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- EMDB-33490: Cryo-EM structure of an apo-form of human DICER -

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Basic information

Entry
Database: EMDB / ID: EMD-33490
TitleCryo-EM structure of an apo-form of human DICER
Map data
Sample
  • Complex: monomeric form of human DICER
    • Protein or peptide: Endoribonuclease Dicer
KeywordsDicer / RNaseIII / RNA-binding / micro-RNA processing / GENE REGULATION
Function / homology
Function and homology information


peripheral nervous system myelin formation / global gene silencing by mRNA cleavage / pre-miRNA binding / tRNA-derived small RNA (tsRNA or tRNA-related fragment, tRF) biogenesis / Small interfering RNA (siRNA) biogenesis / negative regulation of Schwann cell proliferation / tRNA decay / apoptotic DNA fragmentation / positive regulation of myelination / ribonuclease III ...peripheral nervous system myelin formation / global gene silencing by mRNA cleavage / pre-miRNA binding / tRNA-derived small RNA (tsRNA or tRNA-related fragment, tRF) biogenesis / Small interfering RNA (siRNA) biogenesis / negative regulation of Schwann cell proliferation / tRNA decay / apoptotic DNA fragmentation / positive regulation of myelination / ribonuclease III / deoxyribonuclease I activity / positive regulation of Schwann cell differentiation / nerve development / RISC-loading complex / miRNA metabolic process / RISC complex assembly / ribonuclease III activity / miRNA processing / pre-miRNA processing / siRNA processing / siRNA binding / Regulation of MITF-M-dependent genes involved in apoptosis / M-decay: degradation of maternal mRNAs by maternally stored factors / RISC complex / MicroRNA (miRNA) biogenesis / negative regulation of tumor necrosis factor production / negative regulation of tumor necrosis factor-mediated signaling pathway / RNA endonuclease activity / neuron projection morphogenesis / helicase activity / double-stranded RNA binding / protein domain specific binding / negative regulation of gene expression / perinuclear region of cytoplasm / negative regulation of transcription by RNA polymerase II / RNA binding / extracellular exosome / ATP binding / nucleus / metal ion binding / cytosol / cytoplasm
Similarity search - Function
Dicer, double-stranded RNA-binding domain / : / Dicer, partner-binding domain / Dicer, dsRNA-binding domain / : / Dicer, platform domain / Dicer dimerisation domain / Dicer dimerisation domain superfamily / Dicer dimerisation domain / Dicer double-stranded RNA-binding fold domain profile. ...Dicer, double-stranded RNA-binding domain / : / Dicer, partner-binding domain / Dicer, dsRNA-binding domain / : / Dicer, platform domain / Dicer dimerisation domain / Dicer dimerisation domain superfamily / Dicer dimerisation domain / Dicer double-stranded RNA-binding fold domain profile. / Ribonuclease III family signature. / Ribonuclease III domain / Ribonuclease III family domain profile. / Ribonuclease III family / Ribonuclease III domain / PAZ domain superfamily / PAZ / PAZ domain / PAZ domain profile. / PAZ domain / Double-stranded RNA binding motif / Ribonuclease III, endonuclease domain superfamily / Double stranded RNA-binding domain (dsRBD) profile. / Double-stranded RNA-binding domain / DEAD/DEAH box helicase domain / DEAD/DEAH box helicase / Helicase conserved C-terminal domain / helicase superfamily c-terminal domain / Superfamilies 1 and 2 helicase C-terminal domain profile. / Superfamilies 1 and 2 helicase ATP-binding type-1 domain profile. / DEAD-like helicases superfamily / Helicase, C-terminal / Helicase superfamily 1/2, ATP-binding domain / P-loop containing nucleoside triphosphate hydrolase
Similarity search - Domain/homology
Endoribonuclease Dicer
Similarity search - Component
Biological speciesHomo sapiens (human)
Methodsingle particle reconstruction / cryo EM / Resolution: 4.04 Å
AuthorsLee H / Roh S-H
Funding support Korea, Republic Of, 1 items
OrganizationGrant numberCountry
National Research Foundation (NRF, Korea) Korea, Republic Of
CitationJournal: Nature / Year: 2023
Title: Structure of the human DICER-pre-miRNA complex in a dicing state.
Authors: Young-Yoon Lee / Hansol Lee / Haedong Kim / V Narry Kim / Soung-Hun Roh /
Abstract: Dicer has a key role in small RNA biogenesis, processing double-stranded RNAs (dsRNAs). Human DICER (hDICER, also known as DICER1) is specialized for cleaving small hairpin structures such as ...Dicer has a key role in small RNA biogenesis, processing double-stranded RNAs (dsRNAs). Human DICER (hDICER, also known as DICER1) is specialized for cleaving small hairpin structures such as precursor microRNAs (pre-miRNAs) and has limited activity towards long dsRNAs-unlike its homologues in lower eukaryotes and plants, which cleave long dsRNAs. Although the mechanism by which long dsRNAs are cleaved has been well documented, our understanding of pre-miRNA processing is incomplete because structures of hDICER in a catalytic state are lacking. Here we report the cryo-electron microscopy structure of hDICER bound to pre-miRNA in a dicing state and uncover the structural basis of pre-miRNA processing. hDICER undergoes large conformational changes to attain the active state. The helicase domain becomes flexible, which allows the binding of pre-miRNA to the catalytic valley. The double-stranded RNA-binding domain relocates and anchors pre-miRNA in a specific position through both sequence-independent and sequence-specific recognition of the newly identified 'GYM motif'. The DICER-specific PAZ helix is also reoriented to accommodate the RNA. Furthermore, our structure identifies a configuration of the 5' end of pre-miRNA inserted into a basic pocket. In this pocket, a group of arginine residues recognize the 5' terminal base (disfavouring guanine) and terminal monophosphate; this explains the specificity of hDICER and how it determines the cleavage site. We identify cancer-associated mutations in the 5' pocket residues that impair miRNA biogenesis. Our study reveals how hDICER recognizes pre-miRNAs with stringent specificity and enables a mechanistic understanding of hDICER-related diseases.
History
DepositionMay 26, 2022-
Header (metadata) releaseMar 8, 2023-
Map releaseMar 8, 2023-
UpdateJul 3, 2024-
Current statusJul 3, 2024Processing site: PDBj / Status: Released

