[English] 日本語
Yorodumi
- EMDB-23061: Cryo-EM structure of Ephydatia fluviatilis PiwiA-piRNA complex -

+
Open data


ID or keywords:

Loading...

-
Basic information

Entry
Database: EMDB / ID: EMD-23061
TitleCryo-EM structure of Ephydatia fluviatilis PiwiA-piRNA complex
Map dataEf PIWI-piRNA local resolution sharpened map
Sample
  • Complex: Complex between EfPiwi and piRNA
    • Complex: Piwi-A
      • Protein or peptide: Piwi-A
    • Complex: piRNA
      • RNA: RNA (5'-R(P*UP*CP*UP*CP*AP*GP*(OMC))-3')
  • Ligand: MAGNESIUM ION
Function / homology
Function and homology information


Piwi domain / Piwi domain profile. / Piwi domain / Piwi / PAZ domain superfamily / PAZ / PAZ domain / PAZ domain / PAZ domain profile. / Ribonuclease H superfamily / Ribonuclease H-like superfamily
Similarity search - Domain/homology
Biological speciesEphydatia fluviatilis (invertebrata) / synthetic construct (others)
Methodsingle particle reconstruction / cryo EM / Resolution: 3.8 Å
AuthorsChowdhury S / Anzelon TA / MacRae IJ / Lander GC
Funding support United States, 1 items
OrganizationGrant numberCountry
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS) United States
CitationJournal: Nature / Year: 2021
Title: Structural basis for piRNA targeting.
Authors: Todd A Anzelon / Saikat Chowdhury / Siobhan M Hughes / Yao Xiao / Gabriel C Lander / Ian J MacRae /
Abstract: PIWI proteins use PIWI-interacting RNAs (piRNAs) to identify and silence transposable elements and thereby maintain genome integrity between metazoan generations. The targeting of transposable ...PIWI proteins use PIWI-interacting RNAs (piRNAs) to identify and silence transposable elements and thereby maintain genome integrity between metazoan generations. The targeting of transposable elements by PIWI has been compared to mRNA target recognition by Argonaute proteins, which use microRNA (miRNA) guides, but the extent to which piRNAs resemble miRNAs is not known. Here we present cryo-electron microscopy structures of a PIWI-piRNA complex from the sponge Ephydatia fluviatilis with and without target RNAs, and a biochemical analysis of target recognition. Mirroring Argonaute, PIWI identifies targets using the piRNA seed region. However, PIWI creates a much weaker seed so that stable target association requires further piRNA-target pairing, making piRNAs less promiscuous than miRNAs. Beyond the seed, the structure of PIWI facilitates piRNA-target pairing in a manner that is tolerant of mismatches, leading to long-lived PIWI-piRNA-target interactions that may accumulate on transposable-element transcripts. PIWI ensures targeting fidelity by physically blocking the propagation of piRNA-target interactions in the absence of faithful seed pairing, and by requiring an extended piRNA-target duplex to reach an endonucleolytically active conformation. PIWI proteins thereby minimize off-targeting cellular mRNAs while defending against evolving genomic threats.
History
DepositionDec 3, 2020-
Header (metadata) releaseJul 14, 2021-
Map releaseJul 14, 2021-
UpdateSep 22, 2021-
Current statusSep 22, 2021Processing site: RCSB / Status: Released

-
Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 0.0618
  • Imaged by UCSF Chimera
  • Download
  • Surface view colored by radius
  • Surface level: 0.0618
  • Imaged by UCSF Chimera
  • Download
  • Surface view with fitted model
  • Atomic models: PDB-7kx7
  • Surface level: 0.0618
  • Imaged by UCSF Chimera
  • Download
Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

