Transient receptor potential vanilloid member 1 (TRPV1) is a Ca2+-permeable cation channel that serves as the primary heat and capsaicin sensor in humans. Using cryo-EM, we have determined the structures of apo and capsaicin-bound full-length rat TRPV1 reconstituted into lipid nanodiscs over a range of temperatures. This has allowed us to visualize the noxious heat-induced opening of TRPV1 in the presence of capsaicin. Notably, noxious heat-dependent TRPV1 opening comprises stepwise conformational transitions. Global conformational changes across multiple subdomains of TRPV1 are followed by the rearrangement of the outer pore, leading to gate opening. Solvent-accessible surface area analyses and functional studies suggest that a subset of residues form an interaction network that is directly involved in heat sensing. Our study provides a glimpse of the molecular principles underlying noxious physical and chemical stimuli sensing by TRPV1, which can be extended to other thermal sensing ion channels.
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The coordinates are deposited in the Protein Data Bank with PDB IDs 7LP9 (TRPV14C,APO), 7LPA (TRPV14C,CAP), 7LPB (TRPV125C,CAP), 7LPC (TRPV148C,APO), 7LPD (TRPV148C,CAP,INT) and 7LPE (TRPV148C,CAP,OPEN), respectively. The cryo-EM maps are deposited in the Electron Microscopy Data Bank with IDs EMD-23473 (TRPV14C,APO), EMD-23474 (TRPV14C,CAP), EMD-23475 (TRPV125C,CAP), EMD-23476 (TRPV148C,APO), EMD-23477 (TRPV148C,CAP,10sec), EMD-23478 (TRPV148C,CAP,INT) and EMD-23479 (TRPV148C,CAP,OPEN), respectively. Source data are provided with this paper.
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Cryo-EM data were collected at the Duke University Shared Materials Instrumentation Facility (SMIF) and at the Pacific Northwest Center for Cryo-EM (PNCC) at OHSU. We thank J. Myers at PNCC for assistance with data collection, N. Bhattacharya at SMIF for assistance with the microscope operation, L.B. Dillard at NIEHS for assistance with sample screening, and A. Bartesaghi at Duke for a pre-processing interface for data collection. We thank J. Fedor and Y. Yin for critical reading of the manuscript and discussions, W. Borschel for training F. Zhang on the patch-clamp recording, J. Grandl for discussions and W. Im for advice on the solvent accessibility analysis. This research was supported by NIH grant R35NS097241 (to S.-Y.L.) and by the National Institute of Health Intramural Research Program, US National Institutes of Environmental Health Sciences (ZIC ES103326 to M.J.B). A portion of this research was supported by NIH grant U24GM129547 and performed at the PNCC at OHSU and accessed through EMSL (grid.436923.9), a DOE Office of Science User Facility sponsored by the Office of Biological and Environmental Research. DUKE SMIF is affiliated with the North Carolina Research Triangle Nanotechnology Network, which is in part supported by the NSF (ECCS-2025064).
The authors declare no competing interests.
Peer review information Nature Structural & Molecular Biology thanks Chia-Hsueh Lee and the other, anonymous, reviewer(s) for their contribution to the peer review of this work. Peer reviewer reports are available. Florian Ullrich was the primary editor on this article and managed its editorial process and peer review in collaboration with the rest of the editorial team.
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Data processing procedures, a, Data processing flow chart for TRPV14C,APO, TRPV14C,CAP, TRPV125C,CAP, TRPV148C,APO. b, representative micrographs, see Table 1 for details. c, 2D classification images, d, 3D reconstructions, e, local resolution estimation, f, the Euler distribution plot, g, FSC curves for TRPV14C,APO, TRPV14C,CAP, TRPV125C,CAP, TRPV148C,APO, TRPV148C,CAP,OPEN and TRPV148C,CAP,INT, respectively. h, Data processing flow chart for TRPV148C,CAP,INT and TRPV148C,CAP,OPEN.
a-f, cryo-EM density for subdomains in TRPV14C,APO (a, thresholding 0.014), TRPV14C,CAP (b, thresholding 0.014), TRPV125C,CAP (c, thresholding 0.025), TRPV148C,APO (d, thresholding 0.019), TRPV148C,CAP,INT (e, thresholding 0.28), TRPV148C,CAP,OPEN (f, thresholding 0.3). Structural elements are shown as sticks and EM density as gray mesh.
