Browse
Submit Data
Databases
API
Help

Dataset Information

0 Views

0 Connections

0 Citations

0 Reanalyses

0 Downloads

Omics score: 0

Global organelle profiling reveals subcellular localization and remodeling at proteome scale

ABSTRACT: Defining the subcellular distribution of all human proteins and its remodeling across cellular states remains a central goal in cell biology. Here, we present a high-resolution strategy to map subcellular organization using organelle immuno-capture coupled to mass spectrometry. We apply this proteomics workflow to a cell-wide collection of membranous and membrane-less compartments. A graph-based representation of our data reveals the subcellular localization of over 7,600 proteins, defines spatial protein networks, and uncovers interconnections between cellular compartments. We demonstrate that our approach can be deployed to comprehensively profile proteome remodeling during cellular perturbation. By characterizing the cellular landscape following hCoV-OC43 viral infection, we discover that many proteins are regulated by changes in their spatial distribution rather than by changes in their total abundance. Our results establish that proteome-wide analysis of subcellular remodeling provides essential insights for the elucidation of cellular responses. Our dataset can be explored at organelles.czbiohub.org.

INSTRUMENT(S): Orbitrap Fusion Lumos

ORGANISM(S): Human Coronavirus Oc43 Homo Sapiens (human)

SUBMITTER: Frank McCarthy

LAB HEAD: Manuel Leonetti

PROVIDER: PXD046440 | Pride | 2023-12-19

REPOSITORIES: Pride

ACCESS DATA

Json Xml

Similar Datasets

High-resolution spatial proteome map of Toxoplasma gondii extracellular tachyzoite

Project description:Apicomplexans, such as malaria-causing Plasmodium species, Toxoplasma, and Cryptosporidium, are parasitic protists that invade cells of virtually every animal, including humans. Despite the great burden on human healthcare and global economy, and the role in the environment, our understanding of the biology of these microorganisms is still very limited. Apicomplexans have evolved a complex cell architecture featuring a range of subcellular compartments, both common to all eukaryotes and unique to the phylum, that enable invasion into the host cell and exploiting its resources for growth and replication, rendering apicomplexans such remarkably successful parasites. In such an intricate subcellular landscape, protein function and location in the cell are tightly linked, which is why localising proteins in the cell is a major strategy in the apicomplexan research. Yet, the majority of proteins in model apicomplexans is unknown. In this project, a state-of-the-art spatial proteomics technology hyperLOPIT has been successfully adapted and applied to a model apicomplexan Toxoplasma gondii. Three independent hyperLOPIT experiments have been conducted on T. gondii strain RH extracellular tachyzoites, the invasive stage of the parasite's life cycle responsible for acute infection, providing the first cell-wide protein location atlas of any apicomplexan. This provides revolutionary insight into the apicomplexan cell organization and biology, including greatly expanded proteomes of secretory invasion organelles, the range of adaptive and regulatory responses of the organelles, and the subcellular distribution of evolutionary selection pressure, innovation, and conservation.

2020-10-14 | PXD015269 | Pride

Galectin-3 interacts with components of the nuclear ribonucleoprotein complex

Project description:The multifunctional -galactoside-binding protein galectin-3 is found in many distinct subcellular compartments including the cell nucleus. Expression and distribution of galectin-3 between the cell nucleus and the cytosol changes during cell differentiation and cancer development. Nuclear functions of galectin-3 and how they contribute to tumorigenesis are not understood. In order to identify nuclear galectin-3 interaction partners, we used affinity chromatography and co-immunoprecipitation. Spatial proximity in the nucleus was assessed by immunofluorescence and proximity ligation assay. We also investigated the function of galectin-3 on mRNA-export by fluorescence in situ hybridization and on mRNA-processing by RNA-sequencing.

