Browse
Submit Data
Databases
API
Help

Dataset Information

0 Views

0 Connections

0 Citations

0 Reanalyses

0 Downloads

Omics score: 0

Subcellular refinement of RNA binding protein target interactions with colocalization CLIP [CLIP-seq]

ABSTRACT: RNA binding proteins (RBPs) are essential for RNA metabolism and have profound impacts in both health and disease. The subcellular organization of RBP interaction networks with target RNAs remains largely unexplored. Here, we develop colocalization CLIP, a method that combines CrossLinking and ImmunoPrecipitation (CLIP) with proximity labeling, to explore in-depth the subcellular RNA interactions of the well-studied RNA-binding protein HuR. Using this method, we uncover HuR's dynamic and location-specific interactions with RNA, revealing alterations in sequence preferences and interactions in the nucleus, cytosol, or stress granule compartments. We uncover HuR's unique binding preferences within stress granules during arsenite stress, illuminating intricate interactions that conventional methodologies cannot capture. Overall, coCLIP provides a powerful method for revealing RBP:RNA interactions based on localization, and lays the foundation for an advanced understanding of RBP models that incorporate subcellular location as a critical determinant of their functions.

ORGANISM(S): Homo sapiens

PROVIDER: GSE245209 | GEO | 2023/10/25

REPOSITORIES: GEO

ACCESS DATA

Json Xml

Similar Datasets

Subcellular refinement of RNA binding protein target interactions with colocalization CLIP [RNA-seq]

Project description:RNA binding proteins (RBPs) are essential for RNA metabolism and have profound impacts in both health and disease. The subcellular organization of RBP interaction networks with target RNAs remains largely unexplored. Here, we develop colocalization CLIP, a method that combines CrossLinking and ImmunoPrecipitation (CLIP) with proximity labeling, to explore in-depth the subcellular RNA interactions of the well-studied RNA-binding protein HuR. Using this method, we uncover HuR's dynamic and location-specific interactions with RNA, revealing alterations in sequence preferences and interactions in the nucleus, cytosol, or stress granule compartments. We uncover HuR's unique binding preferences within stress granules during arsenite stress, illuminating intricate interactions that conventional methodologies cannot capture. Overall, coCLIP provides a powerful method for revealing RBP:RNA interactions based on localization, and lays the foundation for an advanced understanding of RBP models that incorporate subcellular location as a critical determinant of their functions.

2023-10-25 | GSE245210 | GEO

Global Analysis of Yeast mRNPs

Project description:Proteins regulate gene expression by controlling mRNA biogenesis, localization, translation and decay. Identifying the composition, diversity and function of mRNPs (mRNA protein complexes) is essential to understanding these processes. In a global survey of S. cerevisiae mRNA binding proteins we identified 120 proteins that cross-link to mRNA, including 66 new mRNA binding proteins. These include kinases, RNA modification enzymes, metabolic enzymes, and tRNA and rRNA metabolism factors. These proteins show dynamic subcellular localization during stress, including assembly into stress granules and P-bodies (Processing-bodies). CLIP (cross-linking and immunoprecipitation) analyses of the P-body components Pat1, Lsm1, Dhh1 and Sbp1 identified sites of interaction on specific mRNAs revealing positional binding preferences and co-assembly preferences. Taken together, this work defines the major yeast mRNP proteins, reveals widespread changes in their subcellular location during stress, and begins to define assembly rules for P-body mRNPs. CLIP-seq analysis of Dhh1, Lsm1, Pat1 and Sbp1

2013-04-18 | E-GEOD-46142 | biostudies-arrayexpress

Analysis of CLIP and iCLIP methods for nucleotide-resolution studies of protein-RNA interactions

Project description:UV cross-linking and immunoprecipitation (CLIP) and individual-nucleotide resolution CLIP (iCLIP) are the most frequently used methods to study protein-RNA interactions in the intact cells and tissues, but their relative advantages or inherent biases have not been evaluated. To benchmark CLIP and iCLIP method, we performed iCLIP with Nova protein, which is the most extensively studied protein by CLIP. Further, we assessed UV-C-induced cross-linking preferences, by exploiting the UV-independent formation of covalent RNA cross-links of the mutant RNA methylase NSUN2.

