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- EMDB-4795: Yeast pre-40S Rps20 DeltaLoop C2-Headonly -

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Entry
Database: EMDB / ID: EMD-4795
TitleYeast pre-40S Rps20 DeltaLoop C2-Headonly
Map dataC2-headonly S20Delta loop yeast pre-40S particle Structural state 2 3D reconstruction of the head only(signal subtraction of the body) Local Sharpening with LocScale
Sample
  • Complex: C2-S20DeltaLoop State 2 of yeast pre-40S particles purified with Tsr1-TAP as bait, in mutant conditions (deletion of aa 68-78 of Rps20)
Function / homology
Function and homology information


positive regulation of RNA import into nucleus / 18S rRNA (adenine1779-N6/adenine1780-N6)-dimethyltransferase / 18S rRNA (adenine(1779)-N(6)/adenine(1780)-N(6))-dimethyltransferase activity / rRNA modification / rRNA (adenine-N6,N6-)-dimethyltransferase activity / endonucleolytic cleavage of tricistronic rRNA transcript (SSU-rRNA, 5.8S rRNA, LSU-rRNA) / maturation of SSU-rRNA from tricistronic rRNA transcript (SSU-rRNA, LSU-rRNA,5S) / endonucleolytic cleavage to generate mature 5'-end of SSU-rRNA from (SSU-rRNA, 5.8S rRNA, LSU-rRNA) / negative regulation of glucose mediated signaling pathway / negative regulation of translational frameshifting ...positive regulation of RNA import into nucleus / 18S rRNA (adenine1779-N6/adenine1780-N6)-dimethyltransferase / 18S rRNA (adenine(1779)-N(6)/adenine(1780)-N(6))-dimethyltransferase activity / rRNA modification / rRNA (adenine-N6,N6-)-dimethyltransferase activity / endonucleolytic cleavage of tricistronic rRNA transcript (SSU-rRNA, 5.8S rRNA, LSU-rRNA) / maturation of SSU-rRNA from tricistronic rRNA transcript (SSU-rRNA, LSU-rRNA,5S) / endonucleolytic cleavage to generate mature 5'-end of SSU-rRNA from (SSU-rRNA, 5.8S rRNA, LSU-rRNA) / negative regulation of glucose mediated signaling pathway / negative regulation of translational frameshifting / Negative regulators of DDX58/IFIH1 signaling / positive regulation of translational fidelity / Protein methylation / RMTs methylate histone arginines / mTORC1-mediated signalling / Protein hydroxylation / rRNA methylation / ribosome-associated ubiquitin-dependent protein catabolic process / GDP-dissociation inhibitor activity / positive regulation of nuclear-transcribed mRNA catabolic process, deadenylation-dependent decay / U3 snoRNA binding / Formation of the ternary complex, and subsequently, the 43S complex / Translation initiation complex formation / Ribosomal scanning and start codon recognition / poly(A)+ mRNA export from nucleus / preribosome, small subunit precursor / snoRNA binding / mRNA destabilization / Major pathway of rRNA processing in the nucleolus and cytosol / SRP-dependent cotranslational protein targeting to membrane / GTP hydrolysis and joining of the 60S ribosomal subunit / Formation of a pool of free 40S subunits / Nonsense Mediated Decay (NMD) independent of the Exon Junction Complex (EJC) / Nonsense Mediated Decay (NMD) enhanced by the Exon Junction Complex (EJC) / L13a-mediated translational silencing of Ceruloplasmin expression / endonucleolytic cleavage to generate mature 3'-end of SSU-rRNA from (SSU-rRNA, 5.8S rRNA, LSU-rRNA) / proteasome assembly / G-protein alpha-subunit binding / positive regulation of protein kinase activity / regulation of translational fidelity / Ub-specific processing proteases / ribosomal small subunit