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- EMDB-47157: Yeast post-catalytic P complex spliceosome, state I, overall map -
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Open data
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Basic information
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Title | Yeast post-catalytic P complex spliceosome, state I, overall map | |||||||||
![]() | P-complex spliceosome state I core refinement map, postprocessed | |||||||||
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![]() | spliceosome / snRNA / yeast / ribonucleoprotein complex / SPLICING | |||||||||
Function / homology | ![]() U2-type post-spliceosomal complex / mRNA branch site recognition / spliceosomal complex disassembly / post-spliceosomal complex / U2-type post-mRNA release spliceosomal complex / cellular bud site selection / pre-mRNA 3'-splice site binding / post-mRNA release spliceosomal complex / nuclear mRNA surveillance / generation of catalytic spliceosome for first transesterification step ...U2-type post-spliceosomal complex / mRNA branch site recognition / spliceosomal complex disassembly / post-spliceosomal complex / U2-type post-mRNA release spliceosomal complex / cellular bud site selection / pre-mRNA 3'-splice site binding / post-mRNA release spliceosomal complex / nuclear mRNA surveillance / generation of catalytic spliceosome for first transesterification step / cis assembly of pre-catalytic spliceosome / spliceosome conformational change to release U4 (or U4atac) and U1 (or U11) / U4/U6 snRNP / 7-methylguanosine cap hypermethylation / pICln-Sm protein complex / U2-type catalytic step 1 spliceosome / pre-mRNA binding / snRNP binding / small nuclear ribonucleoprotein complex / splicing factor binding / SMN-Sm protein complex / spliceosomal tri-snRNP complex / U2-type spliceosomal complex / commitment complex / mRNA cis splicing, via spliceosome / U2-type prespliceosome assembly / U2-type catalytic step 2 spliceosome / U4 snRNP / U2 snRNP / U1 snRNP / poly(U) RNA binding / U2-type prespliceosome / precatalytic spliceosome / generation of catalytic spliceosome for second transesterification step / Formation of TC-NER Pre-Incision Complex / spliceosomal complex assembly / mRNA 5'-splice site recognition / Gap-filling DNA repair synthesis and ligation in TC-NER / mRNA 3'-splice site recognition / DNA replication origin binding / spliceosomal tri-snRNP complex assembly / Dual incision in TC-NER / Prp19 complex / protein K63-linked ubiquitination / U5 snRNA binding / U5 snRNP / DNA replication initiation / U2 snRNA binding / U6 snRNA binding / pre-mRNA intronic binding / spliceosomal snRNP assembly / U1 snRNA binding / U4/U6 x U5 tri-snRNP complex / positive regulation of cell cycle / catalytic step 2 spliceosome / nuclear periphery / positive regulation of RNA splicing / spliceosomal complex / mRNA splicing, via spliceosome / RING-type E3 ubiquitin transferase / double-strand break repair via nonhomologous end joining / metallopeptidase activity / ubiquitin-protein transferase activity / ubiquitin protein ligase activity / chromosome, telomeric region / RNA helicase activity / RNA helicase / DNA repair / mRNA binding / GTPase activity / chromatin binding / chromatin / GTP binding / ATP hydrolysis activity / mitochondrion / DNA binding / RNA binding / zinc ion binding / ATP binding / identical protein binding / nucleus / cytosol / cytoplasm Similarity search - Function | |||||||||
Biological species | ![]() ![]() | |||||||||
Method | single particle reconstruction / cryo EM / Resolution: 2.31 Å | |||||||||
![]() | Wilkinson ME / Hoskins AA | |||||||||
Funding support | ![]()
