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19 results for “autophagosome”
Tabular datasets for "In situ structural analysis reveals membrane shape transitions during autophagosome formation"
<p>Tabular source data for all plots in the manuscript "In situ structural analysis reveals membrane shape transitions during autophagosome formation". The article is available at https://doi.org/10.1101/2022.05.02.490291. The naming of the sheets in the .xlsx files corresponds to the figure number and panel.</p>
Faa1 membrane binding drives positive feedback in autophagosome biogenesis via fatty acid activation
<p>Autophagy serves as a stress response pathway by mediating the degradation of cellular material within lysosomes. In autophagy, this material is encapsulated in double-membrane vesicles termed autophagosomes, which form from precursors referred to as phagophores. Phagophores grow by lipid influx from the endoplasmic reticulum into Atg9-positive compartments and local lipid synthesis provides lipids for their expansion. How phagophore nucleation and expansion are coordinated with lipid synthesis is unclear. Here, we show that Faa1, an enzyme activating fatty acids, is recruited to Atg9 vesicles by directly binding to negatively charged membranes with a preference for phosphoinositides such as PI3P and PI4P. We define the membrane-binding surface of Faa1 and show that its direct interaction with the membrane is required for its recruitment to phagophores. Furthermore, the physiological localization of Faa1 is key for its efficient catalysis and promotes phagophore expansion. Our results suggest a positive feedback loop coupling phagophore nucleation and expansion to lipid synthesis.</p>
Autophagosomal/autolysosomal/lysosomal dynamics of FaDu and HGFb cells affected by autophagy modulators (dataset of dual labeling of autophagosomes and labeling lysosomes, confocal microscopy)
<p>In this dataset, the impact of autophagy modulators on the auophagosomes and lysosomes in FaDu and HGF cells was investigated. Experimental details are described in the accompanying paper. This dataset contains total 702 CZI confocal image Z-stacks.</p> <p><strong>Relevant paper</strong>: HANELOVA, Klara, RAUDENSKA, Martina, KRATOCHVILOVA, Monika, NAVRATIL, Jiri, VICAR, Tomas, BUGAJOVA, Maria, GUMULEC, Jaromir, MASARIK, Michal and BALVAN, Jan. Autophagy modulators influence the content of important signalling molecules in PS-positive extracellular vesicles. <em>Cell Communication and Signaling</em>. 24 May 2023. Vol. 21, no. 1, p. 120. DOI <a href="https://doi.org/10.1186/s12964-023-01126-z">10.1186/s12964-023-01126-z</a>.</p> <div> <div> <div> </div> </div> </div> <p><strong>Model cell lines</strong></p> <p> </p> <p>The cell line FaDu (HTB-43TM), derived from a squamous cell carcinoma (SCC) of the hypopharynx, and the human gingival fibroblast cell line HGF (derived from the histologically normal gingival biopsy) were used in this study. The authenticated cell lines were purchased from the American Type Culture Collection (ATCC; Manassas, Virginia, USA) within the last five years. </p> <p><strong>Autophagy modulation</strong></p> <p>For autophagy modulation, FaDu cells were treated for 24 h with 5nM bafilomycin A1 (Sigma-Aldrich, B1793), 50 µM of hydroxychloroquine sulphate (Sigma-Aldrich, H0915), 100 µM of Cpd18 (Calbiochem), 50 nM of autophinib (Sigma, SML2632), 10 µM of EACC (MedChemExpress), 200 nM of rapamycin (Sigma-Aldrich, R0395), 3 nM of Torin-1 (MedChemExpress), or 30 nM of NVP-BEZ235 (MedChemExpress). To induce starvation, cells were cultured in DMEM F12 without glutamine and without FBS (Biosera). Modulation of autophagy did not reduce the viability of FaDu cells.</p> <p><strong>Conditioned media preparation</strong></p> <p>see details in the accompanying paper</p> <p><strong>Fluorescence Microscopy</strong><br>The autophagosomal/autolysosomal/lysosomal dynamics of affected cells were observed using the combination of PremoTM Autophagy Tandem Sensor (P36239, Invitrogen) with the far-red emitting LysoTracker® Deep Red (L12492, Invitrogen) (Ex 647 nm/Em 668 nm). By combining acid-sensitive Emerald GFP (Ex 488 nm/Em 509 nm) with acid-insensitive TagRFP (Ex 555 nm/Em 584 nm) in the PremoTM kit, autophagosomes and autolysosomes labelling (yellow and red, respectively) is possible.<br>Immediately after transduction with 12 µl PremoTM Autophagy Tandem Sensor/2ml cell suspension, cells were seeded at 5 × 10<sup>5</sup> into 35-mm glass-bottomed gelatin-coated dishes (Ibidi, μ-Dish 35 mm, high Glass Bottom) and cultured for 48 h to equilibrate expression levels. Subsequently, cells were exposed to the selected agents for 6, 12, 24 and 48 hours before imaging. LysoTracker® Deep Red staining was performed 1 h before imaging. <br>To monitor the uptake of isolated PS-EVs by fibroblasts, we stained EVs with PKH67 (Sigma, PKH67GL) and then removed the remaining dye using Exosome Spin Columns (MW 3000) (Thermo Scientific, #4484449). The stained EVs were then suspended in 400 ul of cultivation medium and added to HGFB cells growing for 24 h in Ibidi µ-Slide I Luer (Ibidi, 80176). Image acquisition was performed 24 hours after EVs addition. 