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- EMDB-11973: Focused refined map at the interface of the transcribing Pol II-U... -

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Basic information

Entry
Database: EMDB / ID: EMD-11973
TitleFocused refined map at the interface of the transcribing Pol II-U1 snRNP complex with 145-nt RNA
Map dataSharpened map, re-sampled onto the overall map
Sample
  • Complex: The transcribing RNA polymerase II-U1 snRNP complex with 145-nt RNA
    • Complex: RNA polymerase II
    • Complex: Nucleic acidsNucleic acid
    • Complex: U1 snRNP
Function / homology
Function and homology information


negative regulation of protein refolding / regulation of ATP-dependent activity / U2 snRNP binding / U7 snRNA binding / histone pre-mRNA DCP binding / U7 snRNP / histone pre-mRNA 3'end processing complex / SLBP independent Processing of Histone Pre-mRNAs / SLBP Dependent Processing of Replication-Dependent Histone Pre-mRNAs / methylosome ...negative regulation of protein refolding / regulation of ATP-dependent activity / U2 snRNP binding / U7 snRNA binding / histone pre-mRNA DCP binding / U7 snRNP / histone pre-mRNA 3'end processing complex / SLBP independent Processing of Histone Pre-mRNAs / SLBP Dependent Processing of Replication-Dependent Histone Pre-mRNAs / methylosome / protein methylation / U12-type spliceosomal complex / 7-methylguanosine cap hypermethylation / pICln-Sm protein complex / U1 snRNP binding / B-WICH complex positively regulates rRNA expression / RNA Polymerase I Transcription Initiation / RNA Polymerase I Promoter Escape / RNA Polymerase I Transcription Termination / RNA Polymerase III Transcription Initiation From Type 1 Promoter / RNA Polymerase III Transcription Initiation From Type 2 Promoter / RNA Polymerase III Transcription Initiation From Type 3 Promoter / Formation of RNA Pol II elongation complex / Formation of the Early Elongation Complex / Transcriptional regulation by small RNAs / RNA Polymerase II Pre-transcription Events / TP53 Regulates Transcription of DNA Repair Genes / FGFR2 alternative splicing / RNA polymerase II transcribes snRNA genes / mRNA Capping / mRNA Splicing - Major Pathway / mRNA Splicing - Minor Pathway / Processing of Capped Intron-Containing Pre-mRNA / RNA Polymerase II Promoter Escape / RNA Polymerase II Transcription Pre-Initiation And Promoter Opening / RNA Polymerase II Transcription Initiation / RNA Polymerase II Transcription Elongation / RNA Polymerase II Transcription Initiation And Promoter Clearance / RNA Pol II CTD phosphorylation and interaction with CE / Estrogen-dependent gene expression / Formation of TC-NER Pre-Incision Complex / Dual incision in TC-NER / Gap-filling DNA repair synthesis and ligation in TC-NER / spliceosomal tri-snRNP complex / small nuclear ribonucleoprotein complex / P granule / SMN-Sm protein complex / positive regulation of mRNA splicing, via spliceosome / U2-type spliceosomal complex / telomerase RNA binding / telomerase holoenzyme complex / U2-type precatalytic spliceosome / negative regulation of chaperone-mediated autophagy / : / U2-type prespliceosome assembly / commitment complex / U2-type catalytic step 2 spliceosome / U4 snRNP / U2 snRNP / RNA Polymerase II Transcription Termination / U1 snRNP / U2-type prespliceosome / : / positive regulation of nuclear-transcribed mRNA poly(A) tail shortening / precatalytic spliceosome / spliceosomal complex assembly / maintenance of transcriptional fidelity during transcription elongation by RNA polymerase II / mRNA Splicing - Minor Pathway / regulation of RNA splicing / positive regulation of translational initiation / RNA polymerase II activity / organelle membrane / transcription-coupled nucleotide-excision repair / U5 snRNP / RNA polymerase I complex / transcription by RNA polymerase I / RNA polymerase III complex / transcription by RNA polymerase III / spliceosomal snRNP assembly / RNA polymerase II, core complex / translation initiation factor binding / U1 snRNA binding / U4/U6 x U5 tri-snRNP complex / catalytic step 2 spliceosome / mRNA Splicing - Major Pathway / positive regulation of RNA splicing / DNA-directed RNA polymerase complex / RNA splicing / promoter-specific chromatin binding / P-body / transcription initiation at RNA polymerase II promoter / spliceosomal complex / ribonucleoside binding / mRNA splicing, via spliceosome / fibrillar center / DNA-directed 5'-3' RNA polymerase activity / DNA-directed RNA polymerase / single-stranded DNA binding / chromosome / snRNP Assembly
