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- EMDB-18400: GDNF/GFRa1 cell adhesion complex cryo-ET structure -

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Basic information

Entry
Database: EMDB / ID: EMD-18400
TitleGDNF/GFRa1 cell adhesion complex cryo-ET structure
Map dataSub-tomogram average map of zebrafish GDNF/GFRa1 cell adhesion complex assembled on reconstituted liposomes.
Sample
  • Complex: GDNF/GFRa1 cell adhesion complex
    • Protein or peptide: GDNF family receptor alpha
    • Protein or peptide: Glial cell line-derived neurotrophic factor
KeywordsSynaptic / Adhesion / Complex / LiCAM / CELL ADHESION
Function / homology
Function and homology information


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 / glial cell-derived neurotrophic factor receptor signaling pathway / regulation of morphogenesis of a branching structure / regulation of dopamine uptake involved in synaptic transmission / enteric nervous system development ...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 / glial cell-derived neurotrophic factor receptor signaling pathway / regulation of morphogenesis of a branching structure / regulation of dopamine uptake involved in synaptic transmission / enteric nervous system development / peristalsis / positive regulation of branching involved in ureteric bud morphogenesis / sympathetic nervous system development / peripheral nervous system development / metanephros development / mRNA stabilization / 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 / nervous system development / signaling receptor activity / 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)
Methodsubtomogram averaging / cryo EM / Resolution: 21.7 Å
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
DepositionSep 6, 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_18400.png

Downloads & links

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Map

FileDownload / File: emd_18400.map.gz / Format: CCP4 / Size: 2 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationSub-tomogram average map of zebrafish GDNF/GFRa1 cell adhesion complex assembled on reconstituted liposomes.
Voxel sizeX=Y=Z: 3.255 Å
Density
Contour LevelBy AUTHOR: 4.6
Minimum - Maximum-8.949246 - 19.611256000000001
Average (Standard dev.)-0.000000000129563 (±3.0387237)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions808080
Spacing808080
CellA=B=C: 260.40002 Å
α=β=γ: 90.0 °

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Supplemental data

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Mask #1

Fileemd_18400_msk_1.map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: Half Map A

Fileemd_18400_half_map_1.map
AnnotationHalf Map A
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: Half Map B

Fileemd_18400_half_map_2.map
AnnotationHalf Map B
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Sample components

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Entire : GDNF/GFRa1 cell adhesion complex

EntireName: GDNF/GFRa1 cell adhesion complex
Components
  • Complex: GDNF/GFRa1 cell adhesion complex
    • Protein or peptide: GDNF family receptor alpha
    • Protein or peptide: Glial cell line-derived neurotrophic factor

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Supramolecule #1: GDNF/GFRa1 cell adhesion complex

SupramoleculeName: GDNF/GFRa1 cell adhesion complex / 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 sequence and tag present at N- and C-terminus
Enantiomer: LEVO
Source (natural)Organism: Danio rerio (zebrafish)
Recombinant expressionOrganism: Homo sapiens (human)
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: DEXTRO
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
Processingsubtomogram averaging
Aggregation stateparticle

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Sample preparation

Concentration0.133 mg/mL
BufferpH: 7.5
Component:
ConcentrationFormulaName
180.0 mMNaClsodium 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.

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Electron microscopy

MicroscopeFEI TALOS ARCTICA
Image recordingFilm or detector model: FEI FALCON III (4k x 4k) / Detector mode: COUNTING / Number real images: 1 / Average exposure time: 0.72 sec. / Average electron dose: 1.99 e/Å2
Electron beamAcceleration voltage: 200 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / 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
Experimental equipment
Model: Talos Arctica / Image courtesy: FEI Company

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Image processing

Final reconstructionNumber classes used: 1 / Applied symmetry - Point group: D5 (2x5 fold dihedral) / Resolution.type: BY AUTHOR / Resolution: 21.7 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: RELION (ver. 4.0) / Number subtomograms used: 187
ExtractionNumber tomograms: 9 / Number images used: 502
Final angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: RELION (ver. 4.0)
FSC plot (resolution estimation)

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