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- EMDB-47291: Cryo-EM structure of yeast Exportin Msn5 bound to RanGTP and Pho4... -

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Basic information

Entry
Database: EMDB / ID: EMD-47291
TitleCryo-EM structure of yeast Exportin Msn5 bound to RanGTP and Pho4 (not modeled) (State 2-1)
Map datastructure of yeast Exportin Msn5 bound to RanGTP and Pho4 (not modeled) (State 2-1)
Sample
  • Complex: Msn5-Gsp1-pPho4(1-200)
    • Protein or peptide: Protein MSN5
  • Protein or peptide: GTP-binding nuclear protein GSP1/CNR1
  • Ligand: GUANOSINE-5'-TRIPHOSPHATE
  • Ligand: MAGNESIUM ION
KeywordsKaryopherin / Exportin / Nuclear Export / TRANSPORT PROTEIN
Function / homology
Function and homology information


regulation of cell cycle phase transition / regulation of nucleocytoplasmic transport / Postmitotic nuclear pore complex (NPC) reformation / exonucleolytic trimming to generate mature 3'-end of 5.8S rRNA from tricistronic rRNA transcript (SSU-rRNA, 5.8S rRNA, LSU-rRNA) / tRNA re-export from nucleus / RNA nuclear export complex / nuclear export signal receptor activity / RNA export from nucleus / poly(A)+ mRNA export from nucleus / nucleus organization ...regulation of cell cycle phase transition / regulation of nucleocytoplasmic transport / Postmitotic nuclear pore complex (NPC) reformation / exonucleolytic trimming to generate mature 3'-end of 5.8S rRNA from tricistronic rRNA transcript (SSU-rRNA, 5.8S rRNA, LSU-rRNA) / tRNA re-export from nucleus / RNA nuclear export complex / nuclear export signal receptor activity / RNA export from nucleus / poly(A)+ mRNA export from nucleus / nucleus organization / ribosomal subunit export from nucleus / protein export from nucleus / protein import into nucleus / GTPase activity / GTP binding / RNA binding / nucleus / cytosol / cytoplasm
Similarity search - Function
Exportin-5, C-terminal domain / Exportin-5 family / Exportin-1/5 / small GTPase Ran family profile. / Ran GTPase / Ran (Ras-related nuclear proteins) /TC4 subfamily of small GTPases / Rho (Ras homology) subfamily of Ras-like small GTPases / Ras subfamily of RAS small GTPases / Small GTPase / Ras family ...Exportin-5, C-terminal domain / Exportin-5 family / Exportin-1/5 / small GTPase Ran family profile. / Ran GTPase / Ran (Ras-related nuclear proteins) /TC4 subfamily of small GTPases / Rho (Ras homology) subfamily of Ras-like small GTPases / Ras subfamily of RAS small GTPases / Small GTPase / Ras family / Rab subfamily of small GTPases / Armadillo-like helical / Small GTP-binding protein domain / Armadillo-type fold / P-loop containing nucleoside triphosphate hydrolase
Similarity search - Domain/homology
GTP-binding nuclear protein GSP1/CNR1 / Protein MSN5
Similarity search - Component
Biological speciesSaccharomyces cerevisiae (brewer's yeast)
Methodsingle particle reconstruction / cryo EM / Resolution: 3.2 Å
AuthorsFung HYJ / Chook YM
Funding support United States, 5 items
OrganizationGrant numberCountry
Cancer Prevention and Research Institute of Texas (CPRIT)RP220582 United States
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)R35GM141461 United States
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)R01GM069909 United States
Welch FoundationI-1532 United States
Other privateUTSW Gilman Special Opportunities Award
CitationJournal: Nat Commun / Year: 2025
Title: Phosphate-dependent nuclear export via a non-classical NES class recognized by exportin Msn5.
Authors: Ho Yee Joyce Fung / Sanraj R Mittal / Ashley B Niesman / Jenny Jiou / Binita Shakya / Takuya Yoshizawa / Ahmet E Cansizoglu / Michael P Rout / Yuh Min Chook /
Abstract: Gene expression in response to environmental stimuli is dependent on nuclear localization of key signaling components, which can be tightly regulated by phosphorylation. This is exemplified by the ...Gene expression in response to environmental stimuli is dependent on nuclear localization of key signaling components, which can be tightly regulated by phosphorylation. This is exemplified by the phosphate-sensing transcription factor Pho4, which requires phosphorylation for nuclear export by the yeast exportin Msn5. Here, we present a high resolution cryogenic-electron microscopy structure showing the phosphorylated 35-residue nuclear export signal of Pho4, which binds the concave surface of Msn5 through two Pho4 phospho-serines that align with two Msn5 basic patches. These findings characterize a mechanism of phosphate-specific recognition mediated by a non-classical signal distinct from that for Exportin-1. Furthermore, the discovery that unliganded Msn5 is autoinhibited explains the positive cooperativity of Pho4/Ran-binding and proposes a mechanism for Pho4's release in the cytoplasm. These findings advance our understanding of the diversity of signals that drive nuclear export and how cargo phosphorylation is crucial in regulating nuclear transport and controlling cellular signaling pathways.
History
DepositionOct 11, 2024-
Header (metadata) releaseMar 19, 2025-
Map releaseMar 19, 2025-
UpdateApr 2, 2025-
Current statusApr 2, 2025Processing site: RCSB / Status: Released

