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- EMDB-0966: cryo-EM structure of C9ORF72-SMCR8-WDR41 -

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Basic information

Entry
Database: EMDB / ID: EMD-0966
Titlecryo-EM structure of C9ORF72-SMCR8-WDR41
Map dataoverall map
Sample
  • Complex: Dimer of C9-ORF72-SMCR8-WDR41
    • Protein or peptide: WD repeat-containing protein 41
    • Protein or peptide: Guanine nucleotide exchange protein SMCR8
    • Protein or peptide: Guanine nucleotide exchange C9orf72
KeywordsALS / FTD / GTPase / C9ORF72 / SMCR8 / WDR41 / GAP / GEF / PROTEIN BINDING
Function / homology
Function and homology information


Atg1/ULK1 kinase complex / late endosome to lysosome transport / regulation of TORC1 signaling / regulation of autophagosome assembly / negative regulation of autophagosome assembly / regulation of actin filament organization / guanyl-nucleotide exchange factor complex / negative regulation of immune response / axon extension / negative regulation of exocytosis ...Atg1/ULK1 kinase complex / late endosome to lysosome transport / regulation of TORC1 signaling / regulation of autophagosome assembly / negative regulation of autophagosome assembly / regulation of actin filament organization / guanyl-nucleotide exchange factor complex / negative regulation of immune response / axon extension / negative regulation of exocytosis / Flemming body / positive regulation of autophagosome maturation / main axon / negative regulation of macroautophagy / protein kinase inhibitor activity / positive regulation of macroautophagy / positive regulation of TOR signaling / autophagosome / axonal growth cone / stress granule assembly / GTPase activator activity / guanyl-nucleotide exchange factor activity / negative regulation of protein phosphorylation / regulation of autophagy / cell projection / negative regulation of protein kinase activity / P-body / small GTPase binding / autophagy / positive regulation of GTPase activity / cytoplasmic stress granule / endocytosis / regulation of protein localization / presynapse / postsynapse / perikaryon / nuclear membrane / lysosome / endosome / lysosomal membrane / negative regulation of gene expression / intracellular membrane-bounded organelle / dendrite / chromatin / protein kinase binding / extracellular space / nucleoplasm / nucleus / cytosol / cytoplasm
Similarity search - Function
WD repeat-containing protein 41 / Guanine nucleotide exchange factor C9orf72 / C9orf72-like protein family / Tripartite DENN C9ORF72-type domain profile. / Folliculin/SMCR8, longin domain / Folliculin/SMCR8, tripartite DENN domain / Vesicle coat protein involved in Golgi to plasma membrane transport / Tripartite DENN FLCN/SMCR8-type domain profile. / G-protein beta WD-40 repeat / WD40 repeat, conserved site ...WD repeat-containing protein 41 / Guanine nucleotide exchange factor C9orf72 / C9orf72-like protein family / Tripartite DENN C9ORF72-type domain profile. / Folliculin/SMCR8, longin domain / Folliculin/SMCR8, tripartite DENN domain / Vesicle coat protein involved in Golgi to plasma membrane transport / Tripartite DENN FLCN/SMCR8-type domain profile. / G-protein beta WD-40 repeat / WD40 repeat, conserved site / Trp-Asp (WD) repeats signature. / Trp-Asp (WD) repeats profile. / Trp-Asp (WD) repeats circular profile. / WD domain, G-beta repeat / WD40 repeats / WD40 repeat / WD40-repeat-containing domain superfamily / WD40/YVTN repeat-like-containing domain superfamily
Similarity search - Domain/homology
Guanine nucleotide exchange protein SMCR8 / Guanine nucleotide exchange factor C9orf72 / WD repeat-containing protein 41
Similarity search - Component
Biological speciesHomo sapiens (human)
Methodsingle particle reconstruction / cryo EM / Resolution: 3.2 Å
AuthorsTang D / Sheng J / Xu L / Zhan X / Yan C / Qi S
Funding support China, 3 items
OrganizationGrant numberCountry
Ministry of Science and Technology (MoST, China)2017YFA0506300 China
Ministry of Science and Technology (MoST, China)2018YFC1004601 China
National Science Foundation (NSF, China)81671388 China
CitationJournal: Proc Natl Acad Sci U S A / Year: 2020
Title: Cryo-EM structure of C9ORF72-SMCR8-WDR41 reveals the role as a GAP for Rab8a and Rab11a.
Authors: Dan Tang / Jingwen Sheng / Liangting Xu / Xiechao Zhan / Jiaming Liu / Hui Jiang / Xiaoling Shu / Xiaoyu Liu / Tizhong Zhang / Lan Jiang / Cuiyan Zhou / Wenqi Li / Wei Cheng / Zhonghan Li / ...Authors: Dan Tang / Jingwen Sheng / Liangting Xu / Xiechao Zhan / Jiaming Liu / Hui Jiang / Xiaoling Shu / Xiaoyu Liu / Tizhong Zhang / Lan Jiang / Cuiyan Zhou / Wenqi Li / Wei Cheng / Zhonghan Li / Kunjie Wang / Kefeng Lu / Chuangye Yan / Shiqian Qi /
Abstract: A massive intronic hexanucleotide repeat (GGGGCC) expansion in is a genetic origin of amyotrophic lateral sclerosis (ALS) and frontotemporal dementia (FTD). Recently, C9ORF72, together with SMCR8 ...A massive intronic hexanucleotide repeat (GGGGCC) expansion in is a genetic origin of amyotrophic lateral sclerosis (ALS) and frontotemporal dementia (FTD). Recently, C9ORF72, together with SMCR8 and WDR41, has been shown to regulate autophagy and function as Rab GEF. However, the precise function of C9ORF72 remains unclear. Here, we report the cryogenic electron microscopy (cryo-EM) structure of the human C9ORF72-SMCR8-WDR41 complex at a resolution of 3.2 Å. The structure reveals the dimeric assembly of a heterotrimer of C9ORF72-SMCR8-WDR41. Notably, the C-terminal tail of C9ORF72 and the DENN domain of SMCR8 play critical roles in the dimerization of the two protomers of the C9ORF72-SMCR8-WDR41 complex. In the protomer, C9ORF72 and WDR41 are joined by SMCR8 without direct interaction. WDR41 binds to the DENN domain of SMCR8 by the C-terminal helix. Interestingly, the prominent structural feature of C9ORF72-SMCR8 resembles that of the FLNC-FNIP2 complex, the GTPase activating protein (GAP) of RagC/D. Structural comparison and sequence alignment revealed that Arg147 of SMCR8 is conserved and corresponds to the arginine finger of FLCN, and biochemical analysis indicated that the Arg147 of SMCR8 is critical to the stimulatory effect of the C9ORF72-SMCR8 complex on Rab8a and Rab11a. Our study not only illustrates the basis of C9ORF72-SMCR8-WDR41 complex assembly but also reveals the GAP activity of the C9ORF72-SMCR8 complex.
History
DepositionJan 21, 2020-
Header (metadata) releaseApr 15, 2020-
Map releaseApr 15, 2020-
UpdateMar 27, 2024-
Current statusMar 27, 2024Processing site: PDBj / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 0.026
  • Imaged by UCSF Chimera
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  • Surface view colored by radius
  • Surface level: 0.026
  • Imaged by UCSF Chimera
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  • Surface view with fitted model
  • Atomic models: PDB-6lt0
  • Surface level: 0.026
  • Imaged by UCSF Chimera
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Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

