@article{1e46895510e44552a390bda2983b82ef,
title = "Evidence that the TRPV1 S1-S4 membrane domain contributes to thermosensing",
abstract = "Sensing and responding to temperature is crucial in biology. The TRPV1 ion channel is a well-studied heat-sensing receptor that is also activated by vanilloid compounds, including capsaicin. Despite significant interest, the molecular underpinnings of thermosensing have remained elusive. The TRPV1 S1-S4 membrane domain couples chemical ligand binding to the pore domain during channel gating. Here we show that the S1-S4 domain also significantly contributes to thermosensing and couples to heat-activated gating. Evaluation of the isolated human TRPV1 S1-S4 domain by solution NMR, far-UV CD, and intrinsic fluorescence shows that this domain undergoes a non-denaturing temperature-dependent transition with a high thermosensitivity. Further NMR characterization of the temperature-dependent conformational changes suggests the contribution of the S1-S4 domain to thermosensing shares features with known coupling mechanisms between this domain with ligand and pH activation. Taken together, this study shows that the TRPV1 S1-S4 domain contributes to TRPV1 temperature-dependent activation.",
author = "Minjoo Kim and Sisco, {Nicholas J.} and Hilton, {Jacob K.} and Montano, {Camila M.} and Castro, {Manuel A.} and Cherry, {Brian R.} and Marcia Levitus and {Van Horn}, {Wade D.}",
note = "Funding Information: Given the role of the TRPV1 S1–S4 domain (V1-S1S4) in ligand activation and the structural similarity to VSDs in VGICs, we hypothesize that the S1–S4 domain may contribute to the thermosensitivity of TRPV1. We adopt a distinct strategy that is not reliant on mutagenesis but instead focuses on direct temperature-dependent characterization of the hV1-S1S4, an evolutionarily conserved structural domain (Fig. 1). Here, we show that an isolated hV1-S1S4 is in a biologically relevant state, is sufficient for vanilloid ligand binding, and retains the expected secondary structure and membrane topology. Temperature-dependent studies of the hV1-S1S4 using solution nuclear magnetic resonance (NMR) spectroscopy identify a two-state transition between folded conformational states. The magnitude of the NMR-detected temperature sensitivity (ΔH) is supported by far-UV circular dichroism (CD) and intrinsic tryptophan fluorescence spectroscopy. Quantitative comparison with whole-cell patch-clamp electrophysiology experiments in mammalian cells indicates that the hV1-S1S4 significantly contributes to TRPV1 thermosensitivity. Additionally, the temperature-dependent conformational changes were examined by NMR-based temperature-dependent distance measurements from paramagnetic relaxation enhancement (PRE), secondary structure measurements from chemical shift assignment and residual dipolar coupling, and solvent exposure from deuterium/hydrogen exchange and water–protein NOEs. To validate these outcomes, we introduced a mutation that disrupts coupling between the S1–S4 and pore domains and show that both ligand and temperature activation are abrogated in the full-length channel but retained in the isolated domain. Taken together, the data provide thermodynamic and mechanistic insight into the properties of thermosensitive TRP channels and suggest an overlap between TRPV1 ligand, proton (pH), and temperature activation.",
year = "2020",
month = dec,
day = "1",
doi = "10.1038/s41467-020-18026-2",
language = "English (US)",
volume = "11",
journal = "Nature Communications",
issn = "2041-1723",
publisher = "Nature Publishing Group",
number = "1",
}