Browse
Submit Data
Databases
API
Help

Dataset Information

0 Views

0 Connections

0 Citations

0 Reanalyses

0 Downloads

Omics score: 0

Enhanced interactome capture (eRIC) of steady-state and arsenite stressed HuH7 cell cytoplasmic and nuclear fractions

ABSTRACT: HuH7 cells were grown under steady-state conditions or exposed 100 ÂµM Na-arsenite for 1h. Subsequently, the cells were irradiated or not with 253 nm UV light (150 mJ/cm^2), processed to nuclear and cytoplasmic fractions, and subjected to eRIC for determining the nuclear and cytoplasmic RNA-binding proteins from the different conditions.

INSTRUMENT(S): Orbitrap Fusion Lumos

ORGANISM(S): Homo Sapiens (human)

TISSUE(S): Hepatocyte, Permanent Cell Line Cell, Cell Culture

SUBMITTER: Frank Stein

LAB HEAD: Andreas E. Kulozik

PROVIDER: PXD016735 | Pride | 2020-04-06

REPOSITORIES: Pride

ACCESS DATA

Json Xml

Publications

Plasticity of nuclear and cytoplasmic stress responses of RNA-binding proteins.

Backlund Michael M Stein Frank F Rettel Mandy M Schwarzl Thomas T Perez-Perri Joel I JI Brosig Annika A Zhou Yang Y Neu-Yilik Gabriele G Hentze Matthias W MW Kulozik Andreas E AE

Nucleic acids research 20200501 9

Cellular stress causes multifaceted reactions to trigger adaptive responses to environmental cues at all levels of the gene expression pathway. RNA-binding proteins (RBP) are key contributors to stress-induced regulation of RNA fate and function. Here, we uncover the plasticity of the RNA interactome in stressed cells, differentiating between responses in the nucleus and in the cytoplasm. We applied enhanced RNA interactome capture (eRIC) analysis preceded by nucleo-cytoplasmic fractionation fol ...[more]

PMID: 32313943

Similar Datasets

Enhanced RNA Interactome capture (eRIC) and RNA Interactome capture (RIC) of Jurkat cells (DSMZ, ACC-282) exposed to 0.5mM DMOG or vehicle (DMSO) for 6 hours.

Project description:1.0 â€“ 1.3 x 10e8 proliferating Jurkat cells at a density of about 1-1.5 x 10e6 cells/mL were employed per sample. Cells were incubated with 0.5mM DMOG (Cayman Chemical Company, 71210) or an equivalent volume of Dimethyl Sulfoxide (DMSO) (vehicle, Merck 1.02950.0500). The DMSO concentration in the medium was 0.023% v/v. Cells were irradiated or not with 254 nm UV light and subjected to eRIC or RIC to determine the RNA-bound proteome. Data correspond to two biologically independent experiments.

2018-09-03 | PXD010942 | Pride

Enhanced RNA Interactome capture (eRIC) and RNA Interactome capture (RIC) of Jurkat cells (DSMZ, ACC-282).

Project description:1.0 â€“ 1.3 x 10e8 proliferating Jurkat cells at a density of about 1-1.5 x 10e6 cells/mL were employed per sample. Cells were irradiated or not with 254 nm UV light and subjected to eRIC or RIC to determine the RNA-bound proteome. Data correspond to two biologically independent experiments.

2018-09-03 | PXD010941 | Pride

Enhanced RNA interactome capture (eRIC) of human malignant bone tumor cells and normal bone/mesenchymal cells

Project description:Patient-derived bone tumor (osteosarcoma and giant cell tumor of bone) cells, and the normal mesenchymal stem cells and osteoblasts were cultured and subjected to UV crosslinking (UV) at 254 nm or without crosslinking (noUV) as negative controls. Subsequently, RNA-binding proteins (RBPs) were identified by eRIC.

