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- PDB-8rgg: Structure of dynein-2 intermediate chain DYNC2I2 (WDR34) in compl... -

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Entry
Database: PDB / ID: 8rgg
TitleStructure of dynein-2 intermediate chain DYNC2I2 (WDR34) in complex with dynein-2 heavy chain DYNC2H1.
Components
  • Cytoplasmic dynein 2 intermediate chain 1
  • Cytoplasmic dynein 2 intermediate chain 2
  • Dynein light chain 1, cytoplasmic
  • Dynein light chain roadblock-type 1
  • Methylated-DNA--protein-cysteine methyltransferase,Cytoplasmic dynein 2 heavy chain 1
KeywordsTRANSPORT PROTEIN / dynein / cilia / intraflagellar transport / complex
Function / homology
Function and homology information


MGMT-mediated DNA damage reversal / nitric-oxide synthase inhibitor activity / negative regulation of DNA strand resection involved in replication fork processing / deoxyribonuclease inhibitor activity / visual behavior / intraciliary retrograde transport / methylated-DNA-[protein]-cysteine S-methyltransferase / methylated-DNA-[protein]-cysteine S-methyltransferase activity / Activation of BIM and translocation to mitochondria / cilium movement involved in cell motility ...MGMT-mediated DNA damage reversal / nitric-oxide synthase inhibitor activity / negative regulation of DNA strand resection involved in replication fork processing / deoxyribonuclease inhibitor activity / visual behavior / intraciliary retrograde transport / methylated-DNA-[protein]-cysteine S-methyltransferase / methylated-DNA-[protein]-cysteine S-methyltransferase activity / Activation of BIM and translocation to mitochondria / cilium movement involved in cell motility / intraciliary transport / 9+2 motile cilium / dynein light chain binding / dynein heavy chain binding / spinal cord motor neuron differentiation / motile cilium assembly / DNA-methyltransferase activity / embryonic skeletal system morphogenesis / ciliary tip / negative regulation of phosphorylation / Intraflagellar transport / negative regulation of nitric oxide biosynthetic process / dynein complex / DNA ligation / minus-end-directed microtubule motor activity / coronary vasculature development / COPI-independent Golgi-to-ER retrograde traffic / non-motile cilium assembly / protein localization to cilium / DNA alkylation repair / cytoplasmic dynein complex / dynein light intermediate chain binding / positive regulation of smoothened signaling pathway / ciliary plasm / dorsal/ventral pattern formation / enzyme inhibitor activity / determination of left/right symmetry / embryonic limb morphogenesis / positive regulation of double-strand break repair / microtubule motor activity / ciliary base / dynein intermediate chain binding / Macroautophagy / microtubule-based movement / pericentriolar material / Golgi organization / positive regulation of insulin secretion involved in cellular response to glucose stimulus / cytoskeletal motor activity / axoneme / tertiary granule membrane / ficolin-1-rich granule membrane / spermatid development / cilium assembly / Hedgehog 'off' state / COPI-mediated anterograde transport / Amplification of signal from unattached kinetochores via a MAD2 inhibitory signal / Mitotic Prometaphase / EML4 and NUDC in mitotic spindle formation / Loss of Nlp from mitotic centrosomes / Loss of proteins required for interphase microtubule organization from the centrosome / forebrain development / Recruitment of mitotic centrosome proteins and complexes / axon cytoplasm / Recruitment of NuMA to mitotic centrosomes / Anchoring of the basal body to the plasma