Structural Domains Underlying the Activation of Acid-Sensing Ion Channel 2a

Schuhmacher, Laura-Nadine; Srivats, Shyam; Smith, Ewan St. John

doi:10.1124/mol.114.096909

Cited by 23 publications

(36 citation statements)

References 35 publications

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“…4D). For ASIC2, it was shown that regions outside the AcP, mostly the first ∼90 residues after the TM1, are critical for channel function (29,30). These observations are consistent with an important regulatory, but not essential, role of the AcP in ASIC activation.…”

Section: Discussionsupporting

confidence: 74%

Conformational dynamics and role of the acidic pocket in ASIC pH-dependent gating

Vullo

Bonifacio

Roy

et al. 2017

Proc. Natl. Acad. Sci. U.S.A.

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Acid-sensing ion channels (ASICs) are proton-activated Na + channels expressed in the nervous system, where they are involved in learning, fear behaviors, neurodegeneration, and pain sensation. In this work, we study the role in pH sensing of two regions of the ectodomain enriched in acidic residues: the acidic pocket, which faces the outside of the protein and is the binding site of several animal toxins, and the palm, a central channel domain. Using voltage clamp fluorometry, we find that the acidic pocket undergoes conformational changes during both activation and desensitization. Concurrently, we find that, although proton sensing in the acidic pocket is not required for channel function, it does contribute to both activation and desensitization. Furthermore, protonationmimicking mutations of acidic residues in the palm induce a dramatic acceleration of desensitization followed by the appearance of a sustained current. In summary, this work describes the roles of potential pH sensors in two extracellular domains, and it proposes a model of acidification-induced conformational changes occurring in the acidic pocket of ASIC1a.

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Section: Discussionsupporting

confidence: 74%

Conformational dynamics and role of the acidic pocket in ASIC pH-dependent gating

Vullo

Bonifacio

Roy

et al. 2017

Proc. Natl. Acad. Sci. U.S.A.

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“…Of the 3 carboxylate pairs contained within the acidic pocket identified from the crystal structure of cASIC1 that were proposed to be critical for proton activation of ASICs (D238-D350, E239-D346, and E220-D408; cASIC1a numbering) (2), D238 is replaced by glutamate and D346 is actually replaced by a serine in mASIC3 and rASIC3. Lacking the full set of carboxylates in the proton-sensitive mASIC3 and rASIC3 confirms earlier results from ourselves and others highlighting that although the acidic pocket is involved in proton activation of ASICs, it alone is not responsible for proton activation of ASICs (17)(18)(19)(20)(21)(22). In nmrASIC3, E220 becomes D210, E239 becomes D229 and D346 is actually retained D352, thus nmrASIC3 actually has a full set of carboxylates in the acidic pocket, and although two glutamates are replaced by aspartates, which would have a different pKa, they are still protonatable residues.…”

Section: Discussionsupporting

confidence: 87%

“…However, ASIC2a lacks D350 and is still functional, whereas ASIC2b also only lacks the D350 carboxylate of the acidic pocket and is not activated by protons (13,17), results which suggest that regions outside of the acidic pocket must be important for proton activation of ASICs. Indeed, we and others have identified a range of residues on ASIC1a and ASIC2a that when mutated alter proton sensitivity (17)(18)(19)(20)(21) and more recently we have shown that the first 87 amino acids of the extracellular domain of rat ASIC2a are required for its proton sensitivity (22).…”

Section: Introductionmentioning

confidence: 81%

Naked mole-rat acid-sensing ion channel 3 forms nonfunctional homomers, but functional heteromers