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![]() | ![]() Title: Control of 3' splice site selection by the yeast splicing factor Fyv6. Authors: Katherine A Senn / Karli A Lipinski / Natalie J Zeps / Amory F Griffin / Max E Wilkinson / Aaron A Hoskins / ![]() ![]() Abstract: Pre-mRNA splicing is catalyzed in two steps: 5' splice site (SS) cleavage and exon ligation. A number of proteins transiently associate with spliceosomes to specifically impact these steps (1 and 2 ...Pre-mRNA splicing is catalyzed in two steps: 5' splice site (SS) cleavage and exon ligation. A number of proteins transiently associate with spliceosomes to specifically impact these steps (1 and 2 step factors). We recently identified Fyv6 (FAM192A in humans) as a 2 step factor in ; however, we did not determine how widespread Fyv6's impact is on the transcriptome. To answer this question, we have used RNA-seq to analyze changes in splicing. These results show that loss of Fyv6 results in activation of non-consensus, branch point (BP) proximal 3' SS transcriptome-wide. To identify the molecular basis of these observations, we determined a high-resolution cryo-EM structure of a yeast product complex spliceosome containing Fyv6 at 2.3 Å. The structure reveals that Fyv6 is the only 2 step factor that contacts the Prp22 ATPase and that Fyv6 binding is mutually exclusive with that of the 1 step factor Yju2. We then use this structure to dissect Fyv6 functional domains and interpret results of a genetic screen for suppressor mutations. The combined transcriptomic, structural, and genetic studies allow us to propose a model in which Yju2/Fyv6 exchange facilitates exon ligation and Fyv6 promotes usage of consensus, BP distal 3' SS. | |||||||||
History |
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Structure visualization
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Downloads & links
-EMDB archive
Map data | ![]() | 387.5 MB | ![]() | |
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Header (meta data) | ![]() ![]() | 65.4 KB 65.4 KB | Display Display | ![]() |
FSC (resolution estimation) | ![]() | 16.8 KB | Display | ![]() |
Images | ![]() | 144.1 KB | ||
Masks | ![]() | 421.9 MB | ![]() | |
Filedesc metadata | ![]() | 18 KB | ||
Others | ![]() ![]() | 339.3 MB 339.3 MB | ||
Archive directory | ![]() ![]() | HTTPS FTP |
-Validation report
Summary document | ![]() | 1.1 MB | Display | ![]() |
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Full document | ![]() | 1.1 MB | Display | |
Data in XML | ![]() | 24.6 KB | Display | |
Data in CIF | ![]() | 33.1 KB | Display | |
Arichive directory | ![]() ![]() | HTTPS FTP |
-Related structure data
Related structure data | ![]() 9dtrMC 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
File | ![]() | ||||||||||||||||||||
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Annotation | P-complex spliceosome state I core refinement map, postprocessed | ||||||||||||||||||||
Voxel size | X=Y=Z: 0.968 Å | ||||||||||||||||||||
Density |
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Symmetry | Space group: 1 | ||||||||||||||||||||
Details | EMDB XML:
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-Supplemental data
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Sample components
+Entire : Yeast post-catalytic P complex spliceosome
+Supramolecule #1: Yeast post-catalytic P complex spliceosome
+Macromolecule #1: U2 snRNA
+Macromolecule #2: U5 snRNA
+Macromolecule #3: U6 snRNA
+Macromolecule #7: UBC4 mRNA spliced exons
+Macromolecule #10: UBC4 lariat-intron
+Macromolecule #4: Pre-mRNA-splicing factor 8
+Macromolecule #5: Pre-mRNA-splicing factor SNU114
+Macromolecule #6: Protein FYV6
+Macromolecule #8: Pre-mRNA-splicing factor ISY1
+Macromolecule #9: Pre-mRNA-splicing factor CWC22
+Macromolecule #11: Pre-mRNA-splicing factor PRP46
+Macromolecule #12: Pre-mRNA-processing protein 45
+Macromolecule #13: Pre-mRNA-splicing factor BUD31
+Macromolecule #14: Pre-mRNA-splicing factor CWC2
+Macromolecule #15: Pre-mRNA-splicing factor SLT11
+Macromolecule #16: Pre-mRNA-splicing factor CEF1
+Macromolecule #17: Pre-mRNA-splicing factor CWC15
