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- EMDB-46557: Cryo-EM map of yeast Exportin Msn5 bound to cargo Pho4 and RanGTP... -

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Basic information

Entry
Database: EMDB / ID: EMD-46557
TitleCryo-EM map of yeast Exportin Msn5 bound to cargo Pho4 and RanGTP (State 1-2)
Map dataMain map
Sample
  • Complex: Msn5-Gsp1-pPho4(1-200)
    • Protein or peptide: Msn5
    • Protein or peptide: Gsp1
    • Protein or peptide: Pho4
KeywordsKaryopherin / Exportin / Nuclear Export / TRANSPORT PROTEIN
Biological speciesSaccharomyces cerevisiae (brewer's yeast)
Methodsingle particle reconstruction / cryo EM / Resolution: 3.37 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
DepositionAug 12, 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_46557.png

Downloads & links

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Map

FileDownload / File: emd_46557.map.gz / Format: CCP4 / Size: 160.8 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationMain map
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: 0.12
Minimum - Maximum-0.23910686 - 0.5489371
Average (Standard dev.)0.0015390625 (±0.013733728)
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 A

Fileemd_46557_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_46557_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 : Msn5-Gsp1-pPho4(1-200)

EntireName: Msn5-Gsp1-pPho4(1-200)
Components
  • Complex: Msn5-Gsp1-pPho4(1-200)
    • Protein or peptide: Msn5
    • Protein or peptide: Gsp1
    • Protein or peptide: Pho4

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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: all
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: Msn5

MacromoleculeName: Msn5 / type: protein_or_peptide / ID: 1 / Enantiomer: LEVO
Source (natural)Organism: Saccharomyces cerevisiae (brewer's yeast)
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

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Macromolecule #2: Gsp1

MacromoleculeName: Gsp1 / type: protein_or_peptide / ID: 2 / Enantiomer: LEVO
Source (natural)Organism: Saccharomyces cerevisiae (brewer's yeast)
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

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Macromolecule #3: Pho4

MacromoleculeName: Pho4 / type: protein_or_peptide / ID: 3 / Details: In vitro phosphorylated / Enantiomer: LEVO
Source (natural)Organism: Saccharomyces cerevisiae (brewer's yeast)
Recombinant expressionOrganism: Escherichia coli (E. coli)
SequenceString: GSMGRTTSEG IHGFVDDLEP KSSILDKVGD FITVNTKRHD GREDFNEQND ELNSQENHNS SENGNENENE QDSLALDDLD RAFELVEGM DMDWMMPSHA HH(SEP)PATTATI KPRLLY(SEP)PLI HTQSAVPVTI (SEP)PNLVATATS TTSANKVTK NKSNS(SEP)PYLN ...String:
GSMGRTTSEG IHGFVDDLEP KSSILDKVGD FITVNTKRHD GREDFNEQND ELNSQENHNS SENGNENENE QDSLALDDLD RAFELVEGM DMDWMMPSHA HH(SEP)PATTATI KPRLLY(SEP)PLI HTQSAVPVTI (SEP)PNLVATATS TTSANKVTK NKSNS(SEP)PYLN KRRGKPGPDS ATSLFELPDS VIPTPKPKPK PKQYPKVILP SNST

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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
VitrificationCryogen name: ETHANE

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Electron microscopy

MicroscopeFEI TITAN 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: 3.37 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: cryoSPARC (ver. 3) / Number images used: 93291
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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