An autoinhibitory intramolecular interaction proof-reads RNA recognition by the essential splicing factor U2AF2
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ABSTRACT:
PROVIDER: PRJNA522997 | ENA |
REPOSITORIES: ENA
Dataset's files
| Action | DRS | |||
|---|---|---|---|---|
| SRR8587620.fastq.gz | Fastqsanger.gz | |||
| SRR8587690.fastq.gz | Fastqsanger.gz | |||
| SRR8587621.fastq.gz | Fastqsanger.gz | |||
| SRR8587691.fastq.gz | Fastqsanger.gz | |||
| SRR8587622.fastq.gz | Fastqsanger.gz |
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Project description:This SuperSeries is composed of the SubSeries listed below.
2020-03-05 | GSE126694 | GEO
Project description:The recognition of cis-regulatory RNA motifs in human transcripts by RNA binding proteins (RBPs) is essential for gene regulation. The molecular features that determine RBP specificity are often poorly understood. Here, we combined NMR structural biology with high-throughput iCLIP approaches to identify a regulatory mechanism for U2AF2 RNA recognition. We found that the intrinsically disordered linker region connecting the two RNA recognition motif (RRM) domains of U2AF2 mediates autoinhibitory intramolecular interactions to reduce nonproductive binding to weak Py-tract RNAs. This proofreading favors binding of U2AF2 at stronger Py-tracts, as required to define 3' splice sites at early stages of spliceosome assembly. Mutations that impair the linker autoinhibition enhance the affinity for weak Py-tracts result in promiscuous binding of U2AF2 along mRNAs and impact on splicing fidelity. Our findings highlight an important role of intrinsically disordered linkers to modulate RNA interactions of multidomain RBPs.
| S-EPMC7132250 | biostudies-literature
Project description:Proof-of-concept of a new method involving the limited digestion and subsequent ligation of intramolecular RNA structures in situ followed by deep sequencing Proof-of-concept of RPL in S. cerevisiae and H. sapiens tissue culture
2015-08-01 | E-GEOD-69472 | biostudies-arrayexpress
Project description:Proof-of-concept of a new method involving the limited digestion and subsequent ligation of intramolecular RNA structures in situ followed by deep sequencing
2015-08-01 | GSE69472 | GEO
Project description:The essential pre-mRNA splicing factor U2AF2 (also called U2AF65) identifies polypyrimidine (Py) tract signals of nascent transcripts, despite length and sequence variations. Previous studies have shown that the U2AF2 RNA recognition motifs (RRM1 and RRM2) preferentially bind uridine-rich RNAs. Nonetheless, the specificity of the RRM1/RRM2 interface for the central Py tract nucleotide has yet to be investigated. Enhanced crosslinking and immunoprecipitation of endogenous U2AF2 in human erythroleukemia cells showed uridine-sensitive binding sites with lower sequence conservation at the central nucleotide positions of otherwise uridine-rich, U2AF2-bound splice sites. Altogether, these results highlight the importance of RNA flexibility for protein recognition and take a step towards relating splice site motifs to pre-mRNA splicing efficiencies. Keywords: splicing, RNA binding, U2AF2, U2AF65, 3'SS, 3' splice site, eCLIP, polypyrimidine tract, RNA recognition motif, RRM, U-rich