2016-06-07 | E-MTAB-4783 | biostudies-arrayexpress

Small RNA deep sequencing of cellular subcompartments

Project description:RNA deep sequencing efforts have revealed abundant expression of small RNAs derived from small nucleolar (sno) RNAs. We employ here spatial RNA deep sequencing to assess the expression of 10-40 nt small RNAs in subcellular compartments of HeLa cells. Total cellular, cytoplasmic, nuclear and nucleolar fractions were isolated, RNA was purified and size-fractionated in a two-step process to yield 10-40 nt RNA fractions. cDNA libraries were constructed and sequenced on a Ion Torrent platform. Vast majority of cellular, cytoplasmic and nuclear small RNA reads were identified as miRNAs. We found the expression of eleven ten miRNAs in the nucleolar preparations using a cut-off rate of 10 reads. Several miRNAs had a greater relative abundance in the nucleolus compared to the other compartments. The nucleolar small RNAs had a unique size distribution consisting of 19-20 and 25 nt RNAs, which were predominantly composed of small snoRNA-derived box C/D RNAs (termed as sdRNA). Sequences from 47 sdRNAs were identified, which mapped both 5M-bM-^@M-^Y and 3M-bM-^@M-^Y ends of the snoRNAs, and retained conserved box C or D motifs. SdRNA reads from SNORD44 comprised 74% of all nucleolar sdRNAs, and were confirmed by Northern blotting as 20 and 25 nt RNAs. This study reveals a rich representation of cell-compartment specific expression of small RNAs and the distinctive unique composition of the nucleolar small RNAs. Examination of small RNAs in cellular subcompartments

2014-05-07 | E-GEOD-50057 | biostudies-arrayexpress

Characterisation of PRR11 - Global subcellular characterization of protein degradation using quantitative proteomics

Project description:Protein degradation provides an important regulatory mechanism used to control cell cycle progression and many other cellular pathways. To comprehensively analyze the spatial control of protein degradation in U2OS osteosarcoma cells, we have combined drug treatment and SILAC-based quantitative mass spectrometry with subcellular and protein fractionation. The resulting data set analyzed more than 74,000 peptides, corresponding to ~5000 proteins, from nuclear, cytosolic, membrane, and cytoskeletal compartments. These data identified rapidly degraded proteasome targets, such as PRR11 and highlighted a feedback mechanism resulting in translation inhibition, induced by blocking the proteasome. We show this is mediated by activation of the unfolded protein response. We observed compartment-specific differences in protein degradation, including proteins that would not have been characterized as rapidly degraded through analysis of whole cell lysates. Bioinformatic analysis of the entire data set is presented in the Encyclopedia of Proteome Dynamics, a web-based resource, with proteins annotated for stability and subcellular distribution.

2013-07-23 | PXD000262 | Pride

Global Subcellular Characterization of Protein Degradation Using Quantitative Proteomics

Project description:Protein degradation provides an important regulatory mechanism used to control cell cycle progression and many other cellular pathways. To comprehensively analyze the spatial control of protein degradation in U2OS osteosarcoma cells, we have combined drug treatment and SILAC-based quantitative mass spectrometry with subcellular and protein fractionation. The resulting data set analyzed more than 74,000 peptides, corresponding to ∼5000 proteins, from nuclear, cytosolic, membrane, and cytoskeletal compartments. These data identified rapidly degraded proteasome targets, such as PRR11 and highlighted a feedback mechanism resulting in translation inhibition, induced by blocking the proteasome. We show this is mediated by activation of the unfolded protein response. We observed compartment-specific differences in protein degradation, including proteins that would not have been characterized as rapidly degraded through analysis of whole cell lysates. Bioinformatic analysis of the entire data set is presented in the Encyclopedia of Proteome Dynamics, a web-based resource, with proteins annotated for stability and subcellular distribution.