export from nucleus / translation regulator activity / ribonucleoprotein complex binding / ribosomal subunit export from nucleus / endonucleolytic cleavage in ITS1 to separate SSU-rRNA from 5.8S rRNA and LSU-rRNA from tricistronic rRNA transcript (SSU-rRNA, 5.8S rRNA, LSU-rRNA) / DNA-(apurinic or apyrimidinic site) endonuclease activity / cellular response to amino acid starvation / rescue of stalled ribosome / 90S preribosome / maturation of SSU-rRNA from tricistronic rRNA transcript (SSU-rRNA, 5.8S rRNA, LSU-rRNA) / maturation of SSU-rRNA / small-subunit processome / positive regulation of apoptotic signaling pathway / protein kinase C binding / maintenance of translational fidelity / cytoplasmic stress granule / modification-dependent protein catabolic process / rRNA processing / protein tag activity / unfolded protein binding / ribosome biogenesis / ribosome binding / ribosomal small subunit biogenesis / ribosomal small subunit assembly / small ribosomal subunit / small ribosomal subunit rRNA binding / cytosolic small ribosomal subunit / cytoplasmic translation / rRNA binding / negative regulation of translation / non-specific serine/threonine protein kinase / ribosome / protein kinase activity / protein ubiquitination / structural constituent of ribosome / translation / positive regulation of protein phosphorylation / G protein-coupled receptor signaling pathway / phosphorylation / negative regulation of gene expression / protein serine kinase activity / protein serine/threonine kinase activity / GTPase activity / mRNA binding / ubiquitin protein ligase binding / nucleolus / GTP binding / mitochondrion / RNA binding / zinc ion binding / nucleoplasm / ATP binding / nucleus / metal ion binding / cytoplasm / cytosol
Similarity search - Function
Serine/threonine-protein kinase Rio2 / RIO2 kinase winged helix domain, N-terminal / Rio2, N-terminal / RIO kinase / RIO-like kinase / RIO1 family / Ribosomal RNA adenine dimethylase / Krr1, KH1 domain / Krr1 KH1 domain / Bystin ...Serine/threonine-protein kinase Rio2 / RIO2 kinase winged helix domain, N-terminal / Rio2, N-terminal / RIO kinase / RIO-like kinase / RIO1 family / Ribosomal RNA adenine dimethylase / Krr1, KH1 domain / Krr1 KH1 domain / Bystin / Bystin / Ribosomal RNA adenine methylase transferase, conserved site / Ribosomal RNA adenine methylase transferase, N-terminal / Ribosomal RNA adenine dimethylases signature. / Ribosomal RNA adenine dimethylases / Ribosomal RNA adenine methyltransferase KsgA/Erm / Ribosomal RNA adenine dimethylase / rRNA adenine N(6)-methyltransferase family profile. / Ribosome biogenesis protein BMS1/TSR1, C-terminal / AARP2CN / Bms1/Tsr1-type G domain / Ribosome biogenesis protein Bms1/Tsr1 / 40S ribosome biogenesis protein Tsr1 and BMS1 C-terminal / AARP2CN (NUC121) domain / Bms1-type guanine nucleotide-binding (G) domain profile. / AARP2CN (NUC121) domain / Protein of unknown function (DUF663) / K Homology domain, type 1 superfamily / : / Ribosomal protein S21e, conserved site / Ribosomal protein S21e signature. / Ribosomal protein S12e signature. / Ribosomal protein S12e / Ribosomal protein S5, eukaryotic/archaeal / Ribosomal protein S19e, conserved site / Ribosomal protein S19e signature. / Small (40S) ribosomal subunit Asc1/RACK1 / Ribosomal protein S2, eukaryotic / Ribosomal protein S21e / Ribosomal protein S21e superfamily / Ribosomal protein S21e / 40S Ribosomal