Similarity search - Function
snRNP70, RNA recognition motif / U1 small nuclear ribonucleoprotein of 70kDa N-terminal / U1 small nuclear ribonucleoprotein of 70kDa MW N terminal / Small ribonucleoprotein associated, SmB/SmN / U1 small nuclear ribonucleoprotein A, RNA recognition motif 2 / U1 small nuclear ribonucleoprotein A, RNA recognition motif 1 / Small nuclear ribonucleoprotein D1 / RNA-binding domain, S1 / Small nuclear ribonucleoprotein Sm D3 / Small nuclear ribonucleoprotein Sm D2 ...snRNP70, RNA recognition motif / U1 small nuclear ribonucleoprotein of 70kDa N-terminal / U1 small nuclear ribonucleoprotein of 70kDa MW N terminal / Small ribonucleoprotein associated, SmB/SmN / U1 small nuclear ribonucleoprotein A, RNA recognition motif 2 / U1 small nuclear ribonucleoprotein A, RNA recognition motif 1 / Small nuclear ribonucleoprotein D1 / RNA-binding domain, S1 / Small nuclear ribonucleoprotein Sm D3 / Small nuclear ribonucleoprotein Sm D2 / Small nuclear ribonucleoprotein E / Small nuclear ribonucleoprotein G / Small nuclear ribonucleoprotein F / Sm-like protein Lsm7/SmG / Like-Sm (LSM) domain containing protein, LSm4/SmD1/SmD3 / Sm-like protein Lsm6/SmF / LSM domain / LSM domain, eukaryotic/archaea-type / snRNP Sm proteins / : / Sm domain profile. / LSM domain superfamily / DNA-directed RNA polymerase II subunit Rpb4-like / RNA polymerase Rpb4/RPC9, core / DNA-directed RNA-polymerase II subunit / RNA polymerase II, heptapeptide repeat, eukaryotic / RNA polymerase Rpb1, domain 6 / RNA polymerase Rpb1, domain 6 / RNA polymerase Rpb1 C-terminal repeat / Eukaryotic RNA polymerase II heptapeptide repeat. / RNA polymerase Rpb1, domain 7 / RNA polymerase Rpb1, domain 7 superfamily / RNA polymerase Rpb1, domain 7 / Pol II subunit B9, C-terminal zinc ribbon / RNA polymerase RBP11 / Rpb4/RPC9 superfamily / RNA polymerase subunit Rpb4/RPC9 / RNA polymerase Rpb4 / Zinc finger TFIIS-type signature. / HRDC-like superfamily / RNA polymerase subunit Rpb7-like / RNA polymerase Rpb7-like , N-terminal / RNA polymerase Rpb7-like, N-terminal domain superfamily / SHS2 domain found in N terminus of Rpb7p/Rpc25p/MJ0397 / RNA polymerase Rpb2, domain 4 / RNA polymerase Rpb2, domain 4 / RNA polymerase Rpb2, domain 5 / RNA polymerase Rpb2, domain 5 / DNA-directed RNA polymerase subunit RPABC5/Rpb10 / RNA polymerases, subunit N, zinc binding site / RNA polymerase subunit RPB10 / RNA polymerases N / 8 kDa subunit / RNA polymerases N / 8 Kd subunits signature. / DNA-directed RNA polymerase, M/15kDa subunit / RNA polymerases M/15 Kd subunit / RNA polymerase subunit 9 / DNA-directed RNA polymerase M, 15kDa subunit, conserved site / RNA polymerases M / 15 Kd subunits signature. / DNA-directed RNA polymerase subunit/transcription factor S / RNA polymerase, Rpb8 / DNA-directed RNA polymerases I, II, and III subunit RPABC4 / RNA polymerase Rpb8 / RNA polymerase subunit 8 / RNA polymerase, Rpb5, N-terminal / RNA polymerase Rpb5, N-terminal domain superfamily / RNA polymerase Rpb5, N-terminal domain / DNA-directed RNA polymerase, subunit RPB6 / DNA-directed RNA polymerase subunit Rpo11 / DNA directed RNA polymerase, 7 kDa subunit / RNA polymerase archaeal subunit P/eukaryotic subunit RPABC4 / RNA polymerase, subunit H/Rpb5, conserved site / RNA polymerases H / 23 Kd subunits signature. / RNA polymerase subunit CX / DNA-directed RNA polymerase Rpb11, 13-16kDa subunit, conserved site / DNA-directed RNA polymerase subunit Rpo5/Rpb5 / RNA polymerases L / 13 to 16 Kd subunits signature. / DNA-directed RNA polymerase, 30-40kDa subunit, conserved site / DNA-directed RNA polymerase subunit Rpo3/Rpb3/RPAC1 / RNA polymerases D / 30 to 40 Kd subunits signature. / DNA-directed RNA polymerase, RBP11-like dimerisation domain / RNA polymerase Rpb3/Rpb11 dimerisation domain / RNA polymerase, subunit H/Rpb5 C-terminal / RNA polymerase subunit RPABC4/transcription elongation factor Spt4 / RPB5-like RNA polymerase subunit superfamily / RNA polymerase Rpb5, C-terminal domain / Zinc finger, TFIIS-type / Transcription factor S-II (TFIIS) / Zinc finger TFIIS-type profile. / C2C2 Zinc finger / Archaeal Rpo6/eukaryotic RPB6 RNA polymerase subunit / DNA-directed RNA polymerase, 14-18kDa subunit, conserved site / RNA polymerases K / 14 to 18 Kd subunits signature. / Ribosomal protein S1-like RNA-binding domain / S1 RNA binding domain / S1 domain / RNA recognition motif / RNA recognition motif / Eukaryotic RNA Recognition Motif (RRM) profile. / RNA recognition motif domain / RNA polymerase Rpb6
