|Entry||Database: PDB / ID: 6tgy|
|Title||Cryo-EM structure of p62-PB1 filament (L-type)|
|Keywords||APOPTOSIS / Autophagy|
|Function / homology|
Function and homology information
protein localization to perinuclear region of cytoplasm / selective autophagy / regulation of Ras protein signal transduction / Lewy body / amphisome / response to mitochondrial depolarisation / aggrephagy / regulation of protein complex stability / mitophagy / endosome organization ...protein localization to perinuclear region of cytoplasm / selective autophagy / regulation of Ras protein signal transduction / Lewy body / amphisome / response to mitochondrial depolarisation / aggrephagy / regulation of protein complex stability / mitophagy / endosome organization / phagophore assembly site / K63-linked polyubiquitin modification-dependent protein binding / regulation of mitochondrion organization / autolysosome / immune system process / sperm midpiece / aggresome / regulation of I-kappaB kinase/NF-kappaB signaling / autophagy of mitochondrion / ionotropic glutamate receptor binding / autophagosome / inclusion body / sarcomere / negative regulation of protein ubiquitination / mitochondrion organization / response to ischemia / SH2 domain binding / ubiquitin binding / protein kinase C binding / protein localization / P-body / endosomal transport / positive regulation of long-term synaptic potentiation / positive regulation of protein localization to plasma membrane / macroautophagy / receptor tyrosine kinase binding / PML body / interleukin-1-mediated signaling pathway / autophagy / late endosome / ubiquitin-dependent protein catabolic process / cell differentiation / intracellular signal transduction / positive regulation of protein phosphorylation / positive regulation of apoptotic process / intracellular membrane-bounded organelle / apoptotic process / protein-containing complex binding / protein serine/threonine kinase activity / ubiquitin protein ligase binding / protein kinase binding / negative regulation of apoptotic process / endoplasmic reticulum / negative regulation of transcription by RNA polymerase II / enzyme binding / positive regulation of transcription by RNA polymerase II / mitochondrion / extracellular exosome / zinc ion binding / nucleoplasm / identical protein binding / cytosol / cytoplasm
Ubiquitin-associated domain / PB1 domain / Zinc finger, ZZ-type / PB1 domain / Zinc finger, ZZ-type superfamily / Sequestosome-1, PB1 domain / Sequestosome-1, UBA domain / UBA-like superfamily / Phosphatidylinositol 3-kinase Catalytic Subunit; Chain A, domain 1 / Ubiquitin-like (UB roll) ...Ubiquitin-associated domain / PB1 domain / Zinc finger, ZZ-type / PB1 domain / Zinc finger, ZZ-type superfamily / Sequestosome-1, PB1 domain / Sequestosome-1, UBA domain / UBA-like superfamily / Phosphatidylinositol 3-kinase Catalytic Subunit; Chain A, domain 1 / Ubiquitin-like (UB roll) / Roll / Alpha Beta
|Biological species||Homo sapiens (human)|
|Method||ELECTRON MICROSCOPY / helical reconstruction / cryo EM / Resolution: 3.5 Å|
|Authors||Jakobi, A.J. / Huber, S.T. / Mortensen, S.A. / Sachse, C.|
|Funding support|| Germany, 3items |
|Citation||Journal: Nat Commun / Year: 2020|
Title: Structural basis of p62/SQSTM1 helical filaments and their role in cellular cargo uptake.
Authors: Arjen J Jakobi / Stefan T Huber / Simon A Mortensen / Sebastian W Schultz / Anthimi Palara / Tanja Kuhm / Birendra Kumar Shrestha / Trond Lamark / Wim J H Hagen / Matthias Wilmanns / Terje ...Authors: Arjen J Jakobi / Stefan T Huber / Simon A Mortensen / Sebastian W Schultz / Anthimi Palara / Tanja Kuhm / Birendra Kumar Shrestha / Trond Lamark / Wim J H Hagen / Matthias Wilmanns / Terje Johansen / Andreas Brech / Carsten Sachse /
Abstract: p62/SQSTM1 is an autophagy receptor and signaling adaptor with an N-terminal PB1 domain that forms the scaffold of phase-separated p62 bodies in the cell. The molecular determinants that govern PB1 ...p62/SQSTM1 is an autophagy receptor and signaling adaptor with an N-terminal PB1 domain that forms the scaffold of phase-separated p62 bodies in the cell. The molecular determinants that govern PB1 domain filament formation in vitro remain to be determined and the role of p62 filaments inside the cell is currently unclear. We here determine four high-resolution cryo-EM structures of different human and Arabidopsis PB1 domain assemblies and observed a filamentous ultrastructure of p62/SQSTM1 bodies using correlative cellular EM. We show that oligomerization or polymerization, driven by a double arginine finger in the PB1 domain, is a general requirement for lysosomal targeting of p62. Furthermore, the filamentous assembly state of p62 is required for autophagosomal processing of the p62-specific cargo KEAP1. Our results show that using such mechanisms, p62 filaments can be critical for cargo uptake in autophagy and are an integral part of phase-separated p62 bodies.
SummaryFull reportAbout validation report
|Structure viewer||Molecule: |
Downloads & links
A: Sequestosome-1x 120
|#1: Protein|| |
Mass: 13704.609 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: SQSTM1, ORCA, OSIL / Production host: Escherichia coli BL21(DE3) (bacteria) / References: UniProt: Q13501
|Experiment||Method: ELECTRON MICROSCOPY|
|EM experiment||Aggregation state: FILAMENT / 3D reconstruction method: helical reconstruction|
|Component||Name: p62-PB1 domain filament (L-type) / Type: COMPLEX / Entity ID: #1 / Source: RECOMBINANT|
|Molecular weight||Experimental value: NO|
|Source (natural)||Organism: Homo sapiens (human)|
|Source (recombinant)||Organism: Escherichia coli BL21(DE3) (bacteria)|
|Buffer solution||pH: 7.5 / Details: 50 mM TRIS (pH 7.5), 100 mM NaCl, 4 mM DTT|
|Specimen||Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES|
|Specimen support||Grid material: COPPER / Grid mesh size: 400 divisions/in. / Grid type: Quantifoil R2/1|
|Vitrification||Instrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE / Humidity: 100 % / Chamber temperature: 283 K|
-Electron microscopy imaging
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 FIELDBright-field microscopy / Nominal defocus max: 2500 nm / Nominal defocus min: 500 nm|
|Specimen holder||Cryogen: NITROGEN / Model: FEI TITAN KRIOS AUTOGRID HOLDER|
|Image recording||Electron dose: 40 e/Å2 / Detector mode: COUNTING / Film or detector model: GATAN K2 SUMMIT (4k x 4k) / Num. of real images: 2277|
|Image scans||Movie frames/image: 40|
|CTF correction||Type: PHASE FLIPPING AND AMPLITUDE CORRECTION|
|Helical symmerty||Angular rotation/subunit: 77.29 ° / Axial rise/subunit: 4.787 Å / Axial symmetry: C2|
|3D reconstruction||Resolution: 3.5 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 51853 / Symmetry type: HELICAL|
|Atomic model building||Protocol: FLEXIBLE FIT / Space: REAL / Target criteria: Correlation coefficient|
|Atomic model building||PDB-ID: 2KKC|
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