Schuhmacher

Callejo

Srivats

et al. 2017

Preprint

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Acid-sensing ion channels (ASICs) form both homotrimeric and heterotrimeric ion channels that are activated by extracellular protons and are involved in a wide range of physiological and pathophysiological processes, including pain and anxiety. ASIC proteins can form both homotrimeric and heterotrimeric ion channels. The ASIC3 subunit has been shown to be of particular importance in the peripheral nervous system with pharmacological and genetic manipulations demonstrating a role in pain. Naked mole-rats, despite having functional ASICs, are insensitive to acid as a noxious stimulus and show diminished avoidance of acidic fumes, ammonia and carbon dioxide. Here we cloned naked mole-rat ASIC3 (nmrASIC3) and used a cell surface biotinylation assay to demonstrate that it traffics to the plasma membrane, but using whole-cell patch-clamp electrophysiology we observed that nmrASIC3 is insensitive to both protons and the non-proton ASIC3 agonist 2-Guanidine-4-methylquinazoline (GMQ). However, in line with previous reports of ASIC3 mRNA expression in dorsal root ganglia (DRG) neurons, we found that the ASIC3 antagonist APETx2 reversibly inhibits ASIC-like currents in naked mole-rat DRG neurons. We further show that like the proton-insensitive ASIC2b and ASIC4, nmrASIC3 forms functional, proton sensitive heteromers with other ASIC subunits. An amino acid alignment of ASIC3s between 9 relevant rodent species and human identified unique sequence differences that might underlie the proton insensitivity of nmrASIC3. However, introducing nmrASIC3 differences into rat ASIC3 (rASIC3) produced only minor differences in channel function, and replacing nmrASIC3 sequence with that of rASIC3 did not produce a proton-sensitive ion channel. Our observation that nmrASIC3 forms nonfunctional homomers may reflect a further adaptation of the naked mole-rat to living in an environment with high-carbon dioxide levels. IntroductionAcid-sensing ion channels (ASICs) are part of the epithelial sodium channel (ENaC)/degenerin (DEG) superfamily of ion channels and are implicated in a diverse range of physiological and pathophysiological processes, ranging from learning and memory to mechanosensation and pain (1). In mammals, there are 4 ASIC encoding genes, which generate 6 distinct ASIC subunits due to splice variants in the ACCN2 and ACCN1 genes producing a and b variants of the ASIC1 and ASIC2 subunits respectively: ASIC1a, ASIC1b, ASIC2a, ASIC2b, ASIC3 and ASIC4. The crystal structure of ASIC1 demonstrated that ASICs form trimeric ion channels (2) and although evidence exists for the formation of ASIC/ENaC heteromers (3, 4), it is largely thought that functional ASICs are the result of either homo-or heterotrimeric arrangement of ASIC subunits.Unlike transient receptor potential vanilloid 1 (TRPV1) that produces a sustained

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“…Proton sensitivity of ASIC homologs is observed in fishes, but missing in jawless vertebrate lampreys and chordates . ASIC2b, that is a splice variant of ASIC2, is not activated by reduced pH .…”

Section: Functional Comparisons Of Asics and Enacmentioning

confidence: 99%

ASIC and ENaC type sodium channels: conformational states and the structures of the ion selectivity filters

Hanukoglu

2016

The FEBS Journal

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The acid-sensing ion channels (ASICs) and epithelial sodium channels (ENaC) are members of a superfamily of channels that play critical roles in mechanosensation, chemosensation, nociception, and regulation of blood volume and pressure. These channels look and function like a tripartite funnel that directs the flow of Na + ions into the cytoplasm via the channel pore in the membrane. The subunits that form these channels share a common structure with two transmembrane segments (TM1 and TM2) and a large extracellular part. In most vertebrates, there are five paralogous genes that code for ASICs (ASIC1-ASIC5), and four for ENaC subunits alpha, beta, gamma, and delta (a, b, c, and d). While ASICs can form functional channels as a homo-or heterotrimer, ENaC functions as an obligate heterotrimer composed of a-b-c or b-c-d subunits. The structure of ASIC has been determined in several conformations, including desensitized and open states. This review presents a comparison of the structures of these states using easy-to-understand molecular models of the full complex, the central tunnel that includes an outer vestibule, the channel pore, and ion selectivity filter. The differences in the secondary, tertiary, and quaternary structures of the states are summarized to pinpoint the conformational changes responsible for channel opening. Results of site-directed mutagenesis studies of ENaC subunits are examined in light of ASIC1 models. Based on these comparisons, a molecular model for the selectivity filter of ENaC is built by in silico mutagenesis of an ASIC1 structure. These models suggest that Na + ions pass through the filter in a hydrated state.

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Structural Domains Underlying the Activation of Acid-Sensing Ion Channel 2a

Cited by 23 publications

References 35 publications

Conformational dynamics and role of the acidic pocket in ASIC pH-dependent gating

Conformational dynamics and role of the acidic pocket in ASIC pH-dependent gating

Naked mole-rat acid-sensing ion channel 3 forms nonfunctional homomers, but functional heteromers

ASIC and ENaC type sodium channels: conformational states and the structures of the ion selectivity filters

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