neuropeptides neuropeptide FF and RFamide-related peptide-1, which have amidated RF C-termini, did not affect HaFaNaC-expressing neurons. Antagonists of NPFF receptors (BIBP3226) also had no effect on HaFaNaC. Therefore, we suggest that heterologous-expression of HaFaNaC in mammalian neurons could be a useful method to selectively and persistently excite specific subtypes of neurons in intact nervous tissue.

Dataset Information

Exploration of the Peptide Recognition of an Amiloride-sensitive FMRFamide Peptide-gated Sodium Channel.

Publications

Exploration of the Peptide Recognition of an Amiloride-sensitive FMRFamide Peptide-gated Sodium Channel.

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