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Structure visualization

Supplemental images

emd_47291.png

Downloads & links

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Map

FileDownload / File: emd_47291.map.gz / Format: CCP4 / Size: 160.8 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Annotationstructure of yeast Exportin Msn5 bound to RanGTP and Pho4 (not modeled) (State 2-1)
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
0.83 Å/pix.
x 348 pix.
= 287.448 Å		0.83 Å/pix.
x 348 pix.
= 287.448 Å		0.83 Å/pix.
x 348 pix.
= 287.448 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 0.826 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.148
Minimum - Maximum-0.19615178 - 0.57930595
Average (Standard dev.)0.0016406447 (±0.01485548)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions348348348
Spacing348348348
CellA=B=C: 287.448 Å
α=β=γ: 90.0 °

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Supplemental data

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Half map: Half Map B

Fileemd_47291_half_map_1.map
AnnotationHalf Map B
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: Half Map A

Fileemd_47291_half_map_2.map
AnnotationHalf Map A
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Sample components

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Entire : Msn5-Gsp1-pPho4(1-200)

EntireName: Msn5-Gsp1-pPho4(1-200)
Components
  • Complex: Msn5-Gsp1-pPho4(1-200)
    • Protein or peptide: Protein MSN5
  • Protein or peptide: GTP-binding nuclear protein GSP1/CNR1
  • Ligand: GUANOSINE-5'-TRIPHOSPHATE
  • Ligand: MAGNESIUM ION

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Supramolecule #1: Msn5-Gsp1-pPho4(1-200)

SupramoleculeName: Msn5-Gsp1-pPho4(1-200) / type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1
Details: Msn5 bound to RanGTP (Gsp1) and phosphorylated Pho4 fragment (1-200)
Source (natural)Organism: Saccharomyces cerevisiae (brewer's yeast)

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Macromolecule #1: Protein MSN5

MacromoleculeName: Protein MSN5 / type: protein_or_peptide / ID: 1 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Saccharomyces cerevisiae (brewer's yeast)
Molecular weightTheoretical: 143.067188 KDa
Recombinant expressionOrganism: Escherichia coli (E. coli)
SequenceString: MDSTGASQIV SALDVIYSPK SNNSQRQEAQ KFLDEVKLCS ESPFWGYEIA LQNPTNSILK YFGLGLLDHA VKKNWNDYDE GKRVALRKW VMELNFGVQD YDTRYIKEKL ATLWVEVAKR TWGEALKQTN PTEEQLLTSW VDMDNNLFEL WNINQSSREL A LIIFRILF ...String:
MDSTGASQIV SALDVIYSPK SNNSQRQEAQ KFLDEVKLCS ESPFWGYEIA LQNPTNSILK YFGLGLLDHA VKKNWNDYDE GKRVALRKW VMELNFGVQD YDTRYIKEKL ATLWVEVAKR TWGEALKQTN PTEEQLLTSW VDMDNNLFEL WNINQSSREL A LIIFRILF EDVFLLDDLI VLKRMTVIQP LCVMIVCPIE VFAIKYKFSD KWTKFKANEE GWFSVWIPEL NNALQQNNSE YI IRLLETL KTCLNWPLTE VIVRNDVLSS LLTCLSSNIP RAQSMALDSI HILLTRPYSN ESHYQMTIDR VFDNMDLLDS VYE SLLFDP TDDIDETKYP IIKKFVDMIS CLYVCVPKIK ETNGQIQKYF KLVLKTTYNP SLIVSGLTLD LWCTCLRNDE YLPK LEKYV IPDLLQFAAD ALVYYEQIDG HISKKFAEID FQSKSEFQTF CSTYRKRIRD IIRLISCVEL DLTYDWLNNR LNNYF SSPF GQQVLSSTFL DHKLEPYLGA LSQYMIVECF INGCIRWKIW YPTGDDYDEK LDSILQKLEI LSNQLIALNL REPLLL KKQ IQNFALFLTM LKDNVLFTLL EKIITSATMD YPEINLEERG AESDAVRDLR YACGIELNRM ALLMPESLKK IYPDLES VI ARIMPNLSYH EKISFKSFLL IIVLKSSLDM KEERFAAIVD PELLAWSDKT TVVGLSDLHW FMERLGIVQI AEYFQRRD I DENSDLLSIP IDDEGKELKS ELTKRWQSLF PVRATRMFIH YSMQSIKTDE EFKMLQDLWR PRIVPILPYI TRLLYQLQS YHDPDNWKGL PTVVQSFVKY STIERFWEAG ASNKSKDEFI DEHMKAMQTL RDFADSVGHI IRYTREYTLL VLSAISSLGS VFYLLDESP DLLLNSIAIF KPGSNEISPG VSTHGWKHIM NIAIRPILKG CPKDCLGKFM PAFLPKLFEI LDLLLCQKWS S HMNDMDMN PVPTDDDQMT EEILEENLLR QLTTVVVRIV IDCVGQGNAN PNSAKSRLNN HQMEMRKIIF NDLNTLAPFL KL LNHLISF KDTKCSFNSI LVMKCCLTSV LNQNNTVDEY FTFEVMKNLL LNVLCNSAFK DSFHEALYAF TVIFLTLCKE YPS ARAFLF EISNGYNIDE LYRNLRSVDE YKTQRALMID FIDWVKSTSG KEDGNVDHAG DERKRQEKRE AILKKANERL IKKN KENGD MLDDPNIEDG AVGNLFDDNE NLYFQ

