Lipid and protein content profiling of isolated native autophagic vesicles
Actions 
Order Assays
Autophagy is responsible for degradation of an extensive portfolio of cytosolic cargoes that are engulfed in autophagosomes to facilitate their transport to lysosomes. Besides basal autophagy, which constantly degrades cellular material, the pathway is dynamically altered by different conditions, resulting in enhanced autophagosome formation and cargo turnover. The extensive profile of autophagosome content as well as the phospholipid composition of human autophagosome membranes remains elusive. Here, we introduce a novel FACS-based approach for isolation of native autophagosomes and confirm the quality and specificity of the preparations. Employing quantitative proteomics and lipidomics, we obtained a profound cargo and lipid profile of autophagosomes isolated upon basal autophagy conditions, nutrient deprivation, and proteasome inhibition. This resource will allow to identify the impact of different autophagy conditions on the degradation of distinct proteins and to dissect the phospholipid composition of the human autophagosome membrane in detail.
Export PNG
Download
Creators and Submitter
Views: 50
Created: 9th Jul 2024 at 13:37
Last updated: 15th Oct 2024 at 12:22
TagsThis item has not yet been tagged.
Related items
Projects: Published Datasets, Unpublished Datasets
Institutions: DZNE
Please visit the 'Related items' tab within the profile page to explore associated studies in more detail.
Projects: Published Datasets, Unpublished Datasets
Institutions: LMU Klinikum
Research Data Steward
This project serves as a centralized repository for omics datasets published by research groups within the SyNergy Cluster. It encompasses investigations such as proteomics and transcriptomics, which are further divided into individual studies led by SyNergy members. Each study is linked to relevant publications, assays and data files (with links to external repositories).
To explore investigations and their associated studies in more detail, please visit the 'Related items' tab on the Project ...
Public web page: Not specified
Submitter: Rainer Malik
Studies: A TBK1 variant causes autophagolysosomal and motoneuron pathology withou..., A ubiquitin-specific, proximity-based labeling approach for the identifi..., ACSL3 is a novel GABARAPL2 interactor that links ufmylation and lipid dr..., ADAM10-Mediated Ectodomain Shedding Is an Essential Driver of Podocyte D..., ALS-linked loss of Cyclin-F function affects HSP90, AMPK, a Regulator of Metabolism and Autophagy, Is Activated by Lysosomal..., ATG4 family proteins drive phagophore growth independently of the LC3/GA..., An optimized quantitative proteomics method establishes the cell type-re..., Autophagosomal Content Profiling Reveals an LC3C-Dependent Piecemeal Mit..., Autophagosome content profiling using proximity biotinylation proteomics..., Autophagy acts through TRAF3 and RELB to regulate gene expression via an..., Basic Fibroblast Growth Factor 2-Induced Proteome Changes Endorse Lewy B..., Beneficial Effect of ACI-24 Vaccination on Aβ Plaque Pathology and Micro..., Brain injury environment critically influences the connectivity of trans..., C9orf72 protein quality control by UBR5-mediated heterotypic ubiquitin c..., CRISPR-Mediated Induction of Neuron-Enriched Mitochondrial Proteins Boos..., Cell-type-specific profiling of brain mitochondria reveals functional an..., Cellular depletion of major cathepsin proteases reveals their concerted ..., Deciphering sources of PET signals in the tumor microenvironment of glio..., Defining the Adult Neural Stem Cell Niche Proteome Identifies Key Regula..., Development of a Proteomic Workflow for the Identification of Heparan Su..., Distinct molecular profiles of skull bone marrow in health and neurologi..., Excessive local host-graft connectivity in aging and amyloid-loaded brain, Experimental evidence for temporal uncoupling of brain Aβ deposition and..., Fibrillar Aβ triggers microglial proteome alterations and dysfunction in..., Filling the Gaps – A Call for Comprehensive Analysis of Extracellular Ma..., IKKβ binds NLRP3 providing a shortcut to inflammasome activation for rap..., Injury-specific factors in the cerebrospinal fluid regulate astrocyte pl..., Lipid and protein content profiling of isolated native autophagic vesicles, Loss of CLN3 in microglia leads to impaired lipid metabolism and myelin ..., Loss of NPC1 enhances phagocytic uptake and impairs lipid trafficking in..., Lysosomal damage sensing and lysophagy initiation by SPG20-ITCH, Lysosomal targeting of the ABC transporter TAPL is determined by membran..., Mapping autophagosome contents identifies interleukin-7 receptor-alpha a..., Met/HGFR triggers detrimental reactive microglia in TBI, MicroRNAs from extracellular vesicles as a signature for Parkinson's dis..., Multi-omics profiling identifies a deregulated FUS-MAP1B axis in ALS/FTD..., Multiomic ALS signatures highlight subclusters and sex differences sugge..., Neuronal differentiation of LUHMES cells induces substantial changes of ..., Nonvesicular lipid transfer drives myelin growth in the central nervous ..., NrCAM is a marker for substrate-selective activation of ADAM10 in Alzhei..., Pro-inflammatory activation following demyelination is required for myel..., Proteomic Characterization of Ubiquitin Carboxyl-Terminal Hydrolase 19 D..., Proteomic and lipidomic profiling of demyelinating lesions identifies fa..., Proteomic profiling in cerebral amyloid angiopathy reveals an overlap wi..., Proteomics of mouse brain endothelium uncovers dysregulation of vesicula..., Rational correction of pathogenic conformational defects in HTRA1, Reactivated endogenous retroviruses promote protein aggregate spreading, Signal peptide peptidase-like 2c impairs vesicular transport and cleaves..., Signatures of glial activity can be detected in the CSF proteome, Spatial centrosome proteome of human neural cells uncovers disease-relev..., Spatial proteomics in three-dimensional intact specimens, Spatial proteomics reveals secretory pathway disturbances caused by neur..., Systematically defining selective autophagy receptor-specific cargo usin..., Targeting the TCA cycle can ameliorate widespread axonal energy deficien..., The Alzheimer's disease-linked protease BACE1 modulates neuronal IL-6 si..., The Alzheimer's disease-linked protease BACE2 cleaves VEGFR3 and modulat..., The COP9 signalosome reduces neuroinflammation and attenuates ischemic n..., The Hippo network kinase STK38 contributes to protein homeostasis by inh..., The intramembrane protease SPPL2c promotes male germ cell development by..., The pseudoprotease iRhom1 controls ectodomain shedding of membrane prote..., The tetraspanin Tspan15 is an essential subunit of an ADAM10 scissor com..., The ubiquitin-conjugating enzyme UBE2QL1 coordinates lysophagy in respon..., Trnp1 organizes diverse nuclear membrane-less compartments in neural ste..., Ubiquitin profiling of lysophagy identifies actin stabilizer CNN2 as a t...