emd_0966.png

Downloads & links

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Map

FileDownload / File: emd_0966.map.gz / Format: CCP4 / Size: 83.7 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Annotationoverall map
Voxel sizeX=Y=Z: 1.061 Å
Density
Contour LevelBy AUTHOR: 0.026 / Movie #1: 0.026
Minimum - Maximum-0.0935754 - 0.14464785
Average (Standard dev.)0.000040449377 (±0.003347591)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions280280280
Spacing280280280
CellA=B=C: 297.08 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z1.0611.0611.061
M x/y/z280280280
origin x/y/z0.0000.0000.000
length x/y/z297.080297.080297.080
α/β/γ90.00090.00090.000
start NX/NY/NZ000
NX/NY/NZ320320320
MAP C/R/S123
start NC/NR/NS000
NC/NR/NS280280280
D min/max/mean-0.0940.1450.000

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Supplemental data

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Additional map: #1

Fileemd_0966_additional_1.map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Additional map: map for N terminal domains of B chain and C chain

Fileemd_0966_additional_2.map
Annotationmap for N terminal domains of B chain and C chain
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Sample components

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Entire : Dimer of C9-ORF72-SMCR8-WDR41

EntireName: Dimer of C9-ORF72-SMCR8-WDR41
Components
  • Complex: Dimer of C9-ORF72-SMCR8-WDR41
    • Protein or peptide: WD repeat-containing protein 41
    • Protein or peptide: Guanine nucleotide exchange protein SMCR8
    • Protein or peptide: Guanine nucleotide exchange C9orf72

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Supramolecule #1: Dimer of C9-ORF72-SMCR8-WDR41

