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- PDB-7yg6: Cryo-EM structure of the EfPiwi(N959K) in complex with piRNA -

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Basic information

Entry
Database: PDB / ID: 7yg6
TitleCryo-EM structure of the EfPiwi(N959K) in complex with piRNA
Components
  • Piwi
  • piRNA
KeywordsRNA BINDING PROTEIN/RNA / piRNA / Piwi protein / Argonaute / RNA binding protein / RNA BINDING PROTEIN-RNA complex
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
RNA / RNA (> 10) / Piwi
Similarity search - Component
Biological speciesEphydatia fluviatilis (invertebrata)
Homo sapiens (human)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3.2 Å
AuthorsLi, Z.Q. / Liu, H.B. / Wu, J.P. / Shen, E.Z.
Funding support China, 1items
OrganizationGrant numberCountry
Other privateWestlake Education Foundation (101486021901) China
CitationJournal: Nat Struct Mol Biol / Year: 2024
Title: Mammalian PIWI-piRNA-target complexes reveal features for broad and efficient target silencing.
Authors: Zhiqing Li / Zhenzhen Li / Yuqi Zhang / Lunni Zhou / Qikui Xu / Lili Li / Lin Zeng / Junchao Xue / Huilin Niu / Jing Zhong / Qilu Yu / Dengfeng Li / Miao Gui / Yongping Huang / Shikui Tu / ...Authors: Zhiqing Li / Zhenzhen Li / Yuqi Zhang / Lunni Zhou / Qikui Xu / Lili Li / Lin Zeng / Junchao Xue / Huilin Niu / Jing Zhong / Qilu Yu / Dengfeng Li / Miao Gui / Yongping Huang / Shikui Tu / Zhao Zhang / Chun-Qing Song / Jianping Wu / En-Zhi Shen /
Abstract: The PIWI-interacting RNA (piRNA) pathway is an adaptive defense system wherein piRNAs guide PIWI family Argonaute proteins to recognize and silence ever-evolving selfish genetic elements and ensure ...The PIWI-interacting RNA (piRNA) pathway is an adaptive defense system wherein piRNAs guide PIWI family Argonaute proteins to recognize and silence ever-evolving selfish genetic elements and ensure genome integrity. Driven by this intensive host-pathogen arms race, the piRNA pathway and its targeted transposons have coevolved rapidly in a species-specific manner, but how the piRNA pathway adapts specifically to target silencing in mammals remains elusive. Here, we show that mouse MILI and human HILI piRNA-induced silencing complexes (piRISCs) bind and cleave targets more efficiently than their invertebrate counterparts from the sponge Ephydatia fluviatilis. The inherent functional differences comport with structural features identified by cryo-EM studies of piRISCs. In the absence of target, MILI and HILI piRISCs adopt a wider nucleic-acid-binding channel and display an extended prearranged piRNA seed as compared with EfPiwi piRISC, consistent with their ability to capture targets more efficiently than EfPiwi piRISC. In the presence of target, the seed gate-which enforces seed-target fidelity in microRNA RISC-adopts a relaxed state in mammalian piRISC, revealing how MILI and HILI tolerate seed-target mismatches to broaden the target spectrum. A vertebrate-specific lysine distorts the piRNA seed, shifting the trajectory of the piRNA-target duplex out of the central cleft and toward the PAZ lobe. Functional analyses reveal that this lysine promotes target binding and cleavage. Our study therefore provides a molecular basis for the piRNA targeting mechanism in mice and humans, and suggests that mammalian piRNA machinery can achieve broad target silencing using a limited supply of piRNA species.
History
DepositionJul 11, 2022Deposition site: PDBJ / Processing site: PDBJ
Revision 1.0Jan 24, 2024Provider: repository / Type: Initial release
Revision 1.1May 1, 2024Group: Database references / Category: citation / citation_author
Item: _citation.country / _citation.journal_abbrev ..._citation.country / _citation.journal_abbrev / _citation.journal_id_CSD / _citation.journal_id_ISSN / _citation.pdbx_database_id_DOI / _citation.title / _citation.year
Revision 1.2May 8, 2024Group: Database references / Category: citation / citation_author
Item: _citation.pdbx_database_id_PubMed / _citation.title / _citation_author.name

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

Downloads & links

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Assembly

Deposited unit
B: piRNA
A: Piwi
hetero molecules


Theoretical massNumber of molelcules
Total (without water)100,6604
Polymers100,6122
Non-polymers492
Water0
1


  • Idetical with deposited unit
  • defined by author
  • Evidence: gel filtration
TypeNameSymmetry operationNumber
identity operation1_5551

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Components

#1: RNA chain piRNA /


Mass: 7957.792 Da / Num. of mol.: 1 / Source method: obtained synthetically / Source: (synth.) Homo sapiens (human)
#2: Protein Piwi /


Mass: 92654.094 Da / Num. of mol.: 1 / Mutation: N959K
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Ephydatia fluviatilis (invertebrata) / Gene: EfPiwiA / Production host: Homo sapiens (human) / References: UniProt: D5MRY8
#3: Chemical ChemComp-MG / MAGNESIUM ION


Mass: 24.305 Da / Num. of mol.: 2 / Source method: obtained synthetically / Formula: Mg
Has ligand of interestN

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Experimental details

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Experiment

ExperimentMethod: ELECTRON MICROSCOPY
EM experimentAggregation state: PARTICLE / 3D reconstruction method: single particle reconstruction

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Sample preparation

Component
IDNameTypeEntity IDParent-IDSource
1EfPiwi(N959K)-piRNACOMPLEX#1-#20RECOMBINANT
2EfPiwi(N959K)COMPLEX#21RECOMBINANT
3piRNACOMPLEX#11SYNTHETIC
Molecular weightExperimental value: NO
Source (natural)Organism: Ephydatia fluviatilis (invertebrata)
Source (recombinant)Organism: Homo sapiens (human)
Buffer solutionpH: 8
SpecimenEmbedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
VitrificationCryogen name: ETHANE

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Electron microscopy imaging

Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company
MicroscopyModel: FEI TITAN KRIOS
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: FLOOD BEAM
Electron lensMode: BRIGHT FIELDBright-field microscopy / Nominal defocus max: 2500 nm / Nominal defocus min: 1200 nm / C2 aperture diameter: 50 µm
Image recordingElectron dose: 50 e/Å2 / Film or detector model: GATAN K3 (6k x 4k)

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Processing

CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
3D reconstructionResolution: 3.2 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 359119 / Symmetry type: POINT

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