+Open data
-Basic information
Entry | Database: EMDB / ID: EMD-18545 | |||||||||
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Title | Cryo-EM Structure of Pre-B+AMPPNP Complex (core part) | |||||||||
Map data | ||||||||||
Sample |
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Keywords | spliceosome / SPLICING | |||||||||
Function / homology | Function and homology information ribonucleoprotein complex localization / U4atac snRNP / positive regulation of cytotoxic T cell differentiation / maturation of 5S rRNA / RNA localization / R-loop processing / U4atac snRNA binding / cis assembly of pre-catalytic spliceosome / spliceosome conformational change to release U4 (or U4atac) and U1 (or U11) / snRNP binding ...ribonucleoprotein complex localization / U4atac snRNP / positive regulation of cytotoxic T cell differentiation / maturation of 5S rRNA / RNA localization / R-loop processing / U4atac snRNA binding / cis assembly of pre-catalytic spliceosome / spliceosome conformational change to release U4 (or U4atac) and U1 (or U11) / snRNP binding / U2-type catalytic step 1 spliceosome / RNA splicing, via transesterification reactions / U4 snRNA binding / spliceosomal tri-snRNP complex / U2-type spliceosomal complex / U2-type precatalytic spliceosome / mRNA cis splicing, via spliceosome / U2-type prespliceosome assembly / U2-type catalytic step 2 spliceosome / U4 snRNP / U2 snRNP / U2-type prespliceosome / precatalytic spliceosome / K63-linked polyubiquitin modification-dependent protein binding / mRNA Splicing - Minor Pathway / spliceosomal complex assembly / negative regulation of mRNA splicing, via spliceosome / mRNA 3'-splice site recognition / MLL1 complex / spliceosomal tri-snRNP complex assembly / protein deubiquitination / U5 snRNA binding / U5 snRNP / U2 snRNA binding / U6 snRNA binding / ribonucleoprotein complex binding / spliceosomal snRNP assembly / Cajal body / pre-mRNA intronic binding / U1 snRNA binding / U4/U6 x U5 tri-snRNP complex / catalytic step 2 spliceosome / mRNA Splicing - Major Pathway / RNA splicing / response to cocaine / helicase activity / spliceosomal complex / mRNA splicing, via spliceosome / mRNA processing / osteoblast differentiation / cellular response to xenobiotic stimulus / cellular response to tumor necrosis factor / protein-macromolecule adaptor activity / cellular response to lipopolysaccharide / ubiquitinyl hydrolase 1 / nucleic acid binding / RNA helicase activity / hydrolase activity / RNA helicase / nuclear speck / cell division / intracellular membrane-bounded organelle / GTPase activity / mRNA binding / chromatin / nucleolus / GTP binding / Golgi apparatus / positive regulation of transcription by RNA polymerase II / ATP hydrolysis activity / RNA binding / extracellular exosome / zinc ion binding / nucleoplasm / ATP binding / identical protein binding / membrane / nucleus / cytosol / cytoplasm Similarity search - Function | |||||||||
Biological species | Homo sapiens (human) | |||||||||
Method | single particle reconstruction / cryo EM / Resolution: 4.1 Å | |||||||||
Authors | Zhang Z / Kumar V / Dybkov O / Will CL / Zhong J / Ludwig S / Urlaub H / Kastner B / Stark H / Luehrmann R | |||||||||
Funding support | Germany, 1 items
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Citation | Journal: Nature / Year: 2024 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 | emd_18545.map.gz | 387.7 MB | EMDB map data format | |
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Header (meta data) | emd-18545-v30.xml emd-18545.xml | 37.5 KB 37.5 KB | Display Display | EMDB header |
FSC (resolution estimation) | emd_18545_fsc.xml | 16.9 KB | Display | FSC data file |
Images | emd_18545.png | 41.1 KB | ||
Filedesc metadata | emd-18545.cif.gz | 13.6 KB | ||
Others | emd_18545_half_map_1.map.gz emd_18545_half_map_2.map.gz | 337.1 MB 337 MB | ||
Archive directory | http://ftp.pdbj.org/pub/emdb/structures/EMD-18545 ftp://ftp.pdbj.org/pub/emdb/structures/EMD-18545 | HTTPS FTP |
-Validation report
Summary document | emd_18545_validation.pdf.gz | 930.2 KB | Display | EMDB validaton report |
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Full document | emd_18545_full_validation.pdf.gz | 929.7 KB | Display | |
Data in XML | emd_18545_validation.xml.gz | 24.5 KB | Display | |
Data in CIF | emd_18545_validation.cif.gz | 32.7 KB | Display | |
Arichive directory | https://ftp.pdbj.org/pub/emdb/validation_reports/EMD-18545 ftp://ftp.pdbj.org/pub/emdb/validation_reports/EMD-18545 | HTTPS FTP |
-Related structure data
Related structure data | 8qp9MC 8qozC 8qp8C 8qpaC 8qpbC 8qpeC 8qpkC 8qxdC 8qzsC 8r08C 8r09C 8r0aC 8r0bC 8rm5C M: atomic model generated by this map C: citing same article (ref.) |
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Similar structure data | Similarity search - Function & homologyF&H Search |
-Links
EMDB pages | EMDB (EBI/PDBe) / EMDataResource |
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Related items in Molecule of the Month |
-Map
File | Download / File: emd_18545.map.gz / Format: CCP4 / Size: 421.9 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES) | ||||||||||||||||||||||||||||||||||||
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Projections & slices | Image control
Images are generated by Spider. | ||||||||||||||||||||||||||||||||||||
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_18545_half_map_1.map | ||||||||||||
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Projections & Slices |
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Density Histograms |
-Half map: #1
File | emd_18545_half_map_2.map | ||||||||||||
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Projections & Slices |
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Density Histograms |
-Sample components
+Entire : human spliceosomal pre-B+AMPPNP complex
+Supramolecule #1: human spliceosomal pre-B+AMPPNP complex
+Macromolecule #1: Splicing factor 3A subunit 1
+Macromolecule #2: U5 small nuclear ribonucleoprotein 200 kDa helicase
+Macromolecule #3: 116 kDa U5 small nuclear ribonucleoprotein component
+Macromolecule #4: Thioredoxin-like protein 4A
+Macromolecule #5: U4/U6 small nuclear ribonucleoprotein Prp3
+Macromolecule #6: U4/U6 small nuclear ribonucleoprotein Prp31
+Macromolecule #7: RNA-binding protein 42
+Macromolecule #8: U4/U6.U5 tri-snRNP-associated protein 1
+Macromolecule #9: Ubiquitin carboxyl-terminal hydrolase 39
+Macromolecule #10: U4/U6.U5 small nuclear ribonucleoprotein 27 kDa protein
+Macromolecule #11: Probable ATP-dependent RNA helicase DDX23
+Macromolecule #15: Pre-mRNA-processing-splicing factor 8
+Macromolecule #16: Pre-mRNA-processing factor 6
+Macromolecule #12: U6 snRNA
+Macromolecule #13: U5 snRNA
+Macromolecule #14: U4 snRNA
+Macromolecule #17: INOSITOL HEXAKISPHOSPHATE
-Experimental details
-Structure determination
Method | cryo EM |
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Processing | single particle reconstruction |
Aggregation state | particle |
-Sample preparation
Buffer | pH: 7.9 |
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Vitrification | Cryogen name: ETHANE |
-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: FIELD EMISSION GUN |
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 |