2015-11-27 | PXD003239 | Pride

Metabolic incorporation of electron-rich ribonucleosides enhances APEX-seq for profiling spatially restricted nascent transcriptome

Project description:We present MERR APEX-seq, a method for newly transcribed RNAs subcellular profiling combined metabolic incorporation of electron-rich ribonucleosides, 6-thioguanosine and 4-thiouridine, with the peroxidase-mediated RNA labeling method, APEX-seq. MERR APEX-seq offers both high spatial specificity and high coverage in the mitochondrial matrix and at the endoplasmic reticulum membrane. Application of MERR APEX-seq at nuclear lamina of human cells reveals that the mRNA components tend to encode for transcripts processing related proteins. MERR APEX-seq with high spatial specificity and high coverage could be widely used to expand our knowledge of RNA localization and function at subcellular compartments.

2022-01-03 | GSE192739 | GEO

Subcellular antibiotic visualization reveals a dynamic drug reservoir in infected macrophages

Project description:Tuberculosis, caused by the intracellular pathogen Mycobacterium tuberculosis, remains the world's deadliest infectious disease. Sterilizing chemotherapy requires at least 6 months of multidrug therapy. Difficulty visualizing the subcellular localization of antibiotics in infected host cells means that it is unclear whether antibiotics penetrate all mycobacteria-containing compartments in the cell. Here, we combined correlated light, electron, and ion microscopy to image the distribution of bedaquiline in infected human macrophages at submicrometer resolution. Bedaquiline accumulated primarily in host cell lipid droplets, but heterogeneously in mycobacteria within a variety of intracellular compartments. Furthermore, lipid droplets did not sequester antibiotic but constituted a transferable reservoir that enhanced antibacterial efficacy. Thus, strong lipid binding facilitated drug trafficking by host organelles to an intracellular target during antimicrobial treatment.

2020-12-07 | MTBLS959 | MetaboLights

Cross-link assisted spatial proteomics to map sub-organelle proteomes and membrane protein topology

Project description:The specific functions of cellular organelles and sub-compartments depend on their protein content, which can be characterized by spatial proteomics approaches. However, many spatial proteomics methods are limited in their ability to resolve organellar sub-compartments, profile multiple sub-compartments in parallel, and/or characterize membrane-associated proteomes. Here, we develop a cross-linking assisted spatial proteomics (CLASP) strategy that addresses these shortcomings. Using human mitochondria as a model system, we show that CLASP can elucidate spatial proteomes of all mitochondrial sub-compartments and provide topological insight into the mitochondrial membrane proteome in a single experiment. Biochemical and imaging-based follow-up studies demonstrate that CLASP allows discovering mitochondria-associated proteins and revising previous protein sub-compartment localization and membrane topology data. This study extends the scope of cross-linking mass spectrometry beyond protein structure and interaction analysis towards spatial proteomics, establishes a method for concomitant profiling of sub-organelle and membrane proteomes, and provides a resource for mitochondrial spatial biology

2024-02-21 | PXD046382 | Pride

Ribosome footprinting in the cytosol and endoplasmic reticulum

Project description:In eukaryotic cells, the spatial regulation of protein expression is frequently conferred through the coupling of mRNA localization and the local control of translation. mRNA localization to the endoplasmic reticulum (ER) is a prominent example of such regulation and serves a ubiquitous role in segregating the synthesis of secretory and integral membrane proteins to the ER. Recent genomic and biochemical studies have now expanded this view to suggest a role for the ER in global protein synthesis. We have utilized cell fractionation and ribosome profiling to obtain a genomic survey of the subcellular organization of mRNA translation and report that ribosomal loading of mRNAs, a proxy for mRNA translation, is biased to the ER. Notably, ER-associated mRNAs encoding both cytosolic and topogenic signal-encoding proteins display similar ribosome loading densities, suggesting that ER-associated ribosomes serve a global role in mRNA translation. We propose that the partitioning of mRNAs and their translation between the cytosol and ER compartments may represent a novel mechanism for the post-transcriptional regulation of gene expression. HEK293 cells were fractionated between the cytosol and endoplasmic reticulum. Within each fraction, ribosome footprints were generated and sequenced. In parallel, total mRNA was sequenced.

2011-12-20 | E-GEOD-31539 | biostudies-arrayexpress