protein S10 / S27a-like superfamily / Ribosomal protein S10, eukaryotic/archaeal / Plectin/S10, N-terminal / Plectin/S10 domain / Ribosomal protein S25 / S25 ribosomal protein / Ribosomal protein S2, eukaryotic/archaeal / : / Ribosomal protein S17e, conserved site / Ribosomal protein S17e signature. / Ribosomal protein S27a / Ribosomal protein S27a / Ribosomal protein S27a / Ribosomal protein S8e subdomain, eukaryotes / Ribosomal protein S30 / Ribosomal protein S30 / 40S ribosomal protein S29/30S ribosomal protein S14 type Z / Ribosomal protein S7e signature. / Ribosomal protein S3, eukaryotic/archaeal / Ribosomal protein S19e / Ribosomal protein S3Ae, conserved site / Ribosomal protein S19e / Ribosomal protein S3Ae signature. / Ribosomal_S19e / Ribosomal protein S27e signature. / Ribosomal protein S4e, N-terminal, conserved site / Ribosomal protein S4e signature. / 40S ribosomal protein S4, C-terminal domain / 40S ribosomal protein S4 C-terminus / Ribosomal protein S19A/S15e / Ribosomal protein S8e, conserved site / Ribosomal protein S8e signature. / Ribosomal protein S17e / Ribosomal protein S17e-like superfamily / Ribosomal S17 / Ribosomal protein S6, eukaryotic / 40S ribosomal protein S1/3, eukaryotes / Ribosomal protein S4e, N-terminal / RS4NT (NUC023) domain / Ribosomal S24e conserved site / Ribosomal protein S24e signature. / 40S ribosomal protein S11, N-terminal / Ribosomal_S17 N-terminal / Ribosomal protein S7e / Ribosomal protein S7e / Ribosomal protein S4, KOW domain / Ribosomal protein S4e / Ribosomal protein S4e, central region / Ribosomal protein S4e, central domain superfamily / Ribosomal family S4e / Ribosomal protein S27, zinc-binding domain superfamily / Ribosomal protein S24e / Ribosomal protein S23, eukaryotic/archaeal / Ribosomal protein S24e / Ribosomal protein S6/S6e/A/B/2, conserved site / Ribosomal protein S6e signature. / Ribosomal protein S27 / Ribosomal protein S27
Similarity search - Domain/homology
Small ribosomal subunit protein uS4A / Small ribosomal subunit protein eS17A / Small ribosomal subunit protein uS3 / Small ribosomal subunit protein uS15 / Ubiquitin-ribosomal protein eS31 fusion protein / Small ribosomal subunit protein uS11A / Small ribosomal subunit protein eS19A / Small ribosomal subunit protein eS21A / Small ribosomal subunit protein uS8A / Small ribosomal subunit protein uS12A ...Small ribosomal subunit protein uS4A / Small ribosomal subunit protein eS17A / Small ribosomal subunit protein uS3 / Small ribosomal subunit protein uS15 / Ubiquitin-ribosomal protein eS31 fusion protein / Small ribosomal subunit protein uS11A / Small ribosomal subunit protein eS19A / Small ribosomal subunit protein eS21A / Small ribosomal subunit protein uS8A / Small ribosomal subunit protein uS12A / Small ribosomal subunit protein eS24A / Small ribosomal subunit protein eS30A / Small ribosomal subunit protein eS4A / Small ribosomal subunit protein eS6A / Small ribosomal subunit protein eS8A / Small ribosomal subunit protein uS17A / Small ribosomal subunit protein uS9A / Small ribosomal subunit protein uS13A / Small ribosomal subunit protein uS5 / Small ribosomal subunit protein uS7 / Small ribosomal subunit protein eS7A / Small ribosomal subunit protein uS2A / Small ribosomal subunit protein eS1A / Small ribosomal subunit protein eS27A / Small ribosomal subunit protein RACK1 / Essential nuclear protein 1 / Small ribosomal subunit protein uS10 / Serine/threonine-protein kinase RIO2 / Small ribosomal subunit protein uS14A / Dimethyladenosine transferase / Small ribosomal subunit protein eS12 / Small ribosomal subunit protein uS19 / Ribosome biogenesis protein TSR1 / Small ribosomal subunit protein eS10A / Small ribosomal subunit protein eS25A / Small ribosomal subunit protein eS28A / Pre-rRNA-processing protein PNO1