a, Architecture of the TRPV1 protomer with subdomains indicated: ankyrin repeat domain (ARD), coupling domain (CD), transmembrane helices S1-S6, TRP helix, and C-terminal domain (CTD). b, Cryo-EM density (half-map without symmetry) for the selectivity filter of TRPV14C,APO corresponding to putative sodium ions at 0.04 thresholding. c, Cryo-EM density of the turret and turret junction (0.012 thresholding). d, Close-up view of the outer pore and turret junction (0.012 thresholding). e, Interaction networks spanning the outer pore region and the S1-S4 domain (0.03 thresholding). Key residues interacting with E600 and E648 are shown as sticks with surrounding cryo-EM density. f, g, Cryo-EM density of the CD, TRP, ARD (f), and CTD (g). The ARD is colored in gold, the CD and its individual elements (HTHCD, βCD,) in sky blue, the TRP domain in dark green, and the CTD in orange. The cryo-EM density (gray) is shown at 0.012 thresholding. h, Superposition of a single protomer from TRPV14C,APO (blue) and TRPV148C,APO (gold). i, Superposition of a single protomer from TRPV14C,APO (blue) and TRPV14C,CAP (cyan).
a, b, Cryo-EM 3D reconstructions of TRPV14C,APO (a, blue) and TRPV148C,APO (b, gold), respectively. Outlines indicate AR1-AR4. c, Close-up comparison of the cytoplasmic domains between TRPV14C,APO (blue) and TRPV148C,APO (gold).
Extended Data Fig. 5 Comparison of TRPV14C,APO, TRPV14C,CAP, TRPV125C,CAP and the published structure of TRPV1 in the presence of capsaicin.
a, Close-up view of the S1-S4 domain of TRPV14C,APO (blue) and TRPV14C,CAP (cyan). Capsaicin (red) and phosphatidyl inositol (blue) molecules are shown as sticks. b, Close-up view of capsaicin in the vanilloid pocket of TRPV14C,CAP. The cryo-EM density is shown at 0.025 thresholding. c, Side view comparison of TRPV14C,CAP (cyan) and TRPV125C,CAP (green). d, Side view comparison of TRPV14C,CAP (green) and the published TRPV1 structure in the presence of capsaicin (PDB ID: 3J5R, brown).
Extended Data Fig. 6 Comparison between the overall structures of TRPV14C,APO, TRPV148C,CAP,OPEN and DkTx/RTx-bound TRPV1.
a, Comparison of TRPV14C,APO (silver), TRPV148C,CAP,OPEN (red), and DkTx/RTx-TRPV1 (blue) viewed from the intracellular side. ARD/CD movement occurs at an individual protomer level. b, Comparison of the S6b and TRP domain of TRPV14C,APO, TRPV148C,CAP,OPEN, and DkTx/RTx-TRPV1. c, Close-up view of TRPV14C,APO, TRPV148C,CAP,OPEN, and DkTx/RTx-TRPV1 in the cytoplasmic domains. d, Alternate angle and close-up view of TRPV14C,APO, TRPV148C,CAP,OPEN, and DkTx/RTx-TRPV1 in the cytoplasmic domains.
a, The overlapping locations of phospholipid (TRPV148C,CAP,OPEN, red) and DkTx (DkTx/RTx-TRPV1, blue), shown as sticks and spheres, between the pore loop and pore helix. Several side chains are shown as sticks to illustrate the differences in the outer pore of the two structures. b, Structural differences between TRPV148C,CAP,OPEN and DkTx/RTx-TRPV1 at S6, the S4-S5 linker, and the TRP helix.
Extended Data Fig. 8 Solvent accessible surface area-based heat capacity change plots for the first and the second transitions.
a, b, ΔCPpred plots for the first (a) and second (b) transitions. For each transition, residues exhibiting positive ΔCPpred are plotted in the upper graph using log10(ΔCPpred), and residues exhibiting negative ΔCPpred are plotted in the lower graph using -log10(-ΔCPpred). The dotted line denotes the 15 J mol-1 K-1 threshold. ΔCPpred was calculated as described in the Methods. Residues for which the side chains were not resolved were not included in the calculation.
a, Close-up view of the vanilloid binding site in TRPV125C,CAP (green), TRPV148C,CAP,INT (orange), and TRPV148C,CAP,OPEN (red). Several key residues in capsaicin are shown as sticks. Dotted lines denote either H-bond or salt bridge interactions. The 310 helical region of S4 is indicated as 310. b, Close-up view of S5 and S6 in TRPV148C,CAP,INT (orange), and TRPV148C,CAP,OPEN (red). The π helical turn in S6 is denoted by π. c, Comparison of TRPV148C,CAP,OPEN (red) and DkTx/RTx-bound TRPV1 (PDB ID: 5IRX, blue). DkTx is shown as sticks and gray spheres; capsaicin is depicted as sticks only.
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Kwon, D.H., Zhang, F., Suo, Y. et al. Heat-dependent opening of TRPV1 in the presence of capsaicin. Nat Struct Mol Biol 28, 554–563 (2021). https://doi.org/10.1038/s41594-021-00616-3
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