Browse
Submit Data
Databases
API
Help

Dataset Information

0 Views

0 Connections

0 Citations

0 Reanalyses

0 Downloads

Omics score: 0

Organelle-specific RNA Tagging Probes for RNA Binding Proteome

ABSTRACT: The potential of our enrichment method for high-throughput RBP mapping with organelle-level spatial resolution.

ORGANISM(S): Homo Sapiens

SUBMITTER: Weijie Qin

PROVIDER: PXD033927 | iProX | Wed Oct 18 00:00:00 BST 2023

REPOSITORIES: iProX

ACCESS DATA

Json Xml

Publications

Nuclear and cytoplasmic specific RNA binding proteome enrichment and its changes upon ferroptosis induction.

Sun Haofan H Fu Bin B Qian Xiaohong X Xu Ping P Qin Weijie W

Nature communications 20240129 1

The key role of RNA-binding proteins (RBPs) in posttranscriptional regulation of gene expression is intimately tied to their subcellular localization. Here, we show a subcellular-specific RNA labeling method for efficient enrichment and deep profiling of nuclear and cytoplasmic RBPs. A total of 1221 nuclear RBPs and 1333 cytoplasmic RBPs were enriched and identified using nuclear/cytoplasm targeting enrichment probes, representing an increase of 54.4% and 85.7% compared with previous reports. Th ...[more]

PMID: 38286993

Similar Datasets

Organelle-specific RNA Tagging Probes for Spatially Resolved Profiling of RNA Binding Proteome and Investigation upon Ferroptosis Induction

Project description:The key role of RNA-binding proteins (RBPs) in posttranscriptional regulation of gene expression is intimately tied to their subcellular localization. Dysregulated localization may severely disrupt the biological functions of RBPs. To reveal the regulatory mechanisms of RBPs, methods for systematically mapping the subcellular localized RBPs and monitoring their translocations under physiological conditions are in high demand. Herein, a subcellular-specific RNA labeling method was developed for efficient enrichment and deep profiling of nuclear and cytoplasmic RBPs. A total of 1221 nuclear RBPs and 1333 cytoplasmic RBPs were enriched and identified using nuclear/cytoplasm targeting enrichment probes, which represented an increase of 54.4% and 85.7% compared with previous reports. The probes were further applied in the first omics-level investigation of subcellular-specific RBP-RNA interactions upon ferroptosis induction. Interestingly, large-scale RBPs displayed enhanced interaction with RNAs in nucleus but reduced association with RNAs in cytoplasm during ferroptosis process. Among these RBPs with regulated RNA-binding, translation was found as one of the commonly enriched GO functions by different ferroptosis inducers, indicating ferroptosis could disturb protein translation via different pathways. Furthermore, we discovered dozens of nucleoplasmic translocation candidate RBPs upon ferroptosis induction and validated representative ones by immunofluorescence imaging. The enrichment of TCA cycle in the translocation candidate RBPs may provide new insights for investigating their possible roles in ferroptosis induced metabolism dysregulation. The above findings suggest the potential of our enrichment method for high-throughput RBP mapping with organelle-level spatial resolution.

2022-12-01 | GSE205553 | GEO

Organelle-specific RNA Tagging Probes for Spatially Resolved Profiling of RNA Binding Proteome and Investigation upon Ferroptosis Induction

Project description:Organelle-specific RNA Tagging Probes for Spatially Resolved Profiling of RNA Binding Proteome and Investigation upon Ferroptosis Induction

| PRJNA846316 | ENA

Localisation of Organelle Proteins using Data-Independent Acquisition (DIA-LOP)

Project description:Subcellular localisation within the proteome fundamentally influences cellular processes, however the development of high-throughput techniques to allow proteome-wide mapping of the cell has proven difficult. Here we present DIA-LOP, an approach capable of high-throughput spatial proteome mapping with in-depth subcellular resolution. This unified framework integrates differential-ultracentrifugation with ion-mobility-based data independent acquisition (DIA) mass spectrometry, alongside data processing using DIA-NN and spatial analysis within the pRoloc bioinformatics pipeline. Within the same experimental pipeline, we compare Differential-ultraCentrifugation (DC) fractionation with an alternate detergent-based protocol using both data-dependent (DDA) and data-independent acquisition (DIA) mass spectrometry approaches highlighting the increased subcellular resolution of the DC approach and the increased proteome coverage when DIA is applied. This study thus informs potential users of subcellular proteomics strategies that employ biochemical fractionation of the optimal workflows to achieve high proteome coverage and subcellular resolution.

2025-09-29 | PXD063082 | Pride

Functional multi-organelle units control inflammatory lipid metabolism of macrophages

Project description:Eukaryotic cells contain several membrane-separated organelles to compartmentalize distinct metabolic reactions. However, it has remained unclear how these organelle systems are coordinated, when cells adapt metabolic pathways to support their development, survival or effector functions. Here we present OrgaPlexing, a multispectral organelle imaging approach for the comprehensive mapping of six key metabolic organelles and their interactions. We use this analysis on macrophages, immune cells that undergo rapid metabolic switches upon sensing bacterial and inflammatory stimuli. Our results identify lipid droplets (LDs) as primary inflammatory responder organelle, which forms three- and four-way interactions with other organelles. While clusters with endoplasmic reticulum (ER) and mitochondria (M-ER-LD unit) help supply fatty acids for LD growth, the additional recruitment of peroxisomes (M-ER-P-LD unit) supports fatty acid efflux from LDs. Interference with individual components of these units has direct functional consequences for inflammatory lipid synthesis. Together, we show that macrophages form functional multi-organellar units (MOUs) to support metabolic adaptation, and provide an experimental strategy to identify organelle-metabolic signaling hubs.