Similarity search - Component
Biological speciesSaccharomyces cerevisiae S288c (yeast)
Methodsingle particle reconstruction / cryo EM / Resolution: 3.8 Å
AuthorsShayan R / Mitterer V / Ferreira-Cerca S / Murat G / Rinaldi D / Enne T / Weigl S / Omanic H / Gleizes PE / Kressler D ...Shayan R / Mitterer V / Ferreira-Cerca S / Murat G / Rinaldi D / Enne T / Weigl S / Omanic H / Gleizes PE / Kressler D / Pertschy B / Plisson-Chastang C
Funding support France, 1 items
OrganizationGrant numberCountry
French National Research AgencyRIBOMAN France
CitationJournal: Nat Commun / Year: 2019
Title: Conformational proofreading of distant 40S ribosomal subunit maturation events by a long-range communication mechanism.
Authors: Valentin Mitterer / Ramtin Shayan / Sébastien Ferreira-Cerca / Guillaume Murat / Tanja Enne / Dana Rinaldi / Sarah Weigl / Hajrija Omanic / Pierre-Emmanuel Gleizes / Dieter Kressler / Celia ...Authors: Valentin Mitterer / Ramtin Shayan / Sébastien Ferreira-Cerca / Guillaume Murat / Tanja Enne / Dana Rinaldi / Sarah Weigl / Hajrija Omanic / Pierre-Emmanuel Gleizes / Dieter Kressler / Celia Plisson-Chastang / Brigitte Pertschy /
Abstract: Eukaryotic ribosomes are synthesized in a hierarchical process driven by a plethora of assembly factors, but how maturation events at physically distant sites on pre-ribosomes are coordinated is ...Eukaryotic ribosomes are synthesized in a hierarchical process driven by a plethora of assembly factors, but how maturation events at physically distant sites on pre-ribosomes are coordinated is poorly understood. Using functional analyses and cryo-EM, we show that ribosomal protein Rps20 orchestrates communication between two multi-step maturation events across the pre-40S subunit. Our study reveals that during pre-40S maturation, formation of essential contacts between Rps20 and Rps3 permits assembly factor Ltv1 to recruit the Hrr25 kinase, thereby promoting Ltv1 phosphorylation. In parallel, a deeply buried Rps20 loop reaches to the opposite pre-40S side, where it stimulates Rio2 ATPase activity. Both cascades converge to the final maturation steps releasing Rio2 and phosphorylated Ltv1. We propose that conformational proofreading exerted via Rps20 constitutes a checkpoint permitting assembly factor release and progression of pre-40S maturation only after completion of all earlier maturation steps.
History
DepositionApr 10, 2019-
Header (metadata) releaseJun 26, 2019-
Map releaseJun 26, 2019-
UpdateNov 25, 2020-
Current statusNov 25, 2020Processing site: PDBe / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 0.11
  • Imaged by UCSF Chimera
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  • Surface view colored by height
  • Surface level: 0.11
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  • Surface view with fitted model
  • Atomic models: PDB-6rbe
  • Surface level: 0.11
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Structure viewerEM map:
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Supplemental images