+Macromolecule #18: Pre-mRNA-splicing factor CWC21
+Macromolecule #19: Pre-mRNA-splicing factor CLF1
+Macromolecule #20: Pre-mRNA-splicing factor SYF1
+Macromolecule #21: Pre-mRNA-splicing factor ATP-dependent RNA helicase PRP22
+Macromolecule #22: U2 small nuclear ribonucleoprotein A'
+Macromolecule #23: U2 small nuclear ribonucleoprotein B''
+Macromolecule #24: Pre-mRNA-splicing factor NTC20
+Macromolecule #25: Pre-mRNA-splicing factor 18
+Macromolecule #26: Small nuclear ribonucleoprotein-associated protein B
+Macromolecule #27: Pre-mRNA-splicing factor SLU7
+Macromolecule #28: Small nuclear ribonucleoprotein Sm D3
+Macromolecule #29: Small nuclear ribonucleoprotein E
+Macromolecule #30: Small nuclear ribonucleoprotein F
+Macromolecule #31: Small nuclear ribonucleoprotein G
+Macromolecule #32: Small nuclear ribonucleoprotein Sm D1
+Macromolecule #33: Small nuclear ribonucleoprotein Sm D2
+Macromolecule #34: Pre-mRNA-processing factor 17
+Macromolecule #35: Pre-mRNA-splicing factor SNT309
+Macromolecule #36: Pre-mRNA-processing factor 19
+Macromolecule #37: Pre-mRNA-splicing factor SYF2
+Macromolecule #38: MAGNESIUM ION
+Macromolecule #39: POTASSIUM ION
+Macromolecule #40: INOSITOL HEXAKISPHOSPHATE
+Macromolecule #41: GUANOSINE-5'-TRIPHOSPHATE
+Macromolecule #42: ZINC ION
+Macromolecule #43: water
-Experimental details
-Structure determination
Method | cryo EM |
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![]() | single particle reconstruction |
Aggregation state | particle |
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Sample preparation
Concentration | 0.1 mg/mL | ||||||||
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Buffer | pH: 7.9 Component:
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Grid | Model: Quantifoil R2/2 / Material: COPPER / Mesh: 300 / Support film - Material: CARBON / Support film - topology: CONTINUOUS / Support film - Film thickness: 2 / Pretreatment - Type: GLOW DISCHARGE / Pretreatment - Time: 12 sec. / Pretreatment - Atmosphere: AIR | ||||||||
Vitrification | Cryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 277 K / Instrument: FEI VITROBOT MARK IV |
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Electron microscopy
Microscope | TFS KRIOS |
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Image recording | Film or detector model: GATAN K3 BIOQUANTUM (6k x 4k) / Number grids imaged: 2 / Number real images: 51113 / Average exposure time: 0.72 sec. / Average electron dose: 40.0 e/Å2 |
Electron beam | Acceleration voltage: 300 kV / Electron source: ![]() |
Electron optics | C2 aperture diameter: 50.0 µm / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Cs: 2.7 mm / Nominal defocus max: 3.1 µm / Nominal defocus min: 1.3 µm / Nominal magnification: 130000 |
Sample stage | Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN |
Experimental equipment | ![]() Model: Titan Krios / Image courtesy: FEI Company |
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Image processing
-Atomic model buiding 1
Initial model | PDB ID: Chain - Source name: PDB / Chain - Initial model type: experimental model |
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Details | After fixing in ISOLDE, the model was refined in phenix.real_space_refine with the following non-default flags: refinement { run = *minimization_global rigid_body local_grid_search morphing simulated_annealing *adp } reference_model { enabled = True use_starting_model_as_reference = True fix_outliers = False strict_rotamer_matching = True } pdb_interpretation { reference_coordinate_restraints { enabled = True exclude_outliers = False top_out = True } automatic_linking { link_metals = True } peptide_link { restrain_rama_outliers = False restrain_rama_allowed = False } ramachandran_plot_restraints { enabled = False } } |
Refinement | Space: REAL / Protocol: FLEXIBLE FIT |
Output model | ![]() PDB-9dtr: |