UniProtKB: Protein MSN5

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Macromolecule #2: GTP-binding nuclear protein GSP1/CNR1

MacromoleculeName: GTP-binding nuclear protein GSP1/CNR1 / type: protein_or_peptide / ID: 2 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Saccharomyces cerevisiae (brewer's yeast)
Molecular weightTheoretical: 21.28352 KDa
Recombinant expressionOrganism: Escherichia coli (E. coli)
SequenceString:
MASAPAANGE VPTFKLVLVG DGGTGKTTFV KRHLTGEFEK KYIATIGVEV HPLSFYTNFG EIKFDVWDTA GLEKFGGLRD GYYINAQCA IIMFDVTSRI TYKNVPNWHR DLVRVCENIP IVLCGNKVDV KERKVKAKTI TFHRKKNLQY YDISAKSNYN F EKPFLWLA RKLAGNPQLE FVENLYFQ

UniProtKB: GTP-binding nuclear protein GSP1/CNR1

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Macromolecule #3: GUANOSINE-5'-TRIPHOSPHATE

MacromoleculeName: GUANOSINE-5'-TRIPHOSPHATE / type: ligand / ID: 3 / Number of copies: 1 / Formula: GTP
Molecular weightTheoretical: 523.18 Da
Chemical component information

ChemComp-GTP:
GUANOSINE-5'-TRIPHOSPHATE / GTP, energy-carrying molecule*YM

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Macromolecule #4: MAGNESIUM ION

MacromoleculeName: MAGNESIUM ION / type: ligand / ID: 4 / Number of copies: 1 / Formula: MG
Molecular weightTheoretical: 24.305 Da

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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

BufferpH: 7.4
GridModel: Quantifoil R1.2/1.3 / Material: COPPER / Mesh: 300 / Pretreatment - Type: GLOW DISCHARGE / Pretreatment - Time: 60 sec.
VitrificationCryogen name: ETHANE

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Electron microscopy

MicroscopeTFS KRIOS
Image recordingFilm or detector model: GATAN K3 (6k x 4k) / Average electron dose: 50.0 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Nominal defocus max: 2.5 µm / Nominal defocus min: 0.8 µm
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Startup modelType of model: INSILICO MODEL
Final reconstructionResolution.type: BY AUTHOR / Resolution: 3.2 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: cryoSPARC (ver. 3) / Number images used: 102289
Initial angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: cryoSPARC (ver. 3)
Final angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: cryoSPARC (ver. 3)
Final 3D classificationNumber classes: 6
FSC plot (resolution estimation)

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Atomic model buiding 1

Initial model
PDB IDChainDetails

3m1i

chain_id: A, source_name: PDB, initial_model_type: experimental modelGsp1 from PDB entry 3M1I
source_name: SwissModel, initial_model_type: in silico modelMsn5 model generated based on PDB entry 5YU7
Output model

PDB-9dxm:
Cryo-EM structure of yeast Exportin Msn5 bound to RanGTP and Pho4 (not modeled) (State 2-1)

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