SupramoleculeName: Dimer of C9-ORF72-SMCR8-WDR41 / type: complex / ID: 1 / Parent: 0 / Macromolecule list: all
Source (natural)Organism: Homo sapiens (human)
Molecular weightTheoretical: 400 KDa

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Macromolecule #1: WD repeat-containing protein 41

MacromoleculeName: WD repeat-containing protein 41 / type: protein_or_peptide / ID: 1 / Number of copies: 2 / Enantiomer: LEVO
Source (natural)Organism: Homo sapiens (human)
Molecular weightTheoretical: 51.783805 KDa
Recombinant expressionOrganism: Spodoptera frugiperda (fall armyworm)
SequenceString: MLRWLIGGGR EPQGLAEKSP LQTIGEEQTQ NPYTELLVLK AHHDIVRFLV QLDDYRFASA GDDGIVVVWN AQTGEKLLEL NGHTQKITA IITFPSLESC EEKNQLILTA SADRTVIVWD GDTTRQVQRI SCFQSTVKCL TVLQRLDVWL SGGNDLCVWN R KLDLLCKT ...String:
MLRWLIGGGR EPQGLAEKSP LQTIGEEQTQ NPYTELLVLK AHHDIVRFLV QLDDYRFASA GDDGIVVVWN AQTGEKLLEL NGHTQKITA IITFPSLESC EEKNQLILTA SADRTVIVWD GDTTRQVQRI SCFQSTVKCL TVLQRLDVWL SGGNDLCVWN R KLDLLCKT SHLSDTGISA LVEIPKNCVV AAVGKELIIF RLVAPTEGSL EWDILEVKRL LDHQDNILSL INVNDLSFVT GS HVGELII WDALDWTMQA YERNFWDPSP QLDTQQEIKL CQKSNDISIH HFTCDEENVF AAVGRGLYVY SLQMKRVIAC QKT AHDSNV LHVARLPNRQ LISCSEDGSV RIWELREKQQ LAAEPVPTGF FNMWGFGRVS KQASQPVKKQ QENATSCSLE LIGD LIGHS SSVEMFLYFE DHGLVTCSAD HLIILWKNGE RESGLRSLRL FQKLEENGDL YLAV

UniProtKB: WD repeat-containing protein 41

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Macromolecule #2: Guanine nucleotide exchange protein SMCR8

MacromoleculeName: Guanine nucleotide exchange protein SMCR8 / type: protein_or_peptide / ID: 2 / Number of copies: 2 / Enantiomer: LEVO
Source (natural)Organism: Homo sapiens (human)
Molecular weightTheoretical: 105.149094 KDa
Recombinant expressionOrganism: Spodoptera frugiperda (fall armyworm)
SequenceString: MISAPDVVAF TKEEEYEEEP YNEPALPEEY SVPLFPFASQ GANPWSKLSG AKFSRDFILI SEFSEQVGPQ PLLTIPNDTK VFGTFDLNY FSLRIMSVDY QASFVGHPPG SAYPKLNFVE DSKVVLGDSK EGAFAYVHHL TLYDLEARGF VRPFCMAYIS A DQHKIMQQ ...String:
MISAPDVVAF TKEEEYEEEP YNEPALPEEY SVPLFPFASQ GANPWSKLSG AKFSRDFILI SEFSEQVGPQ PLLTIPNDTK VFGTFDLNY FSLRIMSVDY QASFVGHPPG SAYPKLNFVE DSKVVLGDSK EGAFAYVHHL TLYDLEARGF VRPFCMAYIS A DQHKIMQQ FQELSAEFSR ASECLKTGNR KAFAGELEKK LKDLDYTRTV LHTETEIQKK ANDKGFYSSQ AIEKANELAS VE KSIIEHQ DLLKQIRSYP HRKLKGHDLC PGEMEHIQDQ ASQASTTSNP DESADTDLYT CRPAYTPKLI KAKSTKCFDK KLK TLEELC DTEYFTQTLA QLSHIEHMFR GDLCYLLTSQ IDRALLKQQH ITNFLFEDFV EVDDRMVEKQ ESIPSKPSQD RPPS SSLEE CPIPKVLISV GSYKSSVESV LIKMEQELGD EEYKEVEVTE LSSFDPQENL DYLDMDMKGS ISSGESIEVL GTEKS TSVL SKSDSQASLT VPLSPQVVRS KAVSHRTISE DSIEVLSTCP SEALIPDDFK ASYPSAINEE ESYPDGNEGA IRFQAS ISP PELGETEEGS IENTPSQIDS SCCIGKESDG QLVLPSTPAH THSDEDGVVS SPPQRHRQKD QGFRVDFSVE NANPSSR DN SCEGFPAYEL DPSHLLASRD ISKTSLDNYS DTTSYVSSVA STSSDRIPSA YPAGLSSDRH KKRAGQNALK FIRQYPFA H PAIYSLLSGR TLVVLGEDEA IVRKLVTALA IFVPSYGCYA KPVKHWASSP LHIMDFQKWK LIGLQRVASP AGAGTLHAL SRYSRYTSIL DLDNKTLRCP LYRGTLVPRL ADHRTQIKRG STYYLHVQSM LTQLCSKAFL YTFCHHLHLP THDKETEELV ASRQMSFLK LTLGLVNEDV RVVQYLAELL KLHYMQESPG TSHPMLRFDY VPSFLYKI