Assays: Affinity purification coupled with mass spectrometry proteomics (human), Affinity purification coupled with mass spectrometry proteomics (human), Affinity purification coupled with mass spectrometry proteomics (human), Affinity purification coupled with mass spectrometry proteomics (human), Affinity purification coupled with mass spectrometry proteomics (human), Affinity purification coupled with mass spectrometry proteomics (human), Affinity purification coupled with mass spectrometry proteomics (human), Affinity purification coupled with mass spectrometry proteomics (human), Affinity purification coupled with mass spectrometry proteomics (human), Bottom-up proteomics (human), Bottom-up proteomics (mouse), Bottom-up proteomics (mouse), Bottom-up proteomics (mouse), Bottom-up proteomics (mouse), Gel-based experiment (human), Phosphoproteomics / Bottom-up proteomics (mouse), Proximity-proteomics-based autophagosome content profiling (human), SWATH MS (human), SWATH MS (human, mouse), SWATH MS (mouse), Shotgun proteomics (human), Shotgun proteomics (human), Shotgun proteomics (human), Shotgun proteomics (human), Shotgun proteomics (human), Shotgun proteomics (human), Shotgun proteomics (human), Shotgun proteomics (human), Shotgun proteomics (human), Shotgun proteomics (human), Shotgun proteomics (human), Shotgun proteomics (human), Shotgun proteomics (human), Shotgun proteomics (human), Shotgun proteomics (human), Shotgun proteomics (human), Shotgun proteomics (human, mouse), Shotgun proteomics (human, mouse), Shotgun proteomics (human, mouse), Shotgun proteomics (human, mouse), Shotgun proteomics (macaque), Shotgun proteomics (mouse), Shotgun proteomics (mouse), Shotgun proteomics (mouse), Shotgun proteomics (mouse), Shotgun proteomics (mouse), Shotgun proteomics (mouse), Shotgun proteomics (mouse), Shotgun proteomics (mouse), Shotgun proteomics (mouse), Shotgun proteomics (mouse), Shotgun proteomics (mouse), Shotgun proteomics (mouse), Shotgun proteomics (mouse), Shotgun proteomics (mouse), Shotgun proteomics (mouse), Shotgun proteomics (mouse), Shotgun proteomics (mouse), Shotgun proteomics (mouse), Shotgun proteomics (mouse), Shotgun proteomics (mouse), Shotgun proteomics (mouse), Shotgun proteomics (mouse), Shotgun proteomics (mouse), Shotgun proteomics (mouse), Shotgun proteomics (mouse), Shotgun proteomics (rat), Untargeted Proteomics (mouse)
Sample preparation for LC-MS/MS Eight million of Cy3/5-positive autophagosomes (roughly 10ug of proteins) were denatured with 2% sodium deoxycholate, 50 mM Tris-HCl pH 8.5, 2.5 mM TCEP, 10 mM chloroacetamide at 95°C for 10 min. Lysates were prepared with in-StageTip (iST) processing method for LC-MS/MS as previously described by Kulak et al. 2014. Briefly, Proteins were digested overnight at 37°C with 1 volume of 50mM Tris-HCl pH 8.5 containing LysC (Wako Chemicals) at 1:100 (w/w) ratio and Trypsin ...
Submitter: Rainer Malik
Assay type: Proteomics
Technology type: Technology Type
Investigation: Proteomics (Published)
Autophagy is responsible for degradation of an extensive portfolio of cytosolic cargoes that are engulfed in autophagosomes to facilitate their transport to lysosomes. Besides basal autophagy, which constantly degrades cellular material, the pathway is dynamically altered by different conditions, resulting in enhanced autophagosome formation and cargo turnover. The extensive profile of autophagosome content as well as the phospholipid composition of human autophagosome membranes remains elusive. ...
Creators: None
Submitter: Rainer Malik
External Link
Abstract (Expand)
Authors: D. Schmitt, S. Bozkurt, P. Henning-Domres, H. Huesmann, S. Eimer, L. Bindila, C. Behrends, E. Boyle, F. Wilfling, G. Tascher, C. Munch, C. Behl, A. Kern
Date Published: 6th Dec 2022
Publication Type: Journal