2024-05-03 | GSE235484 | GEO

CoLoC-seq, a high-throughput approach to profile organelle transcriptomes

Project description:CoLoC-seq, a high-throughput approach to profile organelle transcriptomes

| PRJNA759431 | ENA

Photoelectrocyclization as an activation mechanism for organelle-specific live-cell imaging probes.

Project description:Photoactivatable fluorophores are useful tools in live-cell imaging owing to their potential for precise spatial and temporal control. In this report, a new photoactivatable organelle-specific live-cell imaging probe based on a 6π electrocyclization/oxidation mechanism is described. It is shown that this new probe is water-soluble, non-cytotoxic, cell-permeable, and useful for mitochondrial imaging. The probe displays large Stokes shifts in both pre-activated and activated forms, allowing simultaneous use with common dyes and fluorescent proteins. Sequential single-cell activation experiments in dense cellular environments demonstrate high spatial precision and utility in single- or multi-cell labeling experiments.

| S-EPMC4485560 | biostudies-literature

Live-cell mapping of organelle-associated RNAs via proximity biotinylation combined with protein-RNA crosslinking

Project description:The spatial organization of RNA within cells is a crucial factor influencing a wide range of biological functions throughout all kingdoms of life. However, a general understanding of RNA localization has been hindered by a lack of simple, high-throughput methods for mapping the transcriptomes of subcellular compartments. Here, we develop such a method, termed APEX-RIP, which combines peroxidase-catalyzed, spatially restricted in situ protein biotinylation with RNA-protein chemical crosslinking. We demonstrate that, using a single protocol, APEX-RIP can isolate RNAs from a variety of subcellular compartments, including the mitochondrial matrix, nucleus, bulk cytosol, and endoplasmic reticulum (ER), with specificity and sensitivity that rival or exceed those of conventional approaches. We further identify candidate RNAs localized to mitochondria-ER junctions and nuclear lamina, two compartments that are recalcitrant to classical biochemical purification. Since APEX-RIP is simple, versatile, and does not require special instrumentation, we envision its broad application in a variety of biological contexts.

2017-12-14 | GSE106493 | GEO

Global organelle profiling reveals subcellular localization and remodeling at proteome scale

Project description: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 workflow to a cell-wide collection of membranous and membrane-less compartments. A graph-based analysis reveals the subcellular localization of over 7,600 proteins, defines spatial networks, and uncovers interconnections between cellular compartments. 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 abundance. Our results establish that proteome-wide analysis of subcellular remodeling provides unique insights for the elucidation of cellular responses, uncovering an essential role for ferroptosis in OC43 infection. Our dataset can be explored at organelles.czbiohub.org.

2024-10-04 | GSE278059 | GEO

A cooperative organelle biogenesis program in mammalian cells

Project description:The compartmentalisation of distinct organelles within eukaryotic cells is essential for their diverse functions, however, how their structures and functions depend on each other has not been systematically explored. We combined a fluorescent reporter of mitochondrial stress with genome-wide CRISPR knockout screening and identified networks of genes involved in the biogenesis and metabolism of diverse organelles. Targeted organelle gene knockouts identified that defects in peroxisomes, Golgi, and ER cause mitochondrial fragmentation and dysfunction. Correlative light and electron microscopy analysed using artificial intelligence-directed voxel extraction revealed in unprecedented detail how impaired mitochondrial interactions with diverse organelles caused cell-wide defects in their morphology and biogenesis. Multi-omics analyses identified a unified proteome stress response and global shifts in lipid and glycoprotein homeostasis that are elicited when organelle biogenesis is compromised. Our comprehensive resource has defined metabolic and morphological interactions between organelles that can be mined to understand how changes in organelle components drive diverse cellular pathologies.

2023-10-13 | GSE201573 | GEO

EC-tagging allows cell type-specific RNA analysis.

Project description:Purification of cell type-specific RNAs remains a significant challenge. One solution involves biosynthetic tagging of target RNAs. RNA tagging via incorporation of 4-thiouracil (TU) in cells expressing transgenic uracil phosphoribosyltransferase (UPRT), a method known as TU-tagging, has been used in multiple systems but can have limited specificity due to endogenous pathways of TU incorporation. Here, we describe an alternative method that requires the activity of two enzymes: cytosine deaminase (CD) and UPRT. We found that the sequential activity of these enzymes converts 5-ethynylcytosine (EC) to 5-ethynyluridine monophosphate that is subsequently incorporated into nascent RNAs. The ethynyl group allows efficient detection and purification of tagged RNAs. We show that 'EC-tagging' occurs in tissue culture cells and Drosophila engineered to express CD and UPRT. Additional control can be achieved through a split-CD approach in which functional CD is reconstituted from independently expressed fragments. We demonstrate the sensitivity and specificity of EC-tagging by obtaining cell type-specific gene expression data from intact Drosophila larvae, including transcriptome measurements from a small population of central brain neurons. EC-tagging provides several advantages over existing techniques and should be broadly useful for investigating the role of differential RNA expression in cell identity, physiology and pathology.

| S-EPMC5587779 | biostudies-literature

OmicsDI is part of the ELIXIR infrastructure

OmicsDI is an Elixir interoperability service. Learn more ›

Tweets

OmicsDI Databases

PRIDE
PeptideAtlas
MassIVE
JPOST Repository
Physiome Model Repository

EGA
EVA
ENA
LINCS
PAXDB
Cell Collective

MetaboLights
Metabolomics Workbench
MetabolomeExpress
GNPS
BioModels
FAIRDOMHub

ArrayExpress
dbGaP
ExpressionAtlas
GEO
NODE

Information

Databases
Help
API
Contact us
Code on GitHub
Terms of use
Submit Data