2024-02-28 | PXD038185 | Pride

Enhanced RNA Interactome capture (eRIC) and total proteome of mouse brain, kidney and liver

Project description:Male mice on a homogenous C57BL6/J genetic background were sacrificed at 11 to 13 weeks of age by cervical dislocation. Brain, kidneys and liver were immediately harvested, flash frozen in liquid nitrogen and stored at -80Ã‚Â°C until use. Animal handling was in accordance with guidelines approved by the European Molecular Biology Laboratory (EMBL). Organ sections (thickness 30 Ã‚Âµm) were prepared in a Cryostat (Leica Biosystems, Leica CM3050 S) set to -20 Ã‚Â°C and deposited onto SuperFrost glass slides (Carl Roth, H880); the glass slides were pre-cooled to -20Ã‚Â°C in the cryostat. ~10 sections were placed on a glass slide, and a total of around 150 sections were prepared for each sample per mouse. Tissue sections on glass slides were then transferred onto metal plates placed on dry ice. The tissue sections were exposed or not to 1 J/cm2 of 254 nm UV light in a XL 1500 UV Spectrolinker (Spectronics Corporation) and subjected to eRIC to determine the RNA-bound proteome. eRIC eluates obtained from the respective organs of two mice were combined per independent experiment. The analyses were conducted in duplicate for each organ studied. For the no crosslink controls, organ sections from four mice were pooled to generate one unique no crosslink eRIC sample per organ studied.

2023-03-02 | PXD032113 | Pride

Drosophila mRNA Interactome - Global changes of the RNA-bound proteome during the maternal-to-zygotic transition in Drosophila

Project description:The maternal-to-zygotic transition (MZT) is a process that occurs in animal embryos at the earliest developmental stages, during which maternally deposited mRNAs and other molecules are degraded and replaced by products of the zygotic genome. The zygotic genome is not activated immediately upon fertilization, therefore post-transcriptional mechanisms control the first steps of development in the early, pre-MZT embryo. To perform unbiased organism-wide identification of Drosophila RNA binding proteins (RPBs), crucial players of post-transcriptional control, we applied the recently developed RNA interactome capture method, which involves cross-linking of RNAs and their direct protein partners by UV light, purification of RNA under stringent conditions and identification of proteins by mass spectrometry. Our analysis yielded 523 high confidence RBP hits, half of which were not previously reported to bind RNA. Our comparison of the RNA interactomes of pre- and post-MZT embryos reveals a highly dynamic behavior of the RNA-bound proteome during early development, and suggests active regulation of RNA binding of some RBPs. This resource provides the first evidence of RNA binding for hundreds of Drosophila proteins, and opens new avenues for study of molecular mechanisms of early development.

2016-07-11 | PXD003882 | Pride

1 A post-transcriptional regulatory code for mRNA stability dur-ing the maternal-to-zygotic transition

Project description:Post-transcriptional regulation is crucial to shape gene expression. During the Maternal-to-Zygotic transition (MZT), thousands of maternal transcripts are regulated upon fertili-zation and genome activation. Transcript stability can be influenced by cis-elements and trans-factors, but how these inputs are integrated to determine the overall mRNA stability is unclear. Here, we show that most transcripts are under combinatorial regulation by multiple decay pathways. Characterization of the cis-regulatory motifs revealed that nu-cleotide composition bias characteristic of 3’-UTRs poly-U is associated with mRNA stability. In contrast, miR-430, CCUC, CUGC, elements appeared as the main destabiliz-ing motifs, with miR-430 and UAUUUAU (ARE) sequences causing mRNA deadenyla-tion depending on the activation of the genome. We comprehensively identify RNA-protein interactions across the transcriptome during MZT, and their associated regulatory activity. We find that poly-U binding proteins are preferentially associated with 3’-UTR sequences and stabilizing motifs. Analysis of differentially regulated regions revealed antagonistic sequence contexts for poly-C and poly-U binding proteins that shape protein binding and magnitude of regulation across the transcriptome. Finally, we integrate these regulatory motifs into a machine learning model, able to predict the stability of mRNA reporters in vivo. Our findings reveal how mechanisms of post-transcriptional regulation are coordinated to direct changes in mRNA stability within the early embryo.