emd_4795.png

Downloads & links

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Map

FileDownload / File: emd_4795.map.gz / Format: CCP4 / Size: 216 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationC2-headonly S20Delta loop yeast pre-40S particle Structural state 2 3D reconstruction of the head only(signal subtraction of the body) Local Sharpening with LocScale
Voxel sizeX=Y=Z: 1.067 Å
Density
Contour LevelBy AUTHOR: 0.11 / Movie #1: 0.11
Minimum - Maximum-0.2522821 - 0.6893848
Average (Standard dev.)0.0012163665 (±0.013330666)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions384384384
Spacing384384384
CellA=B=C: 409.72803 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z1.0671.0671.067
M x/y/z384384384
origin x/y/z0.0000.0000.000
length x/y/z409.728409.728409.728
α/β/γ90.00090.00090.000
start NX/NY/NZ000
NX/NY/NZ320320320
MAP C/R/S123
start NC/NR/NS000
NC/NR/NS384384384
D min/max/mean-0.2520.6890.001

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Supplemental data

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Half map: C2-headonly S20Delta loop yeast pre-40S particle Structural state...

Fileemd_4795_half_map_1.map
AnnotationC2-headonly S20Delta loop yeast pre-40S particle Structural state 2 3D reconstruction of the head only(signal subtraction of the body) unfiltered half map 2
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: C2-headonly S20Delta loop yeast pre-40S particle Structural state...

Fileemd_4795_half_map_2.map
AnnotationC2-headonly S20Delta loop yeast pre-40S particle Structural state 2 3D reconstruction of the head only(signal subtraction of the body) unfiltered half map 1
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Sample components

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Entire : C2-S20DeltaLoop State 2 of yeast pre-40S particles purified with ...

EntireName: C2-S20DeltaLoop State 2 of yeast pre-40S particles purified with Tsr1-TAP as bait, in mutant conditions (deletion of aa 68-78 of Rps20)
Components
  • Complex: C2-S20DeltaLoop State 2 of yeast pre-40S particles purified with Tsr1-TAP as bait, in mutant conditions (deletion of aa 68-78 of Rps20)

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Supramolecule #1: C2-S20DeltaLoop State 2 of yeast pre-40S particles purified with ...

SupramoleculeName: C2-S20DeltaLoop State 2 of yeast pre-40S particles purified with Tsr1-TAP as bait, in mutant conditions (deletion of aa 68-78 of Rps20)
type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1-#35
Source (natural)Organism: Saccharomyces cerevisiae S288c (yeast)
Molecular weightTheoretical: 1.2 MDa

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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
GridSupport film - Material: CARBON / Support film - topology: CONTINUOUS / Support film - Film thickness: 2.0 nm / Pretreatment - Type: GLOW DISCHARGE / Pretreatment - Atmosphere: AIR / Pretreatment - Pressure: 0.045 kPa / Details: 25 mA
VitrificationCryogen name: ETHANE / Chamber humidity: 95 % / Chamber temperature: 293 K / Instrument: LEICA EM GP / Details: blot 1.7-1.9 seconds before plunging.

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Image recordingFilm or detector model: GATAN K2 SUMMIT (4k x 4k) / Detector mode: COUNTING / Digitization - Frames/image: 1-25 / Number grids imaged: 1 / Number real images: 6480 / Average electron dose: 32.4 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Cs: 2.7 mm / Nominal defocus max: 3.0 µm / Nominal defocus min: 0.8 µm
Sample stageSpecimen 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

Particle selectionNumber selected: 645109
CTF correctionSoftware - Name: RELION (ver. 2.1)
Startup modelType of model: PDB ENTRY
PDB model - PDB ID:

4v88

Final reconstructionNumber classes used: 2 / Applied symmetry - Point group: C1 (asymmetric) / Algorithm: EXACT BACK PROJECTION / Resolution.type: BY AUTHOR / Resolution: 3.8 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: RELION (ver. 2.1) / Number images used: 42901
Initial angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: RELION (ver. 2.1)
Final angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: RELION (ver. 2.1)
Final 3D classificationSoftware - Name: RELION (ver. 2.1)
FSC plot (resolution estimation)

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Atomic model buiding 1

Initial modelPDB ID:

4v88

Output model

PDB-6rbd:
State 1 of yeast Tsr1-TAP Rps20-Deltaloop pre-40S particles

PDB-6rbe:
State 2 of yeast Tsr1-TAP Rps20-Deltaloop pre-40S particles

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