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- EMDB-18651: GDNF/GFRa1 cell adhesion complex bridging between adhering liposomes. -

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Basic information

Entry
Database: EMDB / ID: EMD-18651
TitleGDNF/GFRa1 cell adhesion complex bridging between adhering liposomes.
Map dataReconstructed tomogram of zGDNF-zGFRa1 complexes bridging between adhering liposomes. Tomogram reconstructions were generated with a SIRT-like filter with 5 iterations and binned by 4.
Sample
  • Complex: GDNF/GFRa1 cell adhesion complex assembled onto liposomes.
    • Protein or peptide: GDNF family receptor alphaGFRα
    • Protein or peptide: Glial cell line-derived neurotrophic factor
KeywordsSynaptic / Adhesion / Complex / LiCAM / CELL ADHESION
Function / homology
Function and homology information


RAF/MAP kinase cascade / : / diencephalon development / positive regulation of ureteric bud formation / postganglionic parasympathetic fiber development / positive regulation of monooxygenase activity / regulation of dopaminergic neuron differentiation / glial cell-derived neurotrophic factor receptor binding / regulation of morphogenesis of a branching structure / regulation of dopamine uptake involved in synaptic transmission ...RAF/MAP kinase cascade / : / diencephalon development / positive regulation of ureteric bud formation / postganglionic parasympathetic fiber development / positive regulation of monooxygenase activity / regulation of dopaminergic neuron differentiation / glial cell-derived neurotrophic factor receptor binding / regulation of morphogenesis of a branching structure / regulation of dopamine uptake involved in synaptic transmission / enteric nervous system development / positive regulation of branching involved in ureteric bud morphogenesis / peristalsis / sympathetic nervous system development / peripheral nervous system development / mRNA stabilization / metanephros development / neural crest cell migration / branching involved in ureteric bud morphogenesis / MAP kinase kinase kinase activity / growth factor activity / receptor tyrosine kinase binding / neuron projection development / signaling receptor activity / nervous system development / protein-containing complex assembly / negative regulation of neuron apoptotic process / receptor complex / external side of plasma membrane / protein homodimerization activity / positive regulation of transcription by RNA polymerase II / extracellular space / extracellular region
Similarity search - Function
: / Glial cell line-derived neurotrophic factor receptor, alpha 1 / Glial cell line-derived neurotrophic factor / Glial cell line-derived neurotrophic factor receptor, alpha 1/2 / Glial cell line-derived neurotrophic factor receptor / GDNF receptor alpha / Glial cell line-derived neurotrophic factor family / GDNF/GAS1 / GDNF/GAS1 domain / GDNF/GAS1 domain ...: / Glial cell line-derived neurotrophic factor receptor, alpha 1 / Glial cell line-derived neurotrophic factor / Glial cell line-derived neurotrophic factor receptor, alpha 1/2 / Glial cell line-derived neurotrophic factor receptor / GDNF receptor alpha / Glial cell line-derived neurotrophic factor family / GDNF/GAS1 / GDNF/GAS1 domain / GDNF/GAS1 domain / Transforming growth factor-beta, C-terminal / Transforming growth factor beta like domain / TGF-beta family profile. / Cystine-knot cytokine
Similarity search - Domain/homology
GDNF family receptor alpha / Glial cell line-derived neurotrophic factor
Similarity search - Component
Biological speciesDanio rerio (zebrafish)
Methodelectron tomography / cryo EM
AuthorsHoughton FM / Briggs DC / McDonald NQ
Funding support United Kingdom, 1 items
OrganizationGrant numberCountry
The Francis Crick InstituteCC2068 United Kingdom
CitationJournal: Nat Commun / Year: 2023
Title: Architecture and regulation of a GDNF-GFRα1 synaptic adhesion assembly.
Authors: F M Houghton / S E Adams / A S Ríos / L Masino / A G Purkiss / D C Briggs / F Ledda / N Q McDonald /
Abstract: Glial-cell line derived neurotrophic factor (GDNF) bound to its co-receptor GFRα1 stimulates the RET receptor tyrosine kinase, promoting neuronal survival and neuroprotection. The GDNF-GFRα1 ...Glial-cell line derived neurotrophic factor (GDNF) bound to its co-receptor GFRα1 stimulates the RET receptor tyrosine kinase, promoting neuronal survival and neuroprotection. The GDNF-GFRα1 complex also supports synaptic cell adhesion independently of RET. Here, we describe the structure of a decameric GDNF-GFRα1 assembly determined by crystallography and electron microscopy, revealing two GFRα1 pentamers bridged by five GDNF dimers. We reconsitituted the assembly between adhering liposomes and used cryo-electron tomography to visualize how the complex fulfils its membrane adhesion function. The GFRα1:GFRα1 pentameric interface was further validated both in vitro by native PAGE and in cellulo by cell-clustering and dendritic spine assays. Finally, we provide biochemical and cell-based evidence that RET and heparan sulfate cooperate to prevent assembly of the adhesion complex by competing for the adhesion interface. Our results provide a mechanistic framework to understand GDNF-driven cell adhesion, its relationship to trophic signalling, and the central role played by GFRα1.
History
DepositionOct 12, 2023-
Header (metadata) releaseNov 29, 2023-
Map releaseNov 29, 2023-
UpdateMar 27, 2024-
Current statusMar 27, 2024Processing site: PDBe / Status: Released

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Structure visualization

Supplemental images

emd_18651.png

Downloads & links

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Map

FileDownload / File: emd_18651.map.gz / Format: CCP4 / Size: 21.8 MB / Type: IMAGE STORED AS SIGNED BYTE
AnnotationReconstructed tomogram of zGDNF-zGFRa1 complexes bridging between adhering liposomes. Tomogram reconstructions were generated with a SIRT-like filter with 5 iterations and binned by 4.
Voxel sizeX=Y=Z: 13.02 Å
Density
Minimum - Maximum-128.0 - 127.0
Average (Standard dev.)86.419120000000007 (±4.7673326)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin313387-52
Dimensions50859276
Spacing59250876
CellA: 7707.8403 Å / B: 6614.16 Å / C: 989.52 Å
α=β=γ: 90.0 °

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Supplemental data

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Sample components

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Entire : GDNF/GFRa1 cell adhesion complex assembled onto liposomes.