1 µl of 1 µg/ml of Hoechst 33342 (Enzo) (Ex 350 nm/ Em 461 nm) nuclear stain was added 1 hour before imaging.<br>To determine the viability of the cell population prior to isolation of EVs, cells were left in a culture dish with 1 ml of culture medium to which propidium iodide (Sigma-Aldrich) and Hoechst 33342 were added 45 minutes before capturing. <br>To maximize the possibility of comparison between samples, all samples (from a single cell line) were captured on the same day in a single run using the same microscope settings. For each time and each treatment, 10–12 fields of view were captured from randomized sites of the culture dish. Epifluorescent microscopy images and confocal microscopy images were acquired using Laser scanning confocal microscope Zeiss LSM 880 with AiryscanFast module (Carl Zeiss Inc.) using a C-apochromat 40x/1.20 W and C-Apochromat 63 /1.20 W. LysoTracker® Deep Red was excited HeNe 633 nm solid-state laser and emitted light was detected at 638–759 nm. Emerald GFP was excited 488 nm ArgonRemote laser, and emitted light was detected at 493–576 nm. TagRFP was excited DPSS 561 nm laser, and emitted light was detected at 570–650 nm. Hoechst 33342 was excited with a 405 nm solid-state laser, and emitted light was detected at 410–508 nm. Fluorescence images were acquired by the transmitted light detector. <br>Images were analyzed using ImageJ software and custom MATLAB software developed in our laboratory. The analysis process consists of the segmentation of cells from the background and extraction of the intensity of fluorescence channels (TagRFP, GFP, LysoTracker) inside cells. For segmentation, a thresholding-based method was used, where a manually selected threshold was applied. Segmentation was applied to the image created as the sum of all fluorescence channels to achieve segmentation independent of the intensity of individual channels. To achieve better segmentation without noisy pixels, fluorescence images were preprocessed with median filter (7x7) and Gaussian filter (standard deviation 1); additionally, binary segmentation was post-processed with morphological closing and removal of small binary connected components (<5000px). For intensity extraction, the mean value of segmented cell pixels was used for each field of view. Besides individual fluorescence channels, the mean colocalization of TagRFP and GFP was calculated with a pixel-wise multiplication of TagRFP and GFP channels.</p> <p><strong>File Naming</strong></p> <p>CZI images are organised in folders according to cell line, treatment time, and treatment. Names include magnification, cell line measured, treatment, and time of treatment,</p> <blockquote> <p>40x_FADU_BAF_12h_BF_4.czi</p> </blockquote> <p>The abbreviations used for treatments are as follows: BAF, bafilomycin A1; HCQ, hydroxychloroquine sulphate; Cpd18; APB, autophinib; EACC; RAPA, rapamycin; TOR1, Torin-1; BEZ, NVP-BEZ235; starv, starvation; </p>
Data files for Axonal transport of autophagosomes is regulated by dynein activators JIP3/JIP4 and ARF/RAB GTPases
<p>Supporting data files for <strong>Axonal transport of autophagosomes is regulated by dynein activators JIP3/JIP4 and ARF/RAB GTPases</strong></p>
Quantification data for article entitled "ATG9 vesicles comprise the seed membrane of mammalian autophagosomes"
<p>Membrane fractionation, immunoprecipitation, and <em>in vitro</em> experiment densitometric quantification datasheets for quantified data presented in the research article entitled "ATG9 vesicles comprise the seed membrane of mammalian autophagosomes". Membrane fractions (Figures 3, 4, S4, and S5) were from WT cells (starvation + bafiomycin A1 and basal conditions), ATG2 DKO cells, FIP200 KO cells, ATG9A KO cells, ATG3 KO cells, and ATG4 QKO cells (all KO cells under basal conditions). Immunoprecipitations (Figures 6 and S5) were from WT cells (starvation + bafiomycin A1) and ATG2 DKO cells (basal). Guanidine hydrochloride (GdnHCl) sequential immunoprecipitations (Figure 6) were from WT cells (starvation + bafiomycin A1). <em>In vitro</em> experiments (Figure S5) were using recombinant proteins on reconstituted liposomes.</p>
Study on the Correlation Between Alveolar Macrophage-derived Autophagosomes and the Severity of Lung Injury in ARDS
ClinicalTrials.gov study NCT05101694. IPD Sharing: YES. Countries: 1. Publications: 13.
