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Open data
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Basic information
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Title | Cryo-EM structure of the cross-exon pre-B+AMPPNP complex | |||||||||
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![]() | spliceosome / SPLICING | |||||||||
Function / homology | ![]() negative regulation of protein refolding / Lsm2-8 complex / U6 snRNA 3'-end binding / spliceosomal snRNP complex / ribonucleoprotein complex localization / U4atac snRNP / positive regulation of cytotoxic T cell differentiation / maturation of 5S rRNA / positive regulation of hippo signaling / RNA localization ...negative regulation of protein refolding / Lsm2-8 complex / U6 snRNA 3'-end binding / spliceosomal snRNP complex / ribonucleoprotein complex localization / U4atac snRNP / positive regulation of cytotoxic T cell differentiation / maturation of 5S rRNA / positive regulation of hippo signaling / RNA localization / R-loop processing / U4atac snRNA binding / regulation of ATP-dependent activity / mRNA decay by 5' to 3' exoribonuclease / Lsm1-7-Pat1 complex / regulation of mitotic cell cycle spindle assembly checkpoint / U6 snRNP / box C/D sno(s)RNA binding / U11/U12 snRNP / PH domain binding / dense fibrillar component / U2 snRNP binding / U7 snRNA binding / histone pre-mRNA DCP binding / U7 snRNP / B-WICH complex / histone pre-mRNA 3'end processing complex / cis assembly of pre-catalytic spliceosome / SLBP independent Processing of Histone Pre-mRNAs / SLBP Dependent Processing of Replication-Dependent Histone Pre-mRNAs / splicing factor binding / U4/U6 snRNP / spliceosome conformational change to release U4 (or U4atac) and U1 (or U11) / protein methylation / U12-type spliceosomal complex / methylosome / 7-methylguanosine cap hypermethylation / snRNP binding / U1 snRNP binding / P-body assembly / blastocyst formation / pICln-Sm protein complex / RNA splicing, via transesterification reactions / small nuclear ribonucleoprotein complex / U2-type catalytic step 1 spliceosome / U4 snRNA binding / sno(s)RNA-containing ribonucleoprotein complex / box C/D methylation guide snoRNP complex / SMN-Sm protein complex / negative regulation of chaperone-mediated autophagy / spliceosomal tri-snRNP complex / P granule / telomerase holoenzyme complex / mRNA cis splicing, via spliceosome / telomerase RNA binding / U2-type precatalytic spliceosome / U2-type spliceosomal complex / commitment complex / U2-type prespliceosome assembly / U2-type catalytic step 2 spliceosome / positive regulation of mRNA splicing, via spliceosome / box C/D snoRNP assembly / U4 snRNP / rRNA modification in the nucleus and cytosol / U2 snRNP / RNA Polymerase II Transcription Termination / SAGA complex / U3 snoRNA binding / positive regulation of transcription by RNA polymerase III / U1 snRNP / tRNA processing / Cajal body / U2-type prespliceosome / K63-linked polyubiquitin modification-dependent protein binding / precatalytic spliceosome / positive regulation of transcription by RNA polymerase I / spliceosomal complex assembly / mRNA Splicing - Minor Pathway / regulation of RNA splicing / negative regulation of mRNA splicing, via spliceosome / mRNA catabolic process / nuclear-transcribed mRNA catabolic process / mRNA 3'-splice site recognition / MLL1 complex / spliceosomal tri-snRNP complex assembly / single fertilization / Major pathway of rRNA processing in the nucleolus and cytosol / U5 snRNA binding / U5 snRNP / U2 snRNA binding / RNA processing / U6 snRNA binding / ribonucleoprotein complex binding / spliceosomal snRNP assembly / regulation of DNA repair / pre-mRNA intronic binding / U1 snRNA binding / U4/U6 x U5 tri-snRNP complex / Gene and protein expression by JAK-STAT signaling after Interleukin-12 stimulation / catalytic step 2 spliceosome Similarity search - Function | |||||||||
Biological species | ![]() | |||||||||
Method | single particle reconstruction / cryo EM / Resolution: 6.1 Å | |||||||||
![]() | Zhang Z / Kumar V / Dybkov O / Will CL / Zhong J / Ludwig S / Urlaub H / Kastner B / Stark H / Luehrmann R | |||||||||
Funding support | ![]()
