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- EMDB-40961: Closed-state cryo-EM structure of full-length human TRPV4 in the ... -

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Entry
Database: EMDB / ID: EMD-40961
TitleClosed-state cryo-EM structure of full-length human TRPV4 in the presence of 4a-PDD
Map data
Sample
  • Complex: full-length human TRPV4 in the presence of 4a-PDD
    • Protein or peptide: Transient receptor potential cation channel subfamily V member 4,Enhanced green fluorescent protein
  • Ligand: [(2~{R})-2-[(~{Z})-hexadec-9-enoyl]oxy-3-[oxidanyl-[2-(trimethyl-$l^{4}-azanyl)ethoxy]phosphoryl]oxy-propyl] (~{Z})-docos-13-enoate
  • Ligand: (2R)-2-{[(4-O-hexopyranosyl-beta-D-glucopyranosyl)oxy]methyl}-4-{[(25R)-5beta,14beta,17beta-spirostan-3beta-yl]oxy}butyl 4-O-alpha-D-glucopyranosyl-beta-D-glucopyranoside
  • Ligand: SODIUM ION
  • Ligand: water
Keywordstransient receptor potential V family member 4 / TRP / channel / TRPV4 / TRP channels / membrane protein / agonist / 4a-PDD / 4alpha-Phorbol 12 / 13-didecanoate / closed state
Function / homology
Function and homology information