Similarity search - Domain/homology
DNA-directed RNA polymerase subunit beta / RNA polymerase II subunit D / DNA-directed RNA polymerases I, II, and III subunit RPABC5 / DNA-directed RNA polymerase II subunit RPB11-a / DNA-directed RNA polymerases I, II, and III subunit RPABC2 / DNA-directed RNA polymerases I, II, and III subunit RPABC3 / DNA-directed RNA polymerase II subunit RPB3 / DNA-directed RNA polymerase II subunit RPB7 / RNA polymerase II, I and III subunit K / DNA-directed RNA polymerase II subunit E ...DNA-directed RNA polymerase subunit beta / RNA polymerase II subunit D / DNA-directed RNA polymerases I, II, and III subunit RPABC5 / DNA-directed RNA polymerase II subunit RPB11-a / DNA-directed RNA polymerases I, II, and III subunit RPABC2 / DNA-directed RNA polymerases I, II, and III subunit RPABC3 / DNA-directed RNA polymerase II subunit RPB3 / DNA-directed RNA polymerase II subunit RPB7 / RNA polymerase II, I and III subunit K / DNA-directed RNA polymerase II subunit E / U1 small nuclear ribonucleoprotein 70 kDa / U1 small nuclear ribonucleoprotein A / DNA-directed RNA polymerase II subunit RPB1 / DNA-directed RNA polymerase II subunit RPB9 / Small nuclear ribonucleoprotein E / Small nuclear ribonucleoprotein F / Small nuclear ribonucleoprotein G / Small nuclear ribonucleoprotein Sm D1 / Small nuclear ribonucleoprotein Sm D2 / Small nuclear ribonucleoprotein Sm D3 / Small nuclear ribonucleoprotein-associated protein
Similarity search - Component
Biological speciesSus scrofa domesticus (domestic pig) / synthetic construct (others) / Homo sapiens (human)
Methodsingle particle reconstruction / cryo EM / Resolution: 3.2 Å
AuthorsZhang S / Aibara S / Vos SM / Agafonov DE / Luehrmann R / Cramer P
Funding support4 items
OrganizationGrant numberCountry
European Molecular Biology Organization (EMBO)ALTF 830-2018
European Research Council (ERC)882357
H2020 Marie Curie Actions of the European Commission894862
German Research Foundation (DFG)EXC 2067/1-390729940
CitationJournal: Science / Year: 2021
Title: Structure of a transcribing RNA polymerase II-U1 snRNP complex.
Authors: Suyang Zhang / Shintaro Aibara / Seychelle M Vos / Dmitry E Agafonov / Reinhard Lührmann / Patrick Cramer /
Abstract: To initiate cotranscriptional splicing, RNA polymerase II (Pol II) recruits the U1 small nuclear ribonucleoprotein particle (U1 snRNP) to nascent precursor messenger RNA (pre-mRNA). Here, we report ...To initiate cotranscriptional splicing, RNA polymerase II (Pol II) recruits the U1 small nuclear ribonucleoprotein particle (U1 snRNP) to nascent precursor messenger RNA (pre-mRNA). Here, we report the cryo-electron microscopy structure of a mammalian transcribing Pol II-U1 snRNP complex. The structure reveals that Pol II and U1 snRNP interact directly. This interaction positions the pre-mRNA 5' splice site near the RNA exit site of Pol II. Extension of pre-mRNA retains the 5' splice site, leading to the formation of a "growing intron loop." Loop formation may facilitate scanning of nascent pre-mRNA for the 3' splice site, functional pairing of distant intron ends, and prespliceosome assembly. Our results provide a starting point for a mechanistic analysis of cotranscriptional spliceosome assembly and the biogenesis of mRNA isoforms by alternative splicing.
History
DepositionNov 23, 2020-
Header (metadata) releaseJan 27, 2021-
Map releaseJan 27, 2021-
UpdateJan 27, 2021-
Current statusJan 27, 2021Processing site: PDBe / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 5.6
  • Imaged by UCSF Chimera
  • Download
  • Surface view colored by radius
  • Surface level: 5.6
  • Imaged by UCSF Chimera
  • Download
Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