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![]() | ![]() Title: Structural insights into the cross-exon to cross-intron spliceosome switch. Authors: Zhenwei Zhang / Vinay Kumar / Olexandr Dybkov / Cindy L Will / Jiayun Zhong / Sebastian E J Ludwig / Henning Urlaub / Berthold Kastner / Holger Stark / Reinhard Lührmann / ![]() ![]() Abstract: Early spliceosome assembly can occur through an intron-defined pathway, whereby U1 and U2 small nuclear ribonucleoprotein particles (snRNPs) assemble across the intron. Alternatively, it can occur ...Early spliceosome assembly can occur through an intron-defined pathway, whereby U1 and U2 small nuclear ribonucleoprotein particles (snRNPs) assemble across the intron. Alternatively, it can occur through an exon-defined pathway, whereby U2 binds the branch site located upstream of the defined exon and U1 snRNP interacts with the 5' splice site located directly downstream of it. The U4/U6.U5 tri-snRNP subsequently binds to produce a cross-intron (CI) or cross-exon (CE) pre-B complex, which is then converted to the spliceosomal B complex. Exon definition promotes the splicing of upstream introns and plays a key part in alternative splicing regulation. However, the three-dimensional structure of exon-defined spliceosomal complexes and the molecular mechanism of the conversion from a CE-organized to a CI-organized spliceosome, a pre-requisite for splicing catalysis, remain poorly understood. Here cryo-electron microscopy analyses of human CE pre-B complex and B-like complexes reveal extensive structural similarities with their CI counterparts. The results indicate that the CE and CI spliceosome assembly pathways converge already at the pre-B stage. Add-back experiments using purified CE pre-B complexes, coupled with cryo-electron microscopy, elucidate the order of the extensive remodelling events that accompany the formation of B complexes and B-like complexes. The molecular triggers and roles of B-specific proteins in these rearrangements are also identified. We show that CE pre-B complexes can productively bind in trans to a U1 snRNP-bound 5' splice site. Together, our studies provide new mechanistic insights into the CE to CI switch during spliceosome assembly and its effect on pre-mRNA splice site pairing at this stage. | |||||||||
History |
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Structure visualization
Supplemental images |
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Downloads & links
-EMDB archive
Map data | ![]() | 336.4 MB | ![]() | |
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Header (meta data) | ![]() ![]() | 70.8 KB 70.8 KB | Display Display | ![]() |
Images | ![]() | 73.2 KB | ||
Filedesc metadata | ![]() | 21 KB | ||
Others | ![]() ![]() | 338.1 MB 338 MB | ||
Archive directory | ![]() ![]() | HTTPS FTP |
-Validation report
Summary document | ![]() | 924.2 KB | Display | ![]() |
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Full document | ![]() | 923.8 KB | Display | |
Data in XML | ![]() | 17.8 KB | Display | |
Data in CIF | ![]() | 21.3 KB | Display | |
Arichive directory | ![]() ![]() | HTTPS FTP |
-Related structure data
Related structure data | ![]() 8r08MC ![]() 8qozC ![]() 8qp8C ![]() 8qp9C ![]() 8qpaC ![]() 8qpbC ![]() 8qpeC ![]() 8qpkC ![]() 8qxdC ![]() 8qzsC ![]() 8r09C ![]() 8r0aC ![]() 8r0bC ![]() 8rm5C M: atomic model generated by this map C: citing same article ( |
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Similar structure data | Similarity search - Function & homology ![]() |
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Links
EMDB pages | ![]() ![]() |
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Related items in Molecule of the Month |
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Map
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Voxel size | X=Y=Z: 1.35 Å | ||||||||||||||||||||
Density |
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Symmetry | Space group: 1 | ||||||||||||||||||||
Details | EMDB XML:
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-Supplemental data
-Half map: #2
File | emd_18786_half_map_1.map | ||||||||||||
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Projections & Slices |
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Density Histograms |
-Half map: #1
File | emd_18786_half_map_2.map | ||||||||||||
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Projections & Slices |