Data for: Quantitative analysis of autophagy reveals the role of ATG9 and ATG2 in autophagosome formation
Open the record for dataset details and reuse information.
Identification and therapeutic rescue of autophagosome and glutamate receptor defects in C9ORF72 and sporadic ALS neurons
GEO Series GSE133162. Homo sapiens. 4 samples. Type: Expression profiling by high throughput sequencing.
Transcriptional regulation by Pho23 modulates the frequency of autophagosome formation.
GEO Series GSE57031. Saccharomyces cerevisiae. 4 samples. Type: Expression profiling by high throughput sequencing.
Mycobacterium tuberculosis Sensor Kinase DosS Modulates the Autophagosome in a DosR-independent Manner [II]
GEO Series GSE118867. Mycobacterium tuberculosis CDC1551; Mycobacterium tuberculosis; Mycobacterium tuberculosis H37Rv. 6 samples. Type: Expression profiling by array.
Lamp2-Flotillin-2 interaction enhances autophagosome-lysosome fusion to protect the septic heart in response to ILC2
GEO Series GSE280661. Mus musculus. 13 samples. Type: Expression profiling by high throughput sequencing.
Mycobacterium tuberculosis Sensor Kinase DosS Modulates the Autophagosome in a DosR-independent Manner [I]
GEO Series GSE118866. Homo sapiens; Macaca mulatta. 8 samples. Type: Expression profiling by array.
mTOR hyperactivation in Down Syndrome mediates deficits in autophagy induction, autophagosome formation, and mitophagy
GEO Series GSE126910. Homo sapiens. 12 samples. Type: Expression profiling by high throughput sequencing.
Phase Ib Trial of Multivalent Autophagosome Vaccine With or Without GITR Agonist, With Anti-PD-1 Immunotherapy in HNSCC
ClinicalTrials.gov study NCT04470024. IPD Sharing: NO. Countries: 1. Publications: 0.
Mycobacterium tuberculosis Sensor Kinase DosS Modulates the Autophagosome in a DosR-independent Manner
GEO Series GSE118869. Mycobacterium tuberculosis CDC1551; Macaca mulatta; Mycobacterium tuberculosis; Homo sapiens; Mycobacterium tuberculosis H37Rv. 14 samples. Type: Expression profiling by array.
RNA-seq analysis revealed that crizotinib-induced cardiomyocyte death, and cardiac dysfunctionapoptosis is related to autophagosome-lysosome fusion and mitochondrial injury
GEO Series GSE210150. Homo sapiens. 6 samples. Type: Expression profiling by high throughput sequencing.
USP4 depletion-driven RAB7A ubiquitylation impairs autophagosome-lysosome fusion and aggravates periodontitis
GEO Series GSE273165. Homo sapiens. 9 samples. Type: Expression profiling by high throughput sequencing.
AMPK activation in response to hepatitis E virus infection inhibited viral replication by attenuating autophagosome and promoting innate immunity
GEO Series GSE262469. Homo sapiens. 15 samples. Type: Expression profiling by high throughput sequencing.
VPS34 regulates RAB7 activation and Retromer localization to autophagosomes and mitophagy in oocytes
GEO Series GSE216465. Mus musculus. 18 samples. Type: Expression profiling by high throughput sequencing.
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Allen Brain Atlas
Allen Brain Atlas is an Allen Institute collection of brain map atlases, datasets, APIs, and analysis tools covering mouse, human, and non-human primate brain resources.
Annotated Behaviour and Observability Dataset (ABODe)
ABODe is a University of Edinburgh DataShare dataset for behavior classification in group-housed mice using home-cage video, identities, bounding boxes, ground-plate positions, and annotator labels.
DANDI Archive for NWB datasets
DANDI is a BRAIN Initiative archive for publishing and sharing neurophysiology data, including electrophysiology, optophysiology, and behavioral data packaged as NWB and related standards.
International Brain Laboratory public data
The International Brain Laboratory public data releases expose standardized mouse decision-making experiments, including Neuropixels recordings, widefield calcium imaging, behavior, and session metadata accessed through the ONE API.
OpenNeuro
OpenNeuro is a free, open platform for sharing neuroimaging datasets, with public search, dataset pages, and download paths for web, S3, DataLad, and the OpenNeuro CLI.