emd_23061.png

Downloads & links

-
Map

FileDownload / File: emd_23061.map.gz / Format: CCP4 / Size: 2.8 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationEf PIWI-piRNA local resolution sharpened map
Voxel sizeX=Y=Z: 1.15 Å
Density
Contour LevelBy AUTHOR: 0.0618 / Movie #1: 0.0618
Minimum - Maximum-0.24108344 - 0.39020115
Average (Standard dev.)0.0008398991 (±0.028360372)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions909090
Spacing909090
CellA=B=C: 103.5 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z1.151.151.15
M x/y/z909090
origin x/y/z0.0000.0000.000
length x/y/z103.500103.500103.500
α/β/γ90.00090.00090.000
start NX/NY/NZ000
NX/NY/NZ300300300
MAP C/R/S123
start NC/NR/NS000
NC/NR/NS909090
D min/max/mean-0.2410.3900.001

-
Supplemental data

-
Mask #1

Fileemd_23061_msk_1.map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

-
Half map: Ef PIWI-piRNA odd half map

Fileemd_23061_half_map_1.map
AnnotationEf PIWI-piRNA odd half map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

-
Half map: Ef PIWI-piRNA even half map

Fileemd_23061_half_map_2.map
AnnotationEf PIWI-piRNA even half map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

-
Sample components

-
Entire : Complex between EfPiwi and piRNA

EntireName: Complex between EfPiwi and piRNA
Components
  • Complex: Complex between EfPiwi and piRNA
    • Complex: Piwi-A
      • Protein or peptide: Piwi-A
    • Complex: piRNA
      • RNA: RNA (5'-R(P*UP*CP*UP*CP*AP*GP*(OMC))-3')
  • Ligand: MAGNESIUM ION

-
Supramolecule #1: Complex between EfPiwi and piRNA

SupramoleculeName: Complex between EfPiwi and piRNA / type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1-#2
Molecular weightTheoretical: 83 KDa

-
Supramolecule #2: Piwi-A

SupramoleculeName: Piwi-A / type: complex / ID: 2 / Parent: 1 / Macromolecule list: #1
Source (natural)Organism: Ephydatia fluviatilis (invertebrata)
Recombinant expressionOrganism: Spodoptera frugiperda (fall armyworm)

-
Supramolecule #3: piRNA

SupramoleculeName: piRNA / type: complex / ID: 3 / Parent: 1 / Macromolecule list: #2
Source (natural)Organism: synthetic construct (others)

-
Macromolecule #1: Piwi-A

MacromoleculeName: Piwi-A / type: protein_or_peptide / ID: 1 / Number of copies: 1 / Enantiomer: LEVO
EC number: Hydrolases; Acting on ester bonds; Endoribonucleases producing 5'-phosphomonoesters
Source (natural)Organism: Ephydatia fluviatilis (invertebrata)
Molecular weightTheoretical: 88.232594 KDa
Recombinant expressionOrganism: Spodoptera frugiperda (fall armyworm)
SequenceString: SLNVESSMVS QRGSSGQPVP VSANYLPLKG NMDGVFKYAV GFNPPVEDIR SRSQLLNEHK ELIGLTRVFD GSTLYVPKRI CEQRLDLMS TRQTDGASIK VTISLVDSVK NRDVVQLMNV IFKRILRSLK LQRIGRDYYD ANSPLEVPQH KMQLWPGYVT A INRHEGGL ...String:
SLNVESSMVS QRGSSGQPVP VSANYLPLKG NMDGVFKYAV GFNPPVEDIR SRSQLLNEHK ELIGLTRVFD GSTLYVPKRI CEQRLDLMS TRQTDGASIK VTISLVDSVK NRDVVQLMNV IFKRILRSLK LQRIGRDYYD ANSPLEVPQH KMQLWPGYVT A INRHEGGL MLVLDVSHRV MKTDTALDFL YELYHFNQDK FREEAFKQLV GSVVLTRYNN RTYEIDDIAW DKNPRCAFQD HA GSQITFV DYYKRAYDLD ITDLEQPLLI HRPKKKQRGK QDEGRKEVEE MVCLVPELCA MTGLTDAARS DFKVMKDLAV HTR VPPEKR AESFRKFIQR LNTTKEASEL LHSWGLVLDS RMLDMQGRRL PPEKILFKHS SIVANMEADW SRECLKEHVI SAVS LLDWA VLFVRKDQGK ATDFVNMLSK VCPPIGMEVH EPKMVEVVND RTESYLRALR ELIAPRLQMV VIVFPTSRDD RYSAV KKLC CIESPIPSQV LIARTITQQQ KLRSVAQKVA LQMNAKLGGE LWAVEIPLKS CMVVGIDVYH DKSYGNKSIA GFVAST NPS FTRWYSRTAM QEQSQELIHE LKLCMQAALK KYNEMNQSLP ERIIVFRDGV GEGREEYVSE FEVPQFNSCF SIFGENY CP KLAVVVVQKR ITTRIFGRSG HSYDNPPPGV IVDHTITKSY DFYLVSQHVR QGTVSPTYYR VIYDKSGLKP DHLQRLTY K LTHMYYNWPG TIRTPAPCNY AHKLAFLVGK SLHRDPAHEL SDRLFFL