stretch-activated, monoatomic cation-selective, calcium channel activity / blood vessel endothelial cell delamination / regulation of response to osmotic stress / osmosensor activity / vasopressin secretion / positive regulation of striated muscle contraction / calcium ion import into cytosol / negative regulation of brown fat cell differentiation / positive regulation of microtubule depolymerization / positive regulation of macrophage inflammatory protein 1 alpha production ...stretch-activated, monoatomic cation-selective, calcium channel activity / blood vessel endothelial cell delamination / regulation of response to osmotic stress / osmosensor activity / vasopressin secretion / positive regulation of striated muscle contraction / calcium ion import into cytosol / negative regulation of brown fat cell differentiation / positive regulation of microtubule depolymerization / positive regulation of macrophage inflammatory protein 1 alpha production / hyperosmotic salinity response / positive regulation of chemokine (C-X-C motif) ligand 1 production / positive regulation of chemokine (C-C motif) ligand 5 production / cartilage development involved in endochondral bone morphogenesis / cellular hypotonic salinity response / cellular hypotonic response / cortical microtubule organization / multicellular organismal-level water homeostasis / positive regulation of vascular permeability / osmosensory signaling pathway / cell-cell junction assembly / positive regulation of monocyte chemotactic protein-1 production / calcium ion import / cell volume homeostasis / cellular response to osmotic stress / regulation of aerobic respiration / TRP channels / cortical actin cytoskeleton / diet induced thermogenesis / positive regulation of macrophage chemotaxis / microtubule polymerization / calcium ion import across plasma membrane / response to mechanical stimulus / alpha-tubulin binding / beta-tubulin binding / monoatomic cation channel activity / cytoplasmic microtubule / SH2 domain binding / protein kinase C binding / actin filament organization / bioluminescence / generation of precursor metabolites and energy / positive regulation of JNK cascade / filopodium / adherens junction / response to insulin / calcium ion transmembrane transport / positive regulation of interleukin-6 production / calcium channel activity / ruffle membrane / intracellular calcium ion homeostasis / positive regulation of inflammatory response / actin filament binding / calcium ion transport / glucose homeostasis / lamellipodium / negative regulation of neuron projection development / cellular response to heat / actin binding / positive regulation of cytosolic calcium ion concentration / growth cone / actin cytoskeleton organization / High laminar flow shear stress activates signaling by PIEZO1 and PECAM1:CDH5:KDR in endothelial cells / microtubule binding / calmodulin binding / response to hypoxia / positive regulation of ERK1 and ERK2 cascade / cilium / apical plasma membrane / focal adhesion / lipid binding / protein kinase binding / cell surface / endoplasmic reticulum / negative regulation of transcription by RNA polymerase II / ATP binding / metal ion binding / identical protein binding / membrane / plasma membrane
Similarity search - Function
Transient receptor potential cation channel subfamily V member 4 / Transient receptor potential cation channel subfamily V member 1-4 / Transient receptor potential cation channel subfamily V / Green fluorescent protein, GFP / Ankyrin repeat / Green fluorescent protein-related / Green fluorescent protein / Green fluorescent protein / Ankyrin repeat profile. / Ankyrin repeat region circular profile. ...Transient receptor potential cation channel subfamily V member 4 / Transient receptor potential cation channel subfamily V member 1-4 / Transient receptor potential cation channel subfamily V / Green fluorescent protein, GFP / Ankyrin repeat / Green fluorescent protein-related / Green fluorescent protein / Green fluorescent protein / Ankyrin repeat profile. / Ankyrin repeat region circular profile. / ankyrin repeats / Ankyrin repeat / Ankyrin repeat-containing domain superfamily / Ion transport domain / Ion transport protein
Similarity search - Domain/homology
Enhanced green fluorescent protein / Transient receptor potential cation channel subfamily V member 4
Similarity search - Component
Biological speciesHomo sapiens (human) / Human betaherpesvirus 5
Methodsingle particle reconstruction / cryo EM / Resolution: 2.77 Å
AuthorsTalyzina IA / Nadezhdin KD / Neuberger A / Sobolevsky AI
Funding support United States, Germany, 6 items
OrganizationGrant numberCountry
National Institutes of Health/National Institute of Arthritis and Musculoskeletal and Skin Diseases (NIH/NIAMS)R01 AR078814 United States
National Institutes of Health/National Cancer Institute (NIH/NCI)R01 CA206573 United States
National Institutes of Health/National Institute of Neurological Disorders and Stroke (NIH/NINDS)R01 NS083660 United States
National Institutes of Health/National Institute of Neurological Disorders and Stroke (NIH/NINDS)R01 NS107253 United States
German Research Foundation (DFG)464295817 Germany
National Institutes of Health/National Heart, Lung, and Blood Institute (NIH/NHLBI)R01 HL096647 United States
CitationJournal: Nat Commun / Year: 2023
Title: Structure of human TRPV4 in complex with GTPase RhoA.
Authors: Kirill D Nadezhdin / Irina A Talyzina / Aravind Parthasarathy / Arthur Neuberger / David X Zhang / Alexander I Sobolevsky /
Abstract: Transient receptor potential (TRP) channel TRPV4 is a polymodal cellular sensor that responds to moderate heat, cell swelling, shear stress, and small-molecule ligands. It is involved in ...Transient receptor potential (TRP) channel TRPV4 is a polymodal cellular sensor that responds to moderate heat, cell swelling, shear stress, and small-molecule ligands. It is involved in thermogenesis, regulation of vascular tone, bone homeostasis, renal and pulmonary functions. TRPV4 is implicated in neuromuscular and skeletal disorders, pulmonary edema, and cancers, and represents an important drug target. The cytoskeletal remodeling GTPase RhoA has been shown to suppress TRPV4 activity. Here, we present a structure of the human TRPV4-RhoA complex that shows RhoA interaction with the membrane-facing surface of the TRPV4 ankyrin repeat domains. The contact interface reveals residues that are mutated in neuropathies, providing an insight into the disease pathogenesis. We also identify the binding sites of the TRPV4 agonist 4α-PDD and the inhibitor HC-067047 at the base of the S1-S4 bundle, and show that agonist binding leads to pore opening, while channel inhibition involves a π-to-α transition in the pore-forming helix S6. Our structures elucidate the interaction interface between hTRPV4 and RhoA, as well as residues at this interface that are involved in TRPV4 disease-causing mutations. They shed light on TRPV4 activation and inhibition and provide a template for the design of future therapeutics for treatment of TRPV4-related diseases.
History
DepositionJun 2, 2023-
Header (metadata) releaseJul 5, 2023-
Map releaseJul 5, 2023-
UpdateOct 16, 2024-
Current statusOct 16, 2024Processing site: RCSB / Status: Released

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Structure visualization

Supplemental images

emd_40961.png

Downloads & links

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Map

FileDownload / File: emd_40961.map.gz / Format: CCP4 / Size: 125 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
0.79 Å/pix.
x 320 pix.
= 252.416 Å		0.79 Å/pix.
x 320 pix.
= 252.416 Å		0.79 Å/pix.
x 320 pix.
= 252.416 Å

Surface

Projections

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Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 0.7888 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.065
Minimum - Maximum-0.32909533 - 0.49701604
Average (Standard dev.)0.0008339692 (±0.014981232)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions320320320
Spacing320320320
CellA=B=C: 252.416 Å
α=β=γ: 90.0 °

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Supplemental data

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Mask #1

Fileemd_40961_msk_1.map
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Histogram

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Half map: #2

Fileemd_40961_half_map_1.map
Projections & Slices
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Histogram

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Half map: #1

Fileemd_40961_half_map_2.map
Projections & Slices
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Sample components

