Transcriptome-wide analysis of protein–RNA interactions using high-throughput sequencing

Milek, Miha; Wyler, Emanuel; Landthaler, Markus

doi:10.1016/j.semcdb.2011.12.001

Cited by 54 publications

(39 citation statements)

References 76 publications

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“…Recent experiments have mapped RBP-transcriptome interactions using CLIP-seq, which consists of crosslinking of protein and RNA, followed by immunoprecipitation and sequencing of bound RNA fragments [31][32][33][34] . RBPs were typically found to interact reproducibly and specifically with thousands of target genes and tens of thousands of binding sites [35][36][37][38] , which is consistent with the specific recognition of only 4-8 nucleotides by most RBPs that bind to single-stranded RNA 39,40 .…”

Section: Clashmentioning

confidence: 99%

Competition between target sites of regulators shapes post-transcriptional gene regulation

2014

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Post-transcriptional gene regulation (PTGR) of mRNA turnover, localization and translation is mediated by microRNAs (miRNAs) and RNA-binding proteins (RBPs). These regulators exert their effects by binding to specific sequences within their target mRNAs. Increasing evidence suggests that competition for binding is a fundamental principle of PTGR. Not only can miRNAs be sequestered and neutralized by the targets with which they interact through a process termed 'sponging', but competition between binding sites on different RNAs may also lead to regulatory crosstalk between transcripts. Here, we quantitatively model competition effects under physiological conditions and review the role of endogenous sponges for PTGR in light of the key features that emerge.

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Section: Clashmentioning

confidence: 99%

Competition between target sites of regulators shapes post-transcriptional gene regulation

2014

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“…Nucleotide bases absorb UV light at this wavelength, generating free radicals that react with amino acids at very short range, forming covalent bonds. After immunoprecipitation, RNAs bound to the targeted RBP can be identified by next-generation sequencing (crosslinking, immunoprecipitation and sequencing, CLIP-seq) (Licatalosi et al 2008;Milek et al 2012). In a complementary fashion, the repertoire of proteins that bind to polyadenylated RNAs in living cells can be determined by RNA interactome capture (Baltz et al 2012;Castello et al 2012Castello et al , 2013.…”

Section: Introductionmentioning

confidence: 99%

Specific RNP capture with antisense LNA/DNA mixmers

Rogell

Fischer

Rettel

et al. 2017

RNA

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RNA-binding proteins (RBPs) play essential roles in RNA biology, responding to cellular and environmental stimuli to regulate gene expression. Important advances have helped to determine the (near) complete repertoires of cellular RBPs. However, identification of RBPs associated with specific transcripts remains a challenge. Here, we describe "specific ribonucleoprotein (RNP) capture," a versatile method for the determination of the proteins bound to specific transcripts in vitro and in cellular systems. Specific RNP capture uses UV irradiation to covalently stabilize protein-RNA interactions taking place at "zero distance." Proteins bound to the target RNA are captured by hybridization with antisense locked nucleic acid (LNA)/DNA oligonucleotides covalently coupled to a magnetic resin. After stringent washing, interacting proteins are identified by quantitative mass spectrometry. Applied to in vitro extracts, specific RNP capture identifies the RBPs bound to a reporter mRNA containing the Sex-lethal (Sxl) binding motifs, revealing that the Sxl homolog sister of Sex lethal (Ssx) displays similar binding preferences. This method also revealed the repertoire of RBPs binding to 18S or 28S rRNAs in HeLa cells, including previously unknown rRNA-binding proteins.

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“…Furthermore, combining cross-linking and immunoprecipitation (CLIP) of RNA-interacting proteins with RNA-seq enables high-throughput screening and mapping of specific RNA-protein interactions, which play a key role in transcriptome regulation. 231,232 An overview of recently developed CLIP methods is schematically shown in Figure 24.3.…”

Section: Transcriptomicsmentioning

confidence: 99%

The Path to Personalized Cardiovascular Medicine

Marı́n-Garcı́a

2014

Post-Genomic Cardiology

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Transcriptome-wide analysis of protein–RNA interactions using high-throughput sequencing

Cited by 54 publications

References 76 publications

Competition between target sites of regulators shapes post-transcriptional gene regulation

Competition between target sites of regulators shapes post-transcriptional gene regulation

Specific RNP capture with antisense LNA/DNA mixmers

The Path to Personalized Cardiovascular Medicine

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