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Density Histograms |
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Sample components
+Entire : human spliceosomal pre-B+AMPPNP complex
+Supramolecule #1: human spliceosomal pre-B+AMPPNP complex
+Macromolecule #1: U1 snRNA
+Macromolecule #9: pre-mRNA
+Macromolecule #13: U2 snRNA
+Macromolecule #14: U4 snRNA
+Macromolecule #15: U5 snRNA
+Macromolecule #16: U6 snRNA
+Macromolecule #21: oligo 1
+Macromolecule #2: 116 kDa U5 small nuclear ribonucleoprotein component
+Macromolecule #3: Thioredoxin-like protein 4A
+Macromolecule #4: U5 small nuclear ribonucleoprotein 40 kDa protein
+Macromolecule #5: NHP2-like protein 1, N-terminally processed
+Macromolecule #6: RNA-binding protein 42
+Macromolecule #7: Ubiquitin carboxyl-terminal hydrolase 39
+Macromolecule #8: U4/U6.U5 small nuclear ribonucleoprotein 27 kDa protein
+Macromolecule #10: Serine/threonine-protein kinase PRP4 homolog
+Macromolecule #11: Pre-mRNA-processing-splicing factor 8
+Macromolecule #12: U5 small nuclear ribonucleoprotein 200 kDa helicase
+Macromolecule #17: Splicing factor 3A subunit 1
+Macromolecule #18: Splicing factor 3A subunit 2
+Macromolecule #19: U4/U6 small nuclear ribonucleoprotein Prp3
+Macromolecule #20: U4/U6 small nuclear ribonucleoprotein Prp31
+Macromolecule #22: Small nuclear ribonucleoprotein Sm D3
+Macromolecule #23: Small nuclear ribonucleoprotein F
+Macromolecule #24: Small nuclear ribonucleoprotein Sm D1
+Macromolecule #25: Small nuclear ribonucleoprotein-associated proteins B and B'
+Macromolecule #26: Small nuclear ribonucleoprotein G
+Macromolecule #27: Small nuclear ribonucleoprotein E
+Macromolecule #28: Small nuclear ribonucleoprotein Sm D2
+Macromolecule #29: U1 small nuclear ribonucleoprotein 70 kDa
+Macromolecule #30: U4/U6.U5 tri-snRNP-associated protein 1
+Macromolecule #31: U4/U6 small nuclear ribonucleoprotein Prp4
+Macromolecule #32: Pre-mRNA-processing factor 6
+Macromolecule #33: Probable ATP-dependent RNA helicase DDX23
+Macromolecule #34: Splicing factor 3B subunit 4
+Macromolecule #35: Splicing factor 3A subunit 3
+Macromolecule #36: Splicing factor 3B subunit 2
+Macromolecule #37: Splicing factor 3B subunit 5
+Macromolecule #38: Splicing factor 3B subunit 3
+Macromolecule #39: PHD finger-like domain-containing protein 5A
+Macromolecule #40: Splicing factor 3B subunit 1
+Macromolecule #41: Splicing factor 3B subunit 6
+Macromolecule #42: U2 small nuclear ribonucleoprotein B''
+Macromolecule #43: U2 small nuclear ribonucleoprotein A'
+Macromolecule #44: U6 snRNA-associated Sm-like protein LSm6
+Macromolecule #45: U6 snRNA-associated Sm-like protein LSm7
+Macromolecule #46: U6 snRNA-associated Sm-like protein LSm2
+Macromolecule #47: U6 snRNA-associated Sm-like protein LSm3
+Macromolecule #48: U6 snRNA-associated Sm-like protein LSm8
+Macromolecule #49: U6 snRNA-associated Sm-like protein LSm4
+Macromolecule #50: U6 snRNA-associated Sm-like protein LSm5
-Experimental details
-Structure determination
Method | cryo EM |
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![]() | single particle reconstruction |
Aggregation state | particle |
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Sample preparation
Buffer | pH: 7.9 |
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Vitrification | Cryogen name: ETHANE |
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Electron microscopy
Microscope | FEI TITAN KRIOS |
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Image recording | Film or detector model: FEI FALCON III (4k x 4k) / Average electron dose: 45.0 e/Å2 |
Electron beam | Acceleration voltage: 300 kV / Electron source: ![]() |
Electron optics | Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Nominal defocus max: 5.0 µm / Nominal defocus min: 1.5 µm |
Experimental equipment | ![]() Model: Titan Krios / Image courtesy: FEI Company |
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Image processing
Startup model | Type of model: OTHER / Details: ab initio |
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Final reconstruction | Resolution.type: BY AUTHOR / Resolution: 6.1 Å / Resolution method: FSC 0.143 CUT-OFF / Number images used: 53422 |
Initial angle assignment | Type: MAXIMUM LIKELIHOOD |
Final angle assignment | Type: MAXIMUM LIKELIHOOD |