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Structure visualization

Supplemental images

emd_33490.png

Downloads & links

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Map

FileDownload / File: emd_33490.map.gz / Format: CCP4 / Size: 103 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
1.1 Å/pix.
x 300 pix.
= 330.6 Å		1.1 Å/pix.
x 300 pix.
= 330.6 Å		1.1 Å/pix.
x 300 pix.
= 330.6 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

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Images are generated by Spider.

Voxel sizeX=Y=Z: 1.102 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.105
Minimum - Maximum-0.7675448 - 1.1569057
Average (Standard dev.)0.00045772223 (±0.017673321)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions300300300
Spacing300300300
CellA=B=C: 330.6 Å
α=β=γ: 90.0 °

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Supplemental data

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Half map: #2

Fileemd_33490_half_map_1.map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: #1

Fileemd_33490_half_map_2.map
Projections & Slices
AxesZYX

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Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Sample components

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Entire : monomeric form of human DICER

EntireName: monomeric form of human DICER
Components
  • Complex: monomeric form of human DICER
    • Protein or peptide: Endoribonuclease Dicer

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Supramolecule #1: monomeric form of human DICER

SupramoleculeName: monomeric form of human DICER / type: complex / ID: 1 / Parent: 0 / Macromolecule list: all
Source (natural)Organism: Homo sapiens (human)

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Macromolecule #1: Endoribonuclease Dicer

MacromoleculeName: Endoribonuclease Dicer / type: protein_or_peptide / ID: 1 / Number of copies: 1 / Enantiomer: LEVO / EC number: ribonuclease III
Source (natural)Organism: Homo sapiens (human)
Molecular weightTheoretical: 218.947328 KDa
Recombinant expressionOrganism: Homo sapiens (human)
SequenceString: MKSPALQPLS MAGLQLMTPA SSPMGPFFGL PWQQEAIHDN IYTPRKYQVE LLEAALDHNT IVCLNTGSGK TFIAVLLTKE LSYQIRGDF SRNGKRTVFL VNSANQVAQQ VSAVRTHSDL KVGEYSNLEV NASWTKERWN QEFTKHQVLI MTCYVALNVL K NGYLSLSD ...String:
MKSPALQPLS MAGLQLMTPA SSPMGPFFGL PWQQEAIHDN IYTPRKYQVE LLEAALDHNT IVCLNTGSGK TFIAVLLTKE LSYQIRGDF SRNGKRTVFL VNSANQVAQQ VSAVRTHSDL KVGEYSNLEV NASWTKERWN QEFTKHQVLI MTCYVALNVL K NGYLSLSD INLLVFDECH LAILDHPYRE IMKLCENCPS CPRILGLTAS ILNGKCDPEE LEEKIQKLEK ILKSNAETAT DL VVLDRYT SQPCEIVVDC GPFTDRSGLY ERLLMELEEA LNFINDCNIS VHSKERDSTL ISKQILSDCR AVLVVLGPWC ADK VAGMMV RELQKYIKHE QEELHRKFLL FTDTFLRKIH ALCEEHFSPA SLDLKFVTPK VIKLLEILRK YKPYERQQFE SVEW YNNRN QDNYVSWSDS EDDDEDEEIE EKEKPETNFP SPFTNILCGI IFVERRYTAV VLNRLIKEAG KQDPELAYIS SNFIT GHGI GKNQPRNKQM EAEFRKQEEV LRKFRAHETN LLIATSIVEE GVDIPKCNLV VRFDLPTEYR SYVQSKGRAR APISNY IML ADTDKIKSFE EDLKTYKAIE KILRNKCSKS VDTGETDIDP VMDDDDVFPP YVLRPDDGGP RVTINTAIGH INRYCAR LP SDPFTHLAPK CRTRELPDGT FYSTLYLPIN SPLRASIVGP PMSCVRLAER VVALICCEKL HKIGELDDHL MPVGKETV K YEEELDLHDE EETSVPGRPG STKRRQCYPK AIPECLRDSY PRPDQPCYLY VIGMVLTTPL PDELNFRRRK LYPPEDTTR CFGILTAKPI PQIPHFPVYT RSGEVTISIE LKKSGFMLSL QMLELITRLH QYIFSHILRL EKPALEFKPT DADSAYCVLP LNVVNDSST LDIDFKFMED IEKSEARIGI PSTKYTKETP FVFKLEDYQD AVIIPRYRNF DQPHRFYVAD VYTDLTPLSK F PSPEYETF AEYYKTKYNL DLTNLNQPLL DVDHTSSRLN LLTPRHLNQK GKALPLSSAE KRKAKWESLQ NKQILVPELC AI HPIPASL WRKAVCLPSI LYRLHCLLTA EELRAQTASD AGVGVRSLPA DFRYPNLDFG WKKSIDSKSF ISISNSSSAE NDN YCKHST IVPENAAHQG ANRTSSLENH DQMSVNCRTL LSESPGKLHV EVSADLTAIN GLSYNQNLAN GSYDLANRDF CQGN QLNYY KQEIPVQPTT SYSIQNLYSY ENQPQPSDEC TLLSNKYLDG NANKSTSDGS PVMAVMPGTT DTIQVLKGRM DSEQS PSIG YSSRTLGPNP GLILQALTLS NASDGFNLER LEMLGDSFLK HAITTYLFCT YPDAHEGRLS YMRSKKVSNC NLYRLG KKK GLPSRMVVSI FDPPVNWLPP GYVVNQDKSN TDKWEKDEMT KDCMLANGKL DEDYEEEDEE EESLMWRAPK EEADYED DF LEYDQEHIRF IDNMLMGSGA FVKKISLSPF STTDSAYEWK MPKKSSLGSM PFSSDFEDFD YSSWDAMCYL DPSKAVEE D DFVVGFWNPS EENCGVDTGK QSISYDLHTE QCIADKSIAD CVEALLGCYL TSCGERAAQL FLCSLGLKVL PVIKRTDRE KALCPTRENF NSQQKNLSVS CAAASVASSR SSVLKDSEYG CLKIPPRCMF DHPDADKTLN HLISGFENFE KKINYRFKNK AYLLQAFTH ASYHYNTITD CYQRLEFLGD AILDYLITKH LYEDPRQHSP GVLTDLRSAL VNNTIFASLA VKYDYHKYFK A VSPELFHV IDDFVQFQLE KNEMQGMDSE LRRSEEDEEK EEDIEVPKAM GDIFESLAGA IYMDSGMSLE TVWQVYYPMM RP LIEKFSA NVPRSPVREL LEMEPETAKF SPAERTYDGK VRVTVEVVGK GKFKGVGRSY RIAKSAAARR ALRSLKANQP QVP NS

UniProtKB: Endoribonuclease Dicer

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Experimental details

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Structure determination

Methodcryo EM
Processingsingle particle reconstruction
Aggregation state3D array

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Sample preparation

BufferpH: 8
GridModel: UltrAuFoil R1.2/1.3 / Material: GOLD / Mesh: 300 / Pretreatment - Type: GLOW DISCHARGE / Pretreatment - Time: 60 sec. / Pretreatment - Atmosphere: AIR
VitrificationCryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 288 K / Instrument: FEI VITROBOT MARK IV / Details: blot force 5 and blot for 2 seconds.

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Electron microscopy

MicroscopeTFS GLACIOS
Image recordingFilm or detector model: FEI FALCON IV (4k x 4k) / Average electron dose: 40.0 e/Å2
Electron beamAcceleration voltage: 200 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Nominal defocus max: 2.75 µm / Nominal defocus min: 1.0 µm

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Image processing

Startup modelType of model: OTHER
Final reconstructionResolution.type: BY AUTHOR / Resolution: 4.04 Å / Resolution method: FSC 0.143 CUT-OFF / Number images used: 128635
Initial angle assignmentType: ANGULAR RECONSTITUTION
Final angle assignmentType: MAXIMUM LIKELIHOOD
FSC plot (resolution estimation)

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