2012-07-20 | E-MTAB-1008 | biostudies-arrayexpress

GoldCLIP: Gel-omitted Ligation-dependent CLIP

Project description:Protein-RNA interaction networks are essential to understand gene regulation control. Identifying the binding sites of RNA-binding proteins (RBPs) by CLIP (UV-crosslinking and immunoprecipitation) represents one of the most powerful methods to map in vivo protein-RNA interactions. However, the traditional CLIP protocol is technically challenging, which requires radioactive labeling and suffers from material loss during PAGE-membrane transfer procedures. Here we introduce a super-efficient CLIP method (GoldCLIP) that omits all gel purification steps. This nonisotopic method allowed us to perform highly reproducible CLIP experiments with classical RBP such as PTB in human cell lines. In principle, our method guarantees sequencing library constructions, providing the protein of interest can be successfully crosslinked to RNAs in living cells. GoldCLIP is readily applicable to diverse factors to uncover their endogenous targets.

2018-05-20 | GSE111406 | GEO

Implementation of PAR-CLIP to characterize RNA-protein interactions in prokaryotes at nucleotide resolution

Project description:The identification of RNAs that are recognized by RNA-binding proteins (RNA-BPs) using techniques such as Crosslinking and Immunoprecipitation (CLIP) has revolutionized the genome-wide discovery of RNA-BP RNA targets. Among the different versions of CLIP that have been developed, the use of photoactivable nucleoside analogs has resulted in high efficiency photoactivable ribonucleoside-enhanced CLIP (PAR-CLIP) in vivo. Nonetheless, PAR-CLIP has not yet been applied in prokaryotes. To determine if PAR-CLIP can be used in prokaryotes, we determined suitable conditions for the incorporation of 4-thiouridine (4SU), a photoactivable nucleoside, into E. coli RNA and for the isolation of RNA crosslinked to RNA-BPs of interest. Applying this technique to Hfq, a well-characterized regulator of small RNA (sRNA)-messenger RNA (mRNA) interactions, we showed that PAR-CLIP identified most of the known sRNA targets of Hfq, as well as functionally relevant sites of Hfq-mRNA interactions at nucleotide resolution. Based on our findings, PAR-CLIP represents an improved method to identify both the RNAs and the specific regulatory sites that are recognized by RNA-BPs in prokaryotes.

2022-10-18 | MSV000090547 | MassIVE

Global Analysis of Yeast mRNPs

2013-04-18 | GSE46142 | GEO

Discovering the interactions between circRNAs and RNA-binding proteins from RNA-seq and CLIP-seq data

Project description:Although tens of thousands of circular RNAs (circRNAs) have been identified in mammalian genomes, only a few have been functionally characterized. Here, we report a new approach, circScan, to identify regulatory interactions between circRNAs and RNA-binding proteins (RBPs). Our method identifies back-splicing reads from crosslinking and immunoprecipitation followed by high-throughput sequencing (CLIP-seq) data. Using our method, we systematically scanned ~1,500 CLIP-seq datasets and identified ~12,540 and ~1,090 novel circRNA-RBP interactions in the human and mouse genomes, respectively, including all known interactions between circRNAs and Argonaute (AGO) proteins. Furthermore, more than twenty novel interactions were experimentally confirmed by RNA immunoprecipitation combined with quantitative PCR (RIP-qPCR). Importantly, we determined that natural circRNAs interact with the cap-independent translation factors eukaryotic initiation factor 3 (eIF3) and N6-methyladenosine (m6A), thus suggesting that those circRNAs may be translated into proteins. These findings demonstrate that circRNAs are regulated by various RBPs and therefore may play important roles in diverse biological processes.

