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- PDB-8hf2: Cryo-EM structure of WeiTsing -

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Basic information

Entry
Database: PDB / ID: 8hf2
TitleCryo-EM structure of WeiTsing
ComponentsPRA1 family protein
KeywordsMEMBRANE PROTEIN / Pentamer / Channel
Function / homologymembrane / Transmembrane protein
Function and homology information
Biological speciesArabidopsis thaliana (thale cress)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 4.14 Å
AuthorsQin, L. / Tang, L.H. / Chen, Y.H.
Funding support China, 2items
OrganizationGrant numberCountry
National Basic Research Program of China (973 Program)2021YFA1300702 China
Chinese Academy of SciencesXDA24020305 China
CitationJournal: Cell / Year: 2023
Title: WeiTsing, a pericycle-expressed ion channel, safeguards the stele to confer clubroot resistance.
Authors: Wei Wang / Li Qin / Wenjing Zhang / Linghui Tang / Chao Zhang / Xiaojing Dong / Pei Miao / Meng Shen / Huilong Du / Hangyuan Cheng / Ke Wang / Xiangyun Zhang / Min Su / Hongwei Lu / Chang Li ...Authors: Wei Wang / Li Qin / Wenjing Zhang / Linghui Tang / Chao Zhang / Xiaojing Dong / Pei Miao / Meng Shen / Huilong Du / Hangyuan Cheng / Ke Wang / Xiangyun Zhang / Min Su / Hongwei Lu / Chang Li / Qiang Gao / Xiaojuan Zhang / Yun Huang / Chengzhi Liang / Jian-Min Zhou / Yu-Hang Chen /
Abstract: Plant roots encounter numerous pathogenic microbes that often cause devastating diseases. One such pathogen, Plasmodiophora brassicae (Pb), causes clubroot disease and severe yield losses on ...Plant roots encounter numerous pathogenic microbes that often cause devastating diseases. One such pathogen, Plasmodiophora brassicae (Pb), causes clubroot disease and severe yield losses on cruciferous crops worldwide. Here, we report the isolation and characterization of WeiTsing (WTS), a broad-spectrum clubroot resistance gene from Arabidopsis. WTS is transcriptionally activated in the pericycle upon Pb infection to prevent pathogen colonization in the stele. Brassica napus carrying the WTS transgene displayed strong resistance to Pb. WTS encodes a small protein localized in the endoplasmic reticulum (ER), and its expression in plants induces immune responses. The cryoelectron microscopy (cryo-EM) structure of WTS revealed a previously unknown pentameric architecture with a central pore. Electrophysiology analyses demonstrated that WTS is a calcium-permeable cation-selective channel. Structure-guided mutagenesis indicated that channel activity is strictly required for triggering defenses. The findings uncover an ion channel analogous to resistosomes that triggers immune signaling in the pericycle.
History
DepositionNov 9, 2022Deposition site: PDBJ / Processing site: PDBJ
Revision 1.0Jun 21, 2023Provider: repository / Type: Initial release
Revision 1.1Jul 3, 2024Group: Data collection / Category: chem_comp_atom / chem_comp_bond / em_admin / Item: _em_admin.last_update

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

Downloads & links

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Assembly

Deposited unit
A: PRA1 family protein
B: PRA1 family protein
C: PRA1 family protein
D: PRA1 family protein
E: PRA1 family protein


Theoretical massNumber of molelcules
Total (without water)80,0205
Polymers80,0205
Non-polymers00
Water00
1


  • Idetical with deposited unit
  • defined by author
  • Evidence: electron microscopy
TypeNameSymmetry operationNumber
identity operation1_5551

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Components

#1: Protein
PRA1 family protein / WeiTsing


Mass: 16003.990 Da / Num. of mol.: 5
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Arabidopsis thaliana (thale cress) / Gene: AN1_LOCUS3247, C24_LOCUS3162 / Production host: Schizosaccharomyces pombe (fission yeast) / References: UniProt: A0A654EJS8

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Experimental details

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Experiment

ExperimentMethod: ELECTRON MICROSCOPY
EM experimentAggregation state: PARTICLE / 3D reconstruction method: single particle reconstruction

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Sample preparation

ComponentName: WeiTsing / Type: COMPLEX / Entity ID: all / Source: RECOMBINANT
Source (natural)Organism: Arabidopsis thaliana (thale cress)
Source (recombinant)Organism: Schizosaccharomyces pombe (fission yeast)
Buffer solutionpH: 8
SpecimenConc.: 2.5 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
Specimen supportGrid material: COPPER / Grid mesh size: 300 divisions/in. / Grid type: Quantifoil R1.2/1.3
VitrificationCryogen name: ETHANE / Humidity: 100 % / Chamber temperature: 277 K

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Electron microscopy imaging

Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company
MicroscopyModel: FEI TITAN KRIOS
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: FLOOD BEAM
Electron lensMode: BRIGHT FIELD / Nominal defocus max: 2000 nm / Nominal defocus min: 1200 nm / Cs: 2.7 mm
Image recordingElectron dose: 50 e/Å2 / Film or detector model: GATAN K3 (6k x 4k)

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Processing

SoftwareName: PHENIX / Version: dev_3689: / Classification: refinement
EM software
IDNameVersionCategory
2SerialEM4image acquisition
13cryoSPARC3.23D reconstruction
CTF correctionType: NONE
SymmetryPoint symmetry: C5 (5 fold cyclic)
3D reconstructionResolution: 4.14 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 117360 / Symmetry type: POINT
Refine LS restraints
Refine-IDTypeDev idealNumber
ELECTRON MICROSCOPYf_bond_d0.0124770
ELECTRON MICROSCOPYf_angle_d1.0076500
ELECTRON MICROSCOPYf_dihedral_angle_d6.421630
ELECTRON MICROSCOPYf_chiral_restr0.047815
ELECTRON MICROSCOPYf_plane_restr0.004770

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