EntireName: GDNF/GFRa1 cell adhesion complex assembled onto liposomes.
Components
  • Complex: GDNF/GFRa1 cell adhesion complex assembled onto liposomes.
    • Protein or peptide: GDNF family receptor alphaGFRα
    • Protein or peptide: Glial cell line-derived neurotrophic factor

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Supramolecule #1: GDNF/GFRa1 cell adhesion complex assembled onto liposomes.

SupramoleculeName: GDNF/GFRa1 cell adhesion complex assembled onto liposomes.
type: complex / ID: 1 / Parent: 0 / Macromolecule list: all
Source (natural)Organism: Danio rerio (zebrafish)
Molecular weightTheoretical: 700 KDa

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Macromolecule #1: GDNF family receptor alpha

MacromoleculeName: GDNF family receptor alpha / type: protein_or_peptide / ID: 1
Details: Vector-derived sequences and tag present at N- and C-terminus.
Enantiomer: LEVO
Source (natural)Organism: Danio rerio (zebrafish)
SequenceString: APLAEPALAK E NNCLNAAK AC NLNDTCK KYR SAYISP CTSR VSTAE VCNKR KCHK ALRQFF DKV PPKHSYG ML YCSCPLGD Q SACSERRRQ TIVPACSYED KERPNCLTL Q VSCKTNYI CR SRLADFF TNC QPEPLS LSGC LKENY ADCLL SYSG ...String:
APLAEPALAK E NNCLNAAK AC NLNDTCK KYR SAYISP CTSR VSTAE VCNKR KCHK ALRQFF DKV PPKHSYG ML YCSCPLGD Q SACSERRRQ TIVPACSYED KERPNCLTL Q VSCKTNYI CR SRLADFF TNC QPEPLS LSGC LKENY ADCLL SYSG LIGTVM TPN YLRSPKI SV SPFCDCSS S GNSKEECDR FTEFFTDNAC LRNAIQAFG N GTDVSVWH PM PPVQTTT S AAAHHHHH H

UniProtKB: GDNF family receptor alpha

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Macromolecule #2: Glial cell line-derived neurotrophic factor

MacromoleculeName: Glial cell line-derived neurotrophic factor / type: protein_or_peptide / ID: 2 / Details: Vector-derived sequence at N-terminus / Enantiomer: LEVO
Source (natural)Organism: Danio rerio (zebrafish)
Recombinant expressionOrganism: Homo sapiens (human)
SequenceString:
APLA RVKGQ GR GCLLKEI HLN VTDLDL GYRT KEELI FRYCS GPCH DAETNY DKI LNNLTHN KK LDKDTPSR T CCRPIAFDD DISFLDDSLE YHTLKKHSA K KCACV

UniProtKB: Glial cell line-derived neurotrophic factor

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Experimental details

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Structure determination

Methodcryo EM
Processingelectron tomography
Aggregation stateparticle

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Sample preparation

Concentration0.133 mg/mL
BufferpH: 7.5
Component:
ConcentrationFormulaName
180.0 mMNaClSodium chloridesodium chloride
20.0 mMHEPES4-(2-hydroxyethyl)-1-piperazineethanesulfonic acid
GridModel: Quantifoil R2/2 / Material: COPPER / Mesh: 300 / Pretreatment - Type: GLOW DISCHARGE / Pretreatment - Time: 45 sec. / Pretreatment - Atmosphere: AIR
VitrificationCryogen name: ETHANE / Chamber humidity: 85 % / Chamber temperature: 295 K / Instrument: FEI VITROBOT MARK IV / Details: 3 s blot time.
DetailsUncrosslinked GDNF/GFRa1 complex assembled onto liposome surfaces.
SectioningOther: NO SECTIONING
Fiducial markerManufacturer: BBI Solutions / Diameter: 10 nm

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Electron microscopy

MicroscopeFEI TALOS ARCTICA
Electron beamAcceleration voltage: 200 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELDBright-field microscopy / Nominal defocus max: 7.0 µm / Nominal defocus min: 4.0 µm / Nominal magnification: 45000
Sample stageSpecimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN
Image recordingFilm or detector model: FEI FALCON III (4k x 4k) / Detector mode: COUNTING / Number grids imaged: 1 / Number real images: 38 / Average exposure time: 0.72 sec. / Average electron dose: 1.99 e/Å2
Experimental equipment
Model: Talos Arctica / Image courtesy: FEI Company

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Image processing

Final reconstructionAlgorithm: BACK PROJECTION / Software - Name: IMOD (ver. 4.12)
Details: The final alignment was binned by 4 and the final tomogram was generated using a back-projection algorithm with a SIRT-like filter equivalent to 5 iterations of the SIRT algorithm.
Number images used: 38

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