+Open data
-Basic information
Entry | Database: PDB / ID: 7ux2 | ||||||
---|---|---|---|---|---|---|---|
Title | cryo-EM structure of the Raptor-TFEB-Rag-Ragulator complex | ||||||
Components |
| ||||||
Keywords | SIGNALING PROTEIN / mTORC1 / TFEB / Lysosome biogenesis / Autophagy | ||||||
Function / homology | Function and homology information regulation of cholesterol import / positive regulation of protein localization to lysosome / regulation of cell-substrate junction organization / Gtr1-Gtr2 GTPase complex / regulation of cholesterol efflux / positive regulation of RNA polymerase II regulatory region sequence-specific DNA binding / FNIP-folliculin RagC/D GAP / Ragulator complex / protein localization to cell junction / positive regulation of pentose-phosphate shunt ...regulation of cholesterol import / positive regulation of protein localization to lysosome / regulation of cell-substrate junction organization / Gtr1-Gtr2 GTPase complex / regulation of cholesterol efflux / positive regulation of RNA polymerase II regulatory region sequence-specific DNA binding / FNIP-folliculin RagC/D GAP / Ragulator complex / protein localization to cell junction / positive regulation of pentose-phosphate shunt / antibacterial innate immune response / regulation of TORC1 signaling / TORC1 complex / protein localization to lysosome / TORC1 signaling / positive regulation of odontoblast differentiation / regulation of TOR signaling / endosome organization / cellular response to L-leucine / MTOR signalling / Amino acids regulate mTORC1 / fibroblast migration / lysosome localization / Energy dependent regulation of mTOR by LKB1-AMPK / protein serine/threonine kinase inhibitor activity / protein localization to membrane / kinase activator activity / positive regulation of osteoclast differentiation / cellular response to osmotic stress / enzyme-substrate adaptor activity / positive regulation of transcription by RNA polymerase III / azurophil granule membrane / endosomal transport / regulation of cell size / Macroautophagy / small GTPase-mediated signal transduction / lysosome organization / protein kinase activator activity / RHOJ GTPase cycle / RHOQ GTPase cycle / mTORC1-mediated signalling / cellular response to nutrient levels / social behavior / tertiary granule membrane / CDC42 GTPase cycle / RHOH GTPase cycle / humoral immune response / ficolin-1-rich granule membrane / HSF1-dependent transactivation / RHOG GTPase cycle / TOR signaling / positive regulation of TOR signaling / regulation of receptor recycling / RAC2 GTPase cycle / embryonic placenta development / RAC3 GTPase cycle / response to amino acid / positive regulation of G1/S transition of mitotic cell cycle / positive regulation of autophagy / specific granule membrane / positive regulation of lipid biosynthetic process / protein-membrane adaptor activity / 14-3-3 protein binding / positive regulation of endothelial cell proliferation / positive regulation of TORC1 signaling / tumor necrosis factor-mediated signaling pathway / RAC1 GTPase cycle / cellular response to amino acid starvation / cellular response to starvation / positive regulation of glycolytic process / negative regulation of autophagy / viral genome replication / RNA splicing / guanyl-nucleotide exchange factor activity / positive regulation of peptidyl-threonine phosphorylation / Regulation of PTEN gene transcription / regulation of autophagy / positive regulation of interleukin-8 production / cholesterol homeostasis / regulation of cell growth / cellular response to glucose stimulus / phosphoprotein binding / TP53 Regulates Metabolic Genes / cellular response to amino acid stimulus / Hydrolases; Acting on acid anhydrides; Acting on GTP to facilitate cellular and subcellular movement / response to virus / MAP2K and MAPK activation / negative regulation of cysteine-type endopeptidase activity involved in apoptotic process / protein localization / autophagy / small GTPase binding / cytoplasmic stress granule / positive regulation of protein localization to nucleus / GDP binding / sequence-specific double-stranded DNA binding / late endosome / E3 ubiquitin ligases ubiquitinate target proteins / GTPase binding / positive regulation of peptidyl-serine phosphorylation / glucose homeostasis Similarity search - Function | ||||||