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Entire : full-length human TRPV4 in the presence of 4a-PDD

EntireName: full-length human TRPV4 in the presence of 4a-PDD
Components
  • Complex: full-length human TRPV4 in the presence of 4a-PDD
    • Protein or peptide: Transient receptor potential cation channel subfamily V member 4,Enhanced green fluorescent protein
  • Ligand: [(2~{R})-2-[(~{Z})-hexadec-9-enoyl]oxy-3-[oxidanyl-[2-(trimethyl-$l^{4}-azanyl)ethoxy]phosphoryl]oxy-propyl] (~{Z})-docos-13-enoate
  • Ligand: (2R)-2-{[(4-O-hexopyranosyl-beta-D-glucopyranosyl)oxy]methyl}-4-{[(25R)-5beta,14beta,17beta-spirostan-3beta-yl]oxy}butyl 4-O-alpha-D-glucopyranosyl-beta-D-glucopyranoside
  • Ligand: SODIUM ION
  • Ligand: water

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Supramolecule #1: full-length human TRPV4 in the presence of 4a-PDD

SupramoleculeName: full-length human TRPV4 in the presence of 4a-PDD / type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1
Source (natural)Organism: Homo sapiens (human)
Molecular weightTheoretical: 510 KDa

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Macromolecule #1: Transient receptor potential cation channel subfamily V member 4,...

MacromoleculeName: Transient receptor potential cation channel subfamily V member 4,Enhanced green fluorescent protein
type: protein_or_peptide / ID: 1 / Number of copies: 4 / Enantiomer: LEVO
Source (natural)Organism: Human betaherpesvirus 5
Molecular weightTheoretical: 127.717938 KDa
Recombinant expressionOrganism: Homo sapiens (human)
SequenceString: MADSSEGPRA GPGEVAELPG DESGTPGGEA FPLSSLANLF EGEDGSLSPS PADASRPAGP GDGRPNLRMK FQGAFRKGVP NPIDLLEST LYESSVVPGP KKAPMDSLFD YGTYRHHSSD NKRWRKKIIE KQPQSPKAPA PQPPPILKVF NRPILFDIVS R GSTADLDG ...String:
MADSSEGPRA GPGEVAELPG DESGTPGGEA FPLSSLANLF EGEDGSLSPS PADASRPAGP GDGRPNLRMK FQGAFRKGVP NPIDLLEST LYESSVVPGP KKAPMDSLFD YGTYRHHSSD NKRWRKKIIE KQPQSPKAPA PQPPPILKVF NRPILFDIVS R GSTADLDG LLPFLLTHKK RLTDEEFREP STGKTCLPKA LLNLSNGRND TIPVLLDIAE RTGNMREFIN SPFRDIYYRG QT ALHIAIE RRCKHYVELL VAQGADVHAQ ARGRFFQPKD EGGYFYFGEL PLSLAACTNQ PHIVNYLTEN PHKKADMRRQ DSR GNTVLH ALVAIADNTR ENTKFVTKMY DLLLLKCARL FPDSNLEAVL NNDGLSPLMM AAKTGKIGIF QHIIRREVTD EDTR HLSRK FKDWAYGPVY SSLYDLSSLD TCGEEASVLE ILVYNSKIEN RHEMLAVEPI NELLRDKWRK FGAVSFYINV VSYLC AMVI FTLTAYYQPL EGTPPYPYRT TVDYLRLAGE VITLFTGVLF FFTNIKDLFM KKCPGVNSLF IDGSFQLLYF IYSVLV IVS AALYLAGIEA YLAVMVFALV LGWMNALYFT RGLKLTGTYS IMIQKILFKD LFRFLLVYLL FMIGYASALV SLLNPCA NM KVCNEDQTNC TVPTYPSCRD SETFSTFLLD LFKLTIGMGD LEMLSSTKYP VVFIILLVTY IILTFVLLLN MLIALMGE T VGQVSKESKH IWKLQWATTI LDIERSFPVF LRKAFRSGEM VTVGKSSDGT PDRRWCFRVD EVNWSHWNQN LGIINEDPG KNETYQYYGF SHTVGRLRRD RWSSVVPRVV ELNKNSNPDE VVVPLDSMGN PRCDGHQQGY PRKWRTDDAP LLVPRGSAAA AVSKGEELF TGVVPILVEL DGDVNGHKFS VSGEGEGDAT YGKLTLKFIC TTGKLPVPWP TLVTTLTYGV QCFSRYPDHM K QHDFFKSA MPEGYVQERT IFFKDDGNYK TRAEVKFEGD TLVNRIELKG IDFKEDGNIL GHKLEYNYNS HNVYIMADKQ KN GIKVNFK IRHNIEDGSV QLADHYQQNT PIGDGPVLLP DNHYLSTQSK LSKDPNEKRD HMVLLEFVTA AGITLGMDEL YKS GLRSWS HPQFEK

UniProtKB: Transient receptor potential cation channel subfamily V member 4, Enhanced green fluorescent protein

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Macromolecule #2: [(2~{R})-2-[(~{Z})-hexadec-9-enoyl]oxy-3-[oxidanyl-[2-(trimethyl-...