-
Macromolecule #2: RNA (5'-R(P*UP*CP*UP*CP*AP*GP*(OMC))-3')

MacromoleculeName: RNA (5'-R(P*UP*CP*UP*CP*AP*GP*(OMC))-3') / type: rna / ID: 2 / Number of copies: 1
Source (natural)Organism: synthetic construct (others)
Molecular weightTheoretical: 8.029815 KDa
SequenceString:
UCUCUUGAGU UGGACAAAUG GCAG(OMC)

-
Macromolecule #3: MAGNESIUM ION

MacromoleculeName: MAGNESIUM ION / type: ligand / ID: 3 / Number of copies: 2 / Formula: MG
Molecular weightTheoretical: 24.305 Da

-
Experimental details

-
Structure determination

Methodcryo EM
Processingsingle particle reconstruction
Aggregation stateparticle

-
Sample preparation

Concentration2.5 mg/mL
BufferpH: 8
GridModel: UltrAuFoil R1.2/1.3 / Material: GOLD / Mesh: 300 / Support film - Material: GOLD / Support film - topology: HOLEY ARRAY / Pretreatment - Type: PLASMA CLEANING
VitrificationCryogen name: ETHANE / Chamber humidity: 97 % / Chamber temperature: 277.15 K / Instrument: HOMEMADE PLUNGER
Details: Freezing was carried out in a cold room at 4 degree C and relative humidity between 95%-98%. 3.5 uL sample was applied to plasma cleaned grid and manually blotted with Whatman 1 filter paper ...Details: Freezing was carried out in a cold room at 4 degree C and relative humidity between 95%-98%. 3.5 uL sample was applied to plasma cleaned grid and manually blotted with Whatman 1 filter paper for 5-7 sec before plunge freezing in liquid ethane at -179 degree C..

-
Electron microscopy

MicroscopeFEI TALOS ARCTICA
Electron beamAcceleration voltage: 200 kV / Electron source: FIELD EMISSION GUN
Electron opticsC2 aperture diameter: 70.0 µm / Calibrated magnification: 43478 / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELDBright-field microscopy / Cs: 2.7 mm / Nominal defocus max: 1.6 µm / Nominal defocus min: 1.0 µm / Nominal magnification: 36000
Sample stageSpecimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN
TemperatureMin: 77.0 K / Max: 79.0 K
Image recordingFilm or detector model: GATAN K2 SUMMIT (4k x 4k) / Detector mode: COUNTING / Digitization - Dimensions - Width: 3710 pixel / Digitization - Dimensions - Height: 3838 pixel / Digitization - Sampling interval: 5.0 µm / Digitization - Frames/image: 1-48 / Number real images: 1765 / Average exposure time: 12.0 sec. / Average electron dose: 47.33 e/Å2
Experimental equipment
Model: Talos Arctica / Image courtesy: FEI Company