Biological species | Homo sapiens (human) | ||||||
Method | ELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 2.9 Å | ||||||
Authors | Cui, Z. / Hurley, J. | ||||||
Funding support | United States, 1items
| ||||||
Citation | Journal: Nature / Year: 2023 Title: Structure of the lysosomal mTORC1-TFEB-Rag-Ragulator megacomplex. Authors: Zhicheng Cui / Gennaro Napolitano / Mariana E G de Araujo / Alessandra Esposito / Jlenia Monfregola / Lukas A Huber / Andrea Ballabio / James H Hurley / Abstract: The transcription factor TFEB is a master regulator of lysosomal biogenesis and autophagy. The phosphorylation of TFEB by the mechanistic target of rapamycin complex 1 (mTORC1) is unique in its ...The transcription factor TFEB is a master regulator of lysosomal biogenesis and autophagy. The phosphorylation of TFEB by the mechanistic target of rapamycin complex 1 (mTORC1) is unique in its mTORC1 substrate recruitment mechanism, which is strictly dependent on the amino acid-mediated activation of the RagC GTPase activating protein FLCN. TFEB lacks the TOR signalling motif responsible for the recruitment of other mTORC1 substrates. We used cryogenic-electron microscopy to determine the structure of TFEB as presented to mTORC1 for phosphorylation, which we refer to as the 'megacomplex'. Two full Rag-Ragulator complexes present each molecule of TFEB to the mTOR active site. One Rag-Ragulator complex is bound to Raptor in the canonical mode seen previously in the absence of TFEB. A second Rag-Ragulator complex (non-canonical) docks onto the first through a RagC GDP-dependent contact with the second Ragulator complex. The non-canonical Rag dimer binds the first helix of TFEB with a RagC-dependent aspartate clamp in the cleft between the Rag G domains. In cellulo mutation of the clamp drives TFEB constitutively into the nucleus while having no effect on mTORC1 localization. The remainder of the 108-amino acid TFEB docking domain winds around Raptor and then back to RagA. The double use of RagC GDP contacts in both Rag dimers explains the strong dependence of TFEB phosphorylation on FLCN and the RagC GDP state. | ||||||
History |
|
-Structure visualization
Structure viewer | Molecule: MolmilJmol/JSmol |
---|
-Downloads & links
-Download
PDBx/mmCIF format | 7ux2.cif.gz | 1.1 MB | Display | PDBx/mmCIF format |
---|---|---|---|---|
PDB format | pdb7ux2.ent.gz | 940.3 KB | Display | PDB format |
PDBx/mmJSON format | 7ux2.json.gz | Tree view | PDBx/mmJSON format | |
Others | Other downloads |
-Validation report
Summary document | 7ux2_validation.pdf.gz | 1.2 MB | Display | wwPDB validaton report |
---|---|---|---|---|
Full document | 7ux2_full_validation.pdf.gz | 1.2 MB | Display | |
Data in XML | 7ux2_validation.xml.gz | 86.7 KB | Display | |
Data in CIF | 7ux2_validation.cif.gz | 133.2 KB | Display | |
Arichive directory | https://data.pdbj.org/pub/pdb/validation_reports/ux/7ux2 ftp://data.pdbj.org/pub/pdb/validation_reports/ux/7ux2 | HTTPS FTP |
-Related structure data
Related structure data | 26846MC 7uxcC 7uxhC C: citing same article (ref.) M: map data used to model this data |
---|---|
Similar structure data | Similarity search - Function & homologyF&H Search |
-Links
-Assembly
Deposited unit |
|
---|---|
1 |
|
-Components
-Protein , 2 types, 2 molecules AP
#1: Protein | Mass: 149200.016 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Homo sapiens (human) / Gene: RPTOR, KIAA1303, RAPTOR / Production host: Homo sapiens (human) / References: UniProt: Q8N122 |
---|---|