2020-05-26 | PXD009514 | Pride

Poly(A) RNA interactome capture upon HDAC inhibition by Romidepsin

Project description:Immediate early genes (IEGs) represent a unique class of genes with rapid induction kinetics and transient expression patterns, which requires IEG mRNAs to be short-lived. Here, we establish cytoplasmic polyadenylation element-binding protein 4 (CPEB4) as a major determinant of IEG mRNA instability. We identified human CPEB4 as an RNA-binding protein (RBP) with enhanced association to poly(A) RNA upon inhibition of class I histone deacetylases (HDACs), which is known to cause widespread degradation of poly(A)-containing mRNA. Photoactivatable ribonucleoside-enhanced crosslinking and immunoprecipitation (PAR-CLIP) analysis using endogenously tagged CBEP4 in HeLa cells revealed that CPEB4 preferentially binds to the 3' untranslated region (UTR) of IEG mRNAs, at U-rich sequence motifs located in close proximity to the poly(A) site. By transcriptome-wide mRNA decay measurements, we found that the strength of CPEB4 binding correlates with short mRNA half-lives, and that loss of CPEB4 expression leads to the stabilization of IEG mRNAs. Further, we demonstrate that CPEB4 mediates mRNA degradation by recruitment of the evolutionarily conserved CCR4-NOT complex, the major eukaryotic deadenylase. While CPEB4 is primarily known for its ability to stimulate cytoplasmic polyadenylation, our findings establish an additional function for CPEB4 as an RBP that enhances the degradation of short-lived IEG mRNAs.

2022-08-22 | PXD029592 | Pride

Global assessment of protein-protein crosslinking in ex vivo eRIC samples

Project description:Male mice on a homogenous C57BL6/J genetic background were sacrificed at 11 to 13 weeks of age by cervical dislocation. Kidneys were immediately harvested, flash frozen in liquid nitrogen and stored at -80°C until use. Animal handling was in accordance with guidelines approved by the European Molecular Biology Laboratory (EMBL). Organ sections (thickness 30 µm) were prepared in a Cryostat (Leica Biosystems, Leica CM3050 S) set to -20 °C and deposited onto SuperFrost glass slides (Carl Roth, H880); the glass slides were pre-cooled to -20°C in the cryostat. ~10 sections were placed on a glass slide, and a total of around 150 sections were prepared for each sample per mouse. Tissue sections on glass slides were then transferred onto metal plates placed on dry ice. The tissue sections were exposed to 1 J/cm2 of 254 nm UV light in a XL 1500 UV Spectrolinker (Spectronics Corporation) and subjected to eRIC (PMID: 30352994). The proteins captured by eRIC were separated by gel electrophoresis, divided into 7 fractions of defined mass, and identified by MS to determine their distribution across the fractions. Crosslinking to other proteins should shift the signal to higher mass fractions. The analyses were conducted in triplicate.

2023-03-02 | PXD038100 | Pride

Unresolved stalled ribosome complexes restrict cycle progression after genotoxic stress

Project description:Translation of damaged mRNA can lead to ribosome stalling, thereby producing incomplete proteins toxic to the cell. The mechanism of ribosome-associated quality control (RQC) disassembles stalled ribosomes through the actions of the ASC-1 complex (ASCC). Here, we show that some reagents that chemically damage RNA, such as ultraviolet light (UV), cause ribosome stalling, which leads to accumulation of the ASC-1 complex (ASCC) on stalled ribosomes and stable interaction of the ASCC3 helicase with RNA. In contrast, the ASCC was not similarly affected by emetine or anisomycin-induced ribosome stalling. Our work identified two different types of stalled ribosome. Ribosomes arrested by emetine or anisomycin are transient as they are resolved by the ASCC. Whereas the ASCC fails to split some stalled ribosomes, such as those induced by UV, resulting in long-lived stalled ribosome complexes. We show that ribosome stalling activates the G1/S and G2/M cell cycle checkpoints with long-lived stalled ribosomes causing prolonged checkpoint activation. Thus, the cell adjusts this adaptive survival response to match the nature of the stalled ribosome.

2022-02-07 | PXD023861 | Pride

Unresolved stalled ribosome complexes restrict cycle progression after genotoxic stress

2022-02-07 | PXD023976 | Pride

OmicsDI is part of the ELIXIR infrastructure

OmicsDI is an Elixir interoperability service. Learn more ›

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