-
Image processing

Particle selectionNumber selected: 3280351
Details: Laplacian of Gaussian based automated particle picking program in RELION was used.
CTF correctionSoftware - Name: CTFFIND (ver. 4)
Startup modelType of model: PDB ENTRY
PDB model - PDB ID:

5guh


Details: 40 Angstrom low pass filtered map was generated from Siwi crystal structure (PDB ID 5GUH) using the molmap function in UCSF Chimera.
Initial angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: RELION (ver. 2)
Final angle assignmentType: PROJECTION MATCHING / Software - Name: RELION (ver. 2)
Final reconstructionApplied symmetry - Point group: C1 (asymmetric) / Algorithm: BACK PROJECTION / Resolution.type: BY AUTHOR / Resolution: 3.8 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: RELION (ver. 2.0) / Number images used: 1862936
DetailsBeam-induced motion correction and radiation damage compensation over spatial frequencies (dose-weighting) of the raw movies, was performed using UCSF MotionCor2 implemented in the Appion.
FSC plot (resolution estimation)

+
About Yorodumi

-
News

-
Feb 9, 2022. New format data for meta-information of EMDB entries

New format data for meta-information of EMDB entries

  • Version 3 of the EMDB header file is now the official format.
  • The previous official version 1.9 will be removed from the archive.

Related info.:EMDB header

External links:wwPDB to switch to version 3 of the EMDB data model

-
Aug 12, 2020. Covid-19 info

Covid-19 info

URL: https://pdbj.org/emnavi/covid19.php

New page: Covid-19 featured information page in EM Navigator.

Related info.:Covid-19 info / Mar 5, 2020. Novel coronavirus structure data

+
Mar 5, 2020. Novel coronavirus structure data

Novel coronavirus structure data

Related info.:Yorodumi Speices / Aug 12, 2020. Covid-19 info

External links:COVID-19 featured content - PDBj / Molecule of the Month (242)：Coronavirus Proteases

+
Jan 31, 2019. EMDB accession codes are about to change! (news from PDBe EMDB page)

EMDB accession codes are about to change! (news from PDBe EMDB page)

  • The allocation of 4 digits for EMDB accession codes will soon come to an end. Whilst these codes will remain in use, new EMDB accession codes will include an additional digit and will expand incrementally as the available range of codes is exhausted. The current 4-digit format prefixed with “EMD-” (i.e. EMD-XXXX) will advance to a 5-digit format (i.e. EMD-XXXXX), and so on. It is currently estimated that the 4-digit codes will be depleted around Spring 2019, at which point the 5-digit format will come into force.
  • The EM Navigator/Yorodumi systems omit the EMD- prefix.

Related info.:Q: What is EMD? / ID/Accession-code notation in Yorodumi/EM Navigator

External links:EMDB Accession Codes are Changing Soon! / Contact to PDBj

+
Jul 12, 2017. Major update of PDB

Major update of PDB

  • wwPDB released updated PDB data conforming to the new PDBx/mmCIF dictionary.
  • This is a major update changing the version number from 4 to 5, and with Remediation, in which all the entries are updated.
  • In this update, many items about electron microscopy experimental information are reorganized (e.g. em_software).
  • Now, EM Navigator and Yorodumi are based on the updated data.

External links:wwPDB Remediation / Enriched Model Files Conforming to OneDep Data Standards Now Available in the PDB FTP Archive

-
Yorodumi

Thousand views of thousand structures

  • Yorodumi is a browser for structure data from EMDB, PDB, SASBDB, etc.
  • This page is also the successor to EM Navigator detail page, and also detail information page/front-end page for Omokage search.
  • The word "yorodu" (or yorozu) is an old Japanese word meaning "ten thousand". "mi" (miru) is to see.

Related info.:EMDB / PDB / SASBDB / Comparison of 3 databanks / Yorodumi Search / Aug 31, 2016. New EM Navigator & Yorodumi / Yorodumi Papers / Jmol/JSmol / Function and homology information / Changes in new EM Navigator and Yorodumi

Read more