[English] 日本語
Yorodumi- EMDB-30154: Human AAA+ ATPase VCP mutant - T76E, AMP-PNP-bound form, Conforma... -
+Open data
-Basic information
Entry | Database: EMDB / ID: EMD-30154 | |||||||||
---|---|---|---|---|---|---|---|---|---|---|
Title | Human AAA+ ATPase VCP mutant - T76E, AMP-PNP-bound form, Conformation II | |||||||||
Map data | Human AAA ATPase VCP, T76E mutant with AMP-PNP-bound, Conformation II | |||||||||
Sample |
| |||||||||
Biological species | Homo sapiens (human) | |||||||||
Method | single particle reconstruction / cryo EM / Resolution: 4.8 Å | |||||||||
Authors | Yang C / Zhang H | |||||||||
Citation | Journal: Cell Death Differ / Year: 2022 Title: The phosphorylation and dephosphorylation switch of VCP/p97 regulates the architecture of centrosome and spindle. Authors: Kaiyuan Zhu / Yang Cai / Xiaotong Si / Zuodong Ye / Yuanzhu Gao / Chuang Liu / Rui Wang / Zhibin Ma / Huazhang Zhu / Liang Zhang / Shengjin Li / Hongmin Zhang / Jianbo Yue / Abstract: The proper orientation of centrosome and spindle is essential for genome stability; however, the mechanism that governs these processes remains elusive. Here, we demonstrated that polo-like kinase 1 ...The proper orientation of centrosome and spindle is essential for genome stability; however, the mechanism that governs these processes remains elusive. Here, we demonstrated that polo-like kinase 1 (Plk1), a key mitotic kinase, phosphorylates residue Thr76 in VCP/p97 (an AAA-ATPase), at the centrosome from prophase to anaphase. This phosphorylation process recruits VCP to the centrosome and in this way, it regulates centrosome orientation. VCP exhibits strong co-localization with Eg5 (a mitotic kinesin motor), at the mitotic spindle, and the dephosphorylation of Thr76 in VCP is required for the enrichment of both VCP and Eg5 at the spindle, thus ensuring proper spindle architecture and chromosome segregation. We also showed that the phosphatase, PTEN, is responsible for the dephosphorylation of Thr76 in VCP; when PTEN was knocked down, the normal spread of VCP from the centrosome to the spindle was abolished. Cryo-EM structures of VCP and VCP, which represent dephosphorylated and phosphorylated states of VCP, respectively, revealed that the Thr76 phosphorylation modulates VCP by altering the inter-domain and inter-subunit interactions, and ultimately the nucleotide-binding pocket conformation. Interestingly, the tumor growth in nude mice implanted with VCP-reconstituted cancer cells was significantly slower when compared with those implanted with VCP-reconstituted cancer cells. Collectively, our findings demonstrate that the phosphorylation and dephosphorylation switch of VCP regulates the architecture of centrosome and spindle for faithful chromosome segregation. | |||||||||
History |
|
-Structure visualization
Movie |
Movie viewer |
---|---|
Structure viewer | EM map: SurfViewMolmilJmol/JSmol |
Supplemental images |
-Downloads & links
-EMDB archive
Map data | emd_30154.map.gz | 165.4 MB | EMDB map data format | |
---|---|---|---|---|
Header (meta data) | emd-30154-v30.xml emd-30154.xml | 12 KB 12 KB | Display Display | EMDB header |
Images | emd_30154.png | 154 KB | ||
Archive directory | http://ftp.pdbj.org/pub/emdb/structures/EMD-30154 ftp://ftp.pdbj.org/pub/emdb/structures/EMD-30154 | HTTPS FTP |
-Related structure data
-Links
EMDB pages | EMDB (EBI/PDBe) / EMDataResource |
---|
-Map
File | Download / File: emd_30154.map.gz / Format: CCP4 / Size: 178 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES) | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|
Annotation | Human AAA ATPase VCP, T76E mutant with AMP-PNP-bound, Conformation II | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Voxel size | X=Y=Z: 1.073 Å | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Density |
| ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Symmetry | Space group: 1 | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Details | EMDB XML:
CCP4 map header:
|
-Supplemental data
-Sample components
-Entire : Transitional endoplasmic reticulum ATPase, VCP.
Entire | Name: Transitional endoplasmic reticulum ATPase, VCP. |
---|---|
Components |
|
-Supramolecule #1: Transitional endoplasmic reticulum ATPase, VCP.
Supramolecule | Name: Transitional endoplasmic reticulum ATPase, VCP. / type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1 Details: VCP T76E mutant of AMP-PNP-bound form,Conformation II |
---|---|
Source (natural) | Organism: Homo sapiens (human) / Location in cell: cytoplasm nucleus ER |
Recombinant expression | Organism: Escherichia coli (E. coli) / Recombinant strain: T7 SHuffle (NEB C3026) / Recombinant plasmid: pET |
Molecular weight | Experimental: 97 kDa/nm |
-Experimental details
-Structure determination
Method | cryo EM |
---|---|
Processing | single particle reconstruction |
Aggregation state | particle |
-Sample preparation
Concentration | 2 mg/mL | |||||||||||||||
---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|
Buffer | pH: 7.5 Component:
| |||||||||||||||
Vitrification | Cryogen name: ETHANE / Instrument: FEI VITROBOT MARK IV |
-Electron microscopy
Microscope | FEI TITAN KRIOS |
---|---|
Electron beam | Acceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN |
Electron optics | Illumination mode: OTHER / Imaging mode: BRIGHT FIELDBright-field microscopy |
Image recording | Film or detector model: FEI FALCON III (4k x 4k) / Detector mode: COUNTING / Average electron dose: 50.0 e/Å2 |
Experimental equipment | Model: Titan Krios / Image courtesy: FEI Company |
-Image processing
CTF correction | Software - Name: RELION (ver. 3.0.6) |
---|---|
Startup model | Type of model: EMDB MAP EMDB ID: |
Initial angle assignment | Type: RANDOM ASSIGNMENT / Software - Name: cisTEM (ver. Beta -1.0.0) |
Final angle assignment | Type: NOT APPLICABLE / Software - Name: cisTEM (ver. Beta -1.0.0) |
Final reconstruction | Applied symmetry - Point group: C6 (6 fold cyclic) / Resolution.type: BY AUTHOR / Resolution: 4.8 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: cisTEM (ver. Beta -1.0.0) / Number images used: 533495 |
-Atomic model buiding 1
Refinement | Protocol: RIGID BODY FIT |
---|