#9: Protein | Mass: 52926.621 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Homo sapiens (human) / Gene: TFEB, BHLHE35 / Production host: Spodoptera frugiperda (fall armyworm) / References: UniProt: P19484 |
-Ras-related GTP-binding protein ... , 2 types, 4 molecules BICJ
#2: Protein | Mass: 36600.195 Da / Num. of mol.: 2 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Homo sapiens (human) / Gene: RRAGA / Production host: Homo sapiens (human) References: UniProt: Q7L523, Hydrolases; Acting on acid anhydrides; Acting on GTP to facilitate cellular and subcellular movement #3: Protein | Mass: 44298.859 Da / Num. of mol.: 2 / Mutation: S75N Source method: isolated from a genetically manipulated source Source: (gene. exp.) Homo sapiens (human) / Gene: RRAGC / Production host: Homo sapiens (human) / References: UniProt: Q9HB90 |
---|
-Ragulator complex protein ... , 5 types, 10 molecules DKELFMGNHO
#4: Protein | Mass: 17762.775 Da / Num. of mol.: 2 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Homo sapiens (human) / Gene: LAMTOR1, C11orf59, PDRO, PP7157 / Production host: Spodoptera frugiperda (fall armyworm) / References: UniProt: Q6IAA8 #5: Protein | Mass: 13517.450 Da / Num. of mol.: 2 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Homo sapiens (human) / Gene: LAMTOR2, MAPBPIP, ROBLD3, HSPC003 / Production host: Spodoptera frugiperda (fall armyworm) / References: UniProt: Q9Y2Q5 #6: Protein | Mass: 13637.678 Da / Num. of mol.: 2 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Homo sapiens (human) / Gene: LAMTOR3, MAP2K1IP1, MAPKSP1, PRO2783 / Production host: Spodoptera frugiperda (fall armyworm) / References: UniProt: Q9UHA4 #7: Protein | Mass: 10753.236 Da / Num. of mol.: 2 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Homo sapiens (human) / Gene: LAMTOR4, C7orf59 / Production host: Spodoptera frugiperda (fall armyworm) / References: UniProt: Q0VGL1 #8: Protein | Mass: 9622.900 Da / Num. of mol.: 2 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Homo sapiens (human) / Gene: LAMTOR5, HBXIP, XIP / Production host: Spodoptera frugiperda (fall armyworm) / References: UniProt: O43504 |
---|
-Non-polymers , 3 types, 6 molecules
#10: Chemical | #11: Chemical | #12: Chemical | |
---|
-Details
Has ligand of interest | N |
---|
-Experimental details
-Experiment
Experiment | Method: ELECTRON MICROSCOPY |
---|---|
EM experiment | Aggregation state: PARTICLE / 3D reconstruction method: single particle reconstruction |
-Sample preparation
Component | Name: The Raptor-TFEB-Rag-Ragulator complex / Type: COMPLEX / Entity ID: #1-#9 / Source: MULTIPLE SOURCES |
---|---|
Buffer solution | pH: 7.4 |
Specimen | Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES |
Specimen support | Grid material: COPPER / Grid mesh size: 300 divisions/in. / Grid type: C-flat-2/1 |
Vitrification | Cryogen name: ETHANE |
-Electron microscopy imaging
Experimental equipment | Model: Titan Krios / Image courtesy: FEI Company |
---|---|
Microscopy | Model: FEI TITAN KRIOS |
Electron gun | Electron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: FLOOD BEAM |
Electron lens | Mode: BRIGHT FIELD / Nominal defocus max: 2200 nm / Nominal defocus min: 800 nm |
Image recording | Electron dose: 50 e/Å2 / Film or detector model: GATAN K3 BIOQUANTUM (6k x 4k) |
-Processing
Software |
| ||||||||||||||||||||||||
---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|
EM software |
| ||||||||||||||||||||||||
CTF correction | Type: PHASE FLIPPING AND AMPLITUDE CORRECTION | ||||||||||||||||||||||||
3D reconstruction | Resolution: 2.9 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 377569 / Symmetry type: POINT | ||||||||||||||||||||||||
Atomic model building | Protocol: AB INITIO MODEL / Space: REAL | ||||||||||||||||||||||||
Refinement | Cross valid method: NONE Stereochemistry target values: GeoStd + Monomer Library + CDL v1.2 | ||||||||||||||||||||||||
Displacement parameters | Biso mean: 34.77 Å2 | ||||||||||||||||||||||||
Refine LS restraints |
|