UniProtKB: Guanine nucleotide exchange protein SMCR8

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Macromolecule #3: Guanine nucleotide exchange C9orf72

MacromoleculeName: Guanine nucleotide exchange C9orf72 / type: protein_or_peptide / ID: 3 / Number of copies: 2 / Enantiomer: LEVO
Source (natural)Organism: Homo sapiens (human)
Molecular weightTheoretical: 54.391477 KDa
Recombinant expressionOrganism: Spodoptera frugiperda (fall armyworm)
SequenceString: MSTLCPPPSP AVAKTEIALS GKSPLLAATF AYWDNILGPR VRHIWAPKTE QVLLSDGEIT FLANHTLNGE ILRNAESGAI DVKFFVLSE KGVIIVSLIF DGNWNGDRST YGLSIILPQT ELSFYLPLHR VCVDRLTHII RKGRIWMHKE RQENVQKIIL E GTERMEDQ ...String:
MSTLCPPPSP AVAKTEIALS GKSPLLAATF AYWDNILGPR VRHIWAPKTE QVLLSDGEIT FLANHTLNGE ILRNAESGAI DVKFFVLSE KGVIIVSLIF DGNWNGDRST YGLSIILPQT ELSFYLPLHR VCVDRLTHII RKGRIWMHKE RQENVQKIIL E GTERMEDQ GQSIIPMLTG EVIPVMELLS SMKSHSVPEE IDIADTVLND DDIGDSCHEG FLLNAISSHL QTCGCSVVVG SS AEKVNKI VRTLCLFLTP AERKCSRLCE AESSFKYESG LFVQGLLKDS TGSFVLPFRQ VMYAPYPTTH IDVDVNTVKQ MPP CHEHIY NQRRYMRSEL TAFWRATSEE DMAQDTIIYT DESFTPDLNI FQDVLHRDTL VKAFLDQVFQ LKPGLSLRST FLAQ FLLVL HRKALTLIKY IEDDTQKGKK PFKSLRNLKI DLDLTAEGDL NIIMALAEKI KPGLHSFIFG RPFYTSVQER DVLMT F

UniProtKB: Guanine nucleotide exchange factor C9orf72

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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

Concentration1 mg/mL
BufferpH: 8
Component:
ConcentrationName
25.0 mMTirs
150.0 mMNaClSodium chloride
0.5 mMTCEP
GridModel: Quantifoil R1.2/1.3 / Material: GOLD / Mesh: 300 / Support film - Material: CARBON / Support film - topology: HOLEY / Pretreatment - Type: GLOW DISCHARGE
VitrificationCryogen name: ETHANE
Detailsmonodisperse

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: SPOT SCAN / Imaging mode: DIFFRACTION
Sample stageSpecimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: HELIUM
Image recordingFilm or detector model: GATAN K2 SUMMIT (4k x 4k) / Detector mode: SUPER-RESOLUTION / Number grids imaged: 1 / Number real images: 5332 / Average exposure time: 0.25 sec. / Average electron dose: 5.3 e/Å2
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Startup modelType of model: INSILICO MODEL
Initial angle assignmentType: ANGULAR RECONSTITUTION / Software - Name: RELION
Final 3D classificationSoftware - Name: RELION
Final angle assignmentType: ANGULAR RECONSTITUTION / Software - Name: RELION
Final reconstructionApplied symmetry - Point group: C1 (asymmetric) / Resolution.type: BY AUTHOR / Resolution: 3.2 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: RELION / Number images used: 347925

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Atomic model buiding 1

DetailsThe atomic coordinates of the CSW complex was generated by combining homology modelling and de novo model building.
RefinementSpace: REAL / Protocol: RIGID BODY FIT
Output model

PDB-6lt0:
cryo-EM structure of C9ORF72-SMCR8-WDR41

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