Abstract
Temperature-sensitive TRP channels are important for responses to pain and inflammation, to both of which tissue acidosis is a major contributing factor. However, except for TRPV1, acid-sensing by other ThermoTRP channels remains mysterious. We show here that unique among TRPV1–3 channels, TRPV3 is directly activated by protons from cytoplasmic side. This effect is very weak and involves key cytoplasmic residues L508, D512, S518, or A520. However, mutations of these residues did not affect a strong proton induced potentiation of TRPV3 currents elicited by the TRPV1–3 common agonist, 2-aminoethoxydiphenyl borate (2-APB), no matter if the ligand was applied from extracellular or cytoplasmic side. The acid potentiation was common among TRPV1–3 and only seen with 2-APB-related ligands. Using 1H-nuclear magnetic resonance to examine the solution structures of 2-APB and its analogs, we observed striking structural differences of the boron-containing compounds at neutral/basic as compared to acidic pH, suggesting that a pH-dependent configuration switch of 2-APB-based drugs may underlie their functionality. Supporting this notion, protons also enhanced the inhibitory action of 2-APB on TRPM8. Collectively, our findings reveal novel insights into 2-APB action on TRP channels, which should facilitate the design of new drugs for these channels.
Highlights
Four members of the transient receptor potential vanilloid (TRPV) family, TRPV1–4, have been implicated to play important roles in mammalian chemo-somatosensing, such as changes in temperature, pressure, and osmolarity, as well as a large number of endogenous and exogenous chemical activators, including inflammatory mediators[1,2]
Because TRPV3 channels undergo sensitization upon repeated stimulation[21,32], we examined the effect of extracellular acidification after the response had stabilized following repeated applications of 100 μ M 2-aminoethoxydiphenyl borate (2-APB) at the neutral pH (7.4)
The above results suggest that extracellular acid reversibly enhances the response of TRPV3 to 2-APB
Summary
Four members of the transient receptor potential vanilloid (TRPV) family, TRPV1–4, have been implicated to play important roles in mammalian chemo-somatosensing, such as changes in temperature, pressure, and osmolarity, as well as a large number of endogenous and exogenous chemical activators, including inflammatory mediators[1,2]. Critical amino acid residues involved in proton sensing are localized at extracellular loops of the TRPV1 protein[11,12]. Site-directed mutagenesis revealed that an N-terminal histidine residue, H426, known to be critical for TRPV3 activation by its synthetic ligand, 2-aminoethoxydiphenyl borate (2-APB)[19], might be involved in sensing the intracellular pH drop. Given the importance of maintaining proper TRPV3 function to skin health and the inevitable exposure of TRPV3-expressing keratinocytes to acidic pH under physiological conditions of skin, which has a mean pH around 5.5 (ranging from 4 to 7)[31], it is critical to understand how TRPV3 channels respond to pH changes. The proton activation and potentiation employed different mechanisms, with residues in the S2–S3 loop required in the activation and acid-induced structural changes of the boron-containing compound likely involved in the ligand-dependent potentiation
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