2018-03-18 | GSE97382 | GEO

Advancing Functional Utility of PAR-CLIP by Quantifying Background Binding to mRNAs and lncRNAs

Project description:We developed a method for measuring non-specific background in PAR-CLIP data demonstrating that covalently crosslinked background binding is common, reproducible and apparently universal. Furthermore, we show that quantitative determination of background is essential for identifying targets of weakly binding RNA-binding proteins and can substantially improve motif analysis. To define background binding events in PAR-CLIP data we performed the standard PAR-CLIP protocol (Hafner et al., Cell 2010.) on lysates expressing a commonly used non-RBP control, FLAG-GFP. After FLAG-tag immunopurification of UV 365nm irradiated lysates prepared from cells supplemented with 4-thiouridine (4SU), RNA was partially digested with RNase T1, radiolabeled and separated by SDS-PAGE. Reads were sequenced by Illumina HiSeq. PAR-CLIP was also performed for HuR. Included as well is a total from lysates treated like PAR-CLIP, but without immunoprecipitation (see sample description for more detail).

2013-11-07 | E-GEOD-50989 | biostudies-arrayexpress

Tranposable elements modulate human mRNAs and lncRNAs via specific RNA-protein interactions.

Project description:Transposable elements (TEs) have significantly influenced the evolution of transcriptional regulatory networks in the human genome. Post-transcriptional regulation of human genes by TE-derived sequences has been observed in specific contexts, but has yet to be systematically and comprehensively investigated. Here, studied a collection of CLIP-Seq (CrossLinked ImmunoPrecipitation) experiments mapping the RNA binding sites for a diverse set of 46 human proteins across 68 experiments to explore the role of TEs in post-transcriptional regulation genome-wide via RNA-protein interactions. We detected widespread interactions between RNA binding proteins (RBPs) and various families of TE-derived sequence in the CLIP-Seq data. Alignment coverage clustered on specific positions of the TE consensus sequences, illuminating a diversity of TE-specific motifs for many RBPs. Evidence of binding and conservation of these motifs in the nonrepetitive transcriptome suggest that TEs have appropriated existing sequence preferences of the RBP. Upon depletion of the RBPs, transcripts possessing TE-derived binding sites were similarly regulated as those bound in nonrepetitive sequence. However, in a few cases the effect of RBP binding depended on the specific TE family boundM-bM-^@M-^Te.g., the ubiquitously expressed RBP HuR conferred opposite effects on stability to transcripts when bound to Alu elements versus other families. Our meta-analysis suggests a widespread role for TEs in shaping RNA-protein regulatory networks in the human genome. HuR formaldehyde RIP-Seq in K562 cells, with RIP and input sequenced in triplicate.

2014-09-10 | E-GEOD-61238 | biostudies-arrayexpress

rec-Y2H matrix in S. cerevisiae screening reveals a vast potential for direct protein-protein interactions among RNA binding proteins

Project description:RNA-binding proteins (RBPs) are crucial factors of post-transcriptional gene regulation and their modes of action are intensely investigated. At the center of attention are RNA motifs that guide where RBPs bind. However, sequence motifs recognized by RBPs are typically a poor predictor of RBP-RNA interactions in vivo. It is hence believed that many RBPs recognize RNAs as complexes, to increase specificity and regulatory potential. To probe the potential for RBP–RBP complex formation, we assembled a library of 978 mammalian RBPs and used rec-Y2H screening to detect direct interactions between RBPs, sampling >1M possible interactions. We discovered 1994 new interactions and demonstrate that our interaction screening discovers RBP pairs that bind RNAs adjacently. We further find that the mRNA binding region preferences of an RBP can deviate, depending on its adjacently binding interaction partner. Finally, we reveal novel RBP–RBP interaction networks among major RNA processing steps and show that RBP mutations observed in cancer rewire spliceosomal interaction networks.

2021-09-01 | E-MTAB-9612 | biostudies-arrayexpress

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