Downloads & links

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Map

FileDownload / File: emd_11973.map.gz / Format: CCP4 / Size: 244.1 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationSharpened map, re-sampled onto the overall map
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
1.05 Å/pix.
x 400 pix.
= 420. Å
1.05 Å/pix.
x 400 pix.
= 420. Å
1.05 Å/pix.
x 400 pix.
= 420. Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 1.05 Å
Density
Contour LevelBy AUTHOR: 5.6 / Movie #1: 5.6
Minimum - Maximum-29.095215 - 49.45513
Average (Standard dev.)-0.0037216016 (±0.8613858)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions400400400
Spacing400400400
CellA=B=C: 419.99997 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z1.051.051.05
M x/y/z400400400
origin x/y/z0.0000.0000.000
length x/y/z420.000420.000420.000
α/β/γ90.00090.00090.000
start NX/NY/NZ1081340
NX/NY/NZ13584349
MAP C/R/S123
start NC/NR/NS000
NC/NR/NS400400400
D min/max/mean-29.09549.455-0.004

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Supplemental data

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Mask #1

Fileemd_11973_msk_1.map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

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Additional map: Original sharpened map

Fileemd_11973_additional_1.map
AnnotationOriginal sharpened map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

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Half map: Half map 1

Fileemd_11973_half_map_1.map
AnnotationHalf map 1
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

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Half map: Half map 2

Fileemd_11973_half_map_2.map
AnnotationHalf map 2
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

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Sample components

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Entire : The transcribing RNA polymerase II-U1 snRNP complex with 145-nt RNA

EntireName: The transcribing RNA polymerase II-U1 snRNP complex with 145-nt RNA
Components
  • Complex: The transcribing RNA polymerase II-U1 snRNP complex with 145-nt RNA
    • Complex: RNA polymerase II
    • Complex: Nucleic acidsNucleic acid
    • Complex: U1 snRNP

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Supramolecule #1: The transcribing RNA polymerase II-U1 snRNP complex with 145-nt RNA

SupramoleculeName: The transcribing RNA polymerase II-U1 snRNP complex with 145-nt RNA
type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1-#25

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Supramolecule #2: RNA polymerase II

SupramoleculeName: RNA polymerase II / type: complex / ID: 2 / Parent: 1 / Macromolecule list: #1-#12
Source (natural)Organism: Sus scrofa domesticus (domestic pig)

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Supramolecule #3: Nucleic acids

SupramoleculeName: Nucleic acids / type: complex / ID: 3 / Parent: 1 / Macromolecule list: #13-#15
Source (natural)Organism: synthetic construct (others)
Recombinant expressionOrganism: synthetic construct (others)

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Supramolecule #4: U1 snRNP

SupramoleculeName: U1 snRNP / type: complex / ID: 4 / Parent: 1 / Macromolecule list: #17-#25
Source (natural)Organism: Homo sapiens (human)

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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: 7.5
Component:
ConcentrationFormula
20.0 mMHEPES
150.0 mMNaClSodium chloride
3.0 mMMgCl2
0.5 mMTCEP
GridModel: UltrAuFoil R2/2 / Pretreatment - Type: GLOW DISCHARGE
VitrificationCryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 277.15 K / Instrument: FEI VITROBOT MARK IV

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELDBright-field microscopy
Specialist opticsEnergy filter - Slit width: 20 eV
Image recordingFilm or detector model: GATAN K3 (6k x 4k) / Average electron dose: 1.01 e/Å2
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

CTF correctionSoftware: (Name: Warp, RELION)
Initial angle assignmentType: MAXIMUM LIKELIHOOD
Final angle assignmentType: MAXIMUM LIKELIHOOD
Final reconstructionResolution.type: BY AUTHOR / Resolution: 3.2 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: RELION / Number images used: 61596
FSC plot (resolution estimation)

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Atomic model buiding 1

Initial model(PDB ID:
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