MacromoleculeName: [(2~{R})-2-[(~{Z})-hexadec-9-enoyl]oxy-3-[oxidanyl-[2-(trimethyl-$l^{4}-azanyl)ethoxy]phosphoryl]oxy-propyl] (~{Z})-docos-13-enoate
type: ligand / ID: 2 / Number of copies: 32 / Formula: 9ZR
Molecular weightTheoretical: 815.175 Da
Chemical component information

ChemComp-9ZR:
[(2~{R})-2-[(~{Z})-hexadec-9-enoyl]oxy-3-[oxidanyl-[2-(trimethyl-$l^{4}-azanyl)ethoxy]phosphoryl]oxy-propyl] (~{Z})-docos-13-enoate

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Macromolecule #3: (2R)-2-{[(4-O-hexopyranosyl-beta-D-glucopyranosyl)oxy]methyl}-4-{...

MacromoleculeName: (2R)-2-{[(4-O-hexopyranosyl-beta-D-glucopyranosyl)oxy]methyl}-4-{[(25R)-5beta,14beta,17beta-spirostan-3beta-yl]oxy}butyl 4-O-alpha-D-glucopyranosyl-beta-D-glucopyranoside
type: ligand / ID: 3 / Number of copies: 8 / Formula: YJ0
Molecular weightTheoretical: 1.165315 KDa
Chemical component information

ChemComp-YJ0:
(2R)-2-{[(4-O-hexopyranosyl-beta-D-glucopyranosyl)oxy]methyl}-4-{[(25R)-5beta,14beta,17beta-spirostan-3beta-yl]oxy}butyl 4-O-alpha-D-glucopyranosyl-beta-D-glucopyranoside

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Macromolecule #4: SODIUM ION

MacromoleculeName: SODIUM ION / type: ligand / ID: 4 / Number of copies: 3
Molecular weightTheoretical: 22.99 Da

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Macromolecule #5: water

MacromoleculeName: water / type: ligand / ID: 5 / Number of copies: 36 / Formula: HOH
Molecular weightTheoretical: 18.015 Da
Chemical component information

ChemComp-HOH:
WATER

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Experimental details

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Structure determination

Methodcryo EM
Processingsingle particle reconstruction
Aggregation stateparticle

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Sample preparation

Concentration3.2 mg/mL
BufferpH: 8
Component:
ConcentrationFormulaName
150.0 mMNaClsodium chloride
20.0 mMtris(hydroxymethyl)aminomethane
1.0 mMbeta-Mercaptoethanol
0.01 %glyco-diosgenin
10.0 mMGTPgammaS
GridModel: UltrAuFoil R1.2/1.3 / Material: GOLD / Mesh: 300 / Support film - Material: GOLD / Support film - topology: HOLEY
VitrificationCryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 295 K / Instrument: FEI VITROBOT MARK IV
Detailshuman TRPV4

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Electron microscopy

MicroscopeTFS KRIOS
Image recordingFilm or detector model: GATAN K3 (6k x 4k) / Digitization - Dimensions - Width: 11520 pixel / Digitization - Dimensions - Height: 8184 pixel / Number grids imaged: 1 / Number real images: 15624 / Average exposure time: 2.4 sec. / Average electron dose: 60.0 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Cs: 2.7 mm / Nominal defocus max: 1.5 µm / Nominal defocus min: 0.75 µm
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Particle selectionNumber selected: 3935964
Startup modelType of model: OTHER
Final reconstructionResolution.type: BY AUTHOR / Resolution: 2.77 Å / Resolution method: FSC 0.143 CUT-OFF / Number images used: 75146
Initial angle assignmentType: OTHER
Final angle assignmentType: OTHER
FSC plot (resolution estimation)

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Atomic model buiding 1

RefinementSpace: REAL
Output model

PDB-8t1e:
Closed-state cryo-EM structure of full-length human TRPV4 in the presence of 4a-PDD

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