membrane / HSP90 chaperone cycle for steroid hormone receptors (SHR) in the presence of ligand / MHC class II antigen presentation / Resolution of Sister Chromatid Cohesion / substantia nigra development / centriole / AURKA Activation by TPX2 / kidney development / ciliary basal body / filopodium / methyltransferase activity / RHO GTPases Activate Formins / cilium / mitotic spindle / kinetochore / protein processing / Aggrephagy / HCMV Early Events / Separation of Sister Chromatids / Regulation of PLK1 Activity at G2/M Transition / apical part of cell / site of double-strand break / scaffold protein binding / methylation / microtubule / cytoskeleton / DNA repair / centrosome / DNA damage response / Neutrophil degranulation / protein-containing complex binding / negative regulation of apoptotic process / apoptotic process / Golgi apparatus / enzyme binding / ATP hydrolysis activity
Similarity search - Function
Cytoplasmic dynein 2 intermediate chain 1 / : / Cytoplasmic dynein 2 heavy chain 1, AAA+ ATPase domain / Dynein light chain roadblock-type 1/2 / Methylguanine DNA methyltransferase, ribonuclease-like domain / 6-O-methylguanine DNA methyltransferase, ribonuclease-like domain / Methylated DNA-protein cysteine methyltransferase domain superfamily / Methylated-DNA-[protein]-cysteine S-methyltransferase, active site / Methylated-DNA--protein-cysteine methyltransferase active site. / Methylated-DNA-[protein]-cysteine S-methyltransferase, DNA binding ...Cytoplasmic dynein 2 intermediate chain 1 / : / Cytoplasmic dynein 2 heavy chain 1, AAA+ ATPase domain / Dynein light chain roadblock-type 1/2 / Methylguanine DNA methyltransferase, ribonuclease-like domain / 6-O-methylguanine DNA methyltransferase, ribonuclease-like domain / Methylated DNA-protein cysteine methyltransferase domain superfamily / Methylated-DNA-[protein]-cysteine S-methyltransferase, active site / Methylated-DNA--protein-cysteine methyltransferase active site. / Methylated-DNA-[protein]-cysteine S-methyltransferase, DNA binding / Methylated DNA-protein cysteine methyltransferase, DNA binding domain / 6-O-methylguanine DNA methyltransferase, DNA binding domain / Dynein light chain, type 1/2, conserved site / Dynein light chain type 1 signature. / : / Dynein light chain, type 1/2 / Dynein light chain type 1 / Dynein light chain type 1 / Dynein light chain superfamily / Roadblock/LAMTOR2 domain / Roadblock/LC7 domain / Roadblock/LC7 domain / Dynein heavy chain, C-terminal domain / Dynein heavy chain, C-terminal domain, barrel region / Dynein heavy chain C-terminal domain / : / Dynein heavy chain, ATPase lid domain / P-loop containing dynein motor region / Dynein heavy chain, tail / Dynein heavy chain, N-terminal region 1 / Dynein heavy chain region D6 P-loop domain / Dynein heavy chain, linker / Dynein heavy chain, AAA module D4 / Dynein heavy chain, coiled coil stalk / Dynein heavy chain / Dynein heavy chain, hydrolytic ATP-binding dynein motor region / Dynein heavy chain, ATP-binding dynein motor region / Dynein heavy chain AAA lid domain / Dynein heavy chain AAA lid domain superfamily / Dynein heavy chain, domain 2, N-terminal / Dynein heavy chain, linker, subdomain 3 / Dynein heavy chain, AAA1 domain, small subdomain / Dynein heavy chain region D6 P-loop domain / Dynein heavy chain, N-terminal region 2 / Hydrolytic ATP binding site of dynein motor region / Microtubule-binding stalk of dynein motor / P-loop containing dynein motor region D4 / ATP-binding dynein motor region / Dynein heavy chain AAA lid domain / Trp-Asp (WD) repeats circular profile. / WD domain, G-beta repeat / WD40 repeats / WD40 repeat / WD40-repeat-containing domain superfamily / Winged helix-like DNA-binding domain superfamily / WD40/YVTN repeat-like-containing domain superfamily / ATPases associated with a variety of cellular activities / AAA+ ATPase domain / P-loop containing nucleoside triphosphate hydrolase
Similarity search - Domain/homology
Methylated-DNA--protein-cysteine methyltransferase / Dynein light chain 1, cytoplasmic / Cytoplasmic dynein 2 heavy chain 1 / Cytoplasmic dynein 2 intermediate chain 1 / Cytoplasmic dynein 2 intermediate chain 2 / Dynein light chain roadblock-type 1
Similarity search - Component
Biological speciesHomo sapiens (human)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 4 Å
AuthorsMukhopadhyay, A.G. / Toropova, K. / Daly, L. / Wells, J. / Vuolo, L. / Mladenov, M. / Seda, M. / Jenkins, D. / Stephens, D.J. / Roberts, A.J.
Funding support United Kingdom, 9items
OrganizationGrant numberCountry
Wellcome Trust217186/Z/19/Z United Kingdom
Wellcome Trust206166/Z/17/Z United Kingdom
Wellcome Trust202679/Z/16/Z United Kingdom
Biotechnology and Biological Sciences Research Council (BBSRC)BB/P008348/1 United Kingdom
Biotechnology and Biological Sciences Research Council (BBSRC)BB/S007202/1 United Kingdom
Royal SocietyRG170260 United Kingdom
Biotechnology and Biological Sciences Research Council (BBSRC)BB/S013024/1 United Kingdom
Biotechnology and Biological Sciences Research Council (BBSRC)BB/S005390/1 United Kingdom
Wellcome Trust210585/Z/18/Z United Kingdom
CitationJournal: EMBO J / Year: 2024
Title: Structure and tethering mechanism of dynein-2 intermediate chains in intraflagellar transport.
Authors: Aakash G Mukhopadhyay / Katerina Toropova / Lydia Daly / Jennifer N Wells / Laura Vuolo / Miroslav Mladenov / Marian Seda / Dagan Jenkins / David J Stephens / Anthony J Roberts /
Abstract: Dynein-2 is a large multiprotein complex that powers retrograde intraflagellar transport (IFT) of cargoes within cilia/flagella, but the molecular mechanism underlying this function is still emerging. ...Dynein-2 is a large multiprotein complex that powers retrograde intraflagellar transport (IFT) of cargoes within cilia/flagella, but the molecular mechanism underlying this function is still emerging. Distinctively, dynein-2 contains two identical force-generating heavy chains that interact with two different intermediate chains (WDR34 and WDR60). Here, we dissect regulation of dynein-2 function by WDR34 and WDR60 using an integrative approach including cryo-electron microscopy and CRISPR/Cas9-enabled cell biology. A 3.9 Å resolution structure shows how WDR34 and WDR60 use surprisingly different interactions to engage equivalent sites of the two heavy chains. We show that cilia can assemble in the absence of either WDR34 or WDR60 individually, but not both subunits. Dynein-2-dependent distribution of cargoes depends more strongly on WDR60, because the unique N-terminal extension of WDR60 facilitates dynein-2 targeting to cilia. Strikingly, this N-terminal extension can be transplanted onto WDR34 and retain function, suggesting it acts as a flexible tether to the IFT "trains" that assemble at the ciliary base. We discuss how use of unstructured tethers represents an emerging theme in IFT train interactions.
History
DepositionDec 13, 2023Deposition site: PDBE / Processing site: PDBE
Revision 1.0Mar 20, 2024Provider: repository / Type: Initial release
Revision 1.1Apr 10, 2024Group: Database references / Category: citation
Item: _citation.journal_volume / _citation.page_first / _citation.page_last

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

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Assembly

Deposited unit
B: Methylated-DNA--protein-cysteine methyltransferase,Cytoplasmic dynein 2 heavy chain 1
C: Cytoplasmic dynein 2 intermediate chain 1
D: Cytoplasmic dynein 2 intermediate chain 2
G: Dynein light chain roadblock-type 1
H: Dynein light chain roadblock-type 1
I: Dynein light chain 1, cytoplasmic
J: Dynein light chain 1, cytoplasmic


Theoretical massNumber of molelcules
Total (without water)741,3607
Polymers741,3607
Non-polymers00
Water00
1


  • Idetical with deposited unit
  • defined by author&software
  • Evidence: electron microscopy, not applicable
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1

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Components

#1: Protein Methylated-DNA--protein-cysteine methyltransferase,Cytoplasmic dynein 2 heavy chain 1 / 6-O-methylguanine-DNA methyltransferase / MGMT / O-6-methylguanine-DNA-alkyltransferase / ...6-O-methylguanine-DNA methyltransferase / MGMT / O-6-methylguanine-DNA-alkyltransferase / Cytoplasmic dynein 2 heavy chain / Dynein cytoplasmic heavy chain 2 / Dynein heavy chain 11 / hDHC11 / Dynein heavy chain isotype 1B


Mass: 515223.031 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Details: DYNC2H1 with N-terminal SNAPf tag / Source: (gene. exp.) Homo sapiens (human) / Gene: MGMT, DYNC2H1, DHC1B, DHC2, DNCH2, DYH1B, KIAA1997 / Production host: Spodoptera frugiperda (fall armyworm)
References: UniProt: P16455, UniProt: Q8NCM8, methylated-DNA-[protein]-cysteine S-methyltransferase
#2: Protein Cytoplasmic dynein 2 intermediate chain 1 / Dynein 2 intermediate chain 1 / WD repeat-containing protein 60


Mass: 122865.156 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: DYNC2I1, WDR60 / Production host: Spodoptera frugiperda (fall armyworm) / References: UniProt: Q8WVS4
#3: Protein Cytoplasmic dynein 2 intermediate chain 2 / Dynein 2 intermediate chain 2 / WD repeat-containing protein 34


Mass: 60639.129 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Details: DYNC2I2 (also known as WDR34) with C-terminal Strep tag
Source: (gene. exp.) Homo sapiens (human) / Gene: DYNC2I2, WDR34 / Production host: Spodoptera frugiperda (fall armyworm) / References: UniProt: Q96EX3
#4: Protein Dynein light chain roadblock-type 1 / Bithoraxoid-like protein / BLP / Dynein light chain 2A / cytoplasmic / Dynein-associated protein ...Bithoraxoid-like protein / BLP / Dynein light chain 2A / cytoplasmic / Dynein-associated protein Km23 / Roadblock domain-containing protein 1


Mass: 10934.576 Da / Num. of mol.: 2
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: DYNLRB1, BITH, DNCL2A, DNLC2A, ROBLD1, HSPC162 / Production host: Spodoptera frugiperda (fall armyworm) / References: UniProt: Q9NP97
#5: Protein Dynein light chain 1, cytoplasmic / 8 kDa dynein light chain / DLC8 / Dynein light chain LC8-type 1 / Protein inhibitor of neuronal ...8 kDa dynein light chain / DLC8 / Dynein light chain LC8-type 1 / Protein inhibitor of neuronal nitric oxide synthase / PIN


Mass: 10381.899 Da / Num. of mol.: 2
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: DYNLL1, DLC1, DNCL1, DNCLC1, HDLC1 / Production host: Spodoptera frugiperda (fall armyworm) / References: UniProt: P63167

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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

ComponentName: Dynein-2 complex / Type: COMPLEX / Entity ID: all / Source: RECOMBINANT
Molecular weightExperimental value: NO
Source (natural)Organism: Homo sapiens (human)
Source (recombinant)Organism: Spodoptera frugiperda (fall armyworm)
Buffer solutionpH: 7.5
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 FIELD / Nominal defocus max: 3500 nm / Nominal defocus min: 1500 nm
Image recordingElectron dose: 50.6 e/Å2 / Film or detector model: GATAN K3 BIOQUANTUM (6k x 4k)

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Processing

CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
3D reconstructionResolution: 4 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 113479 / Details: Global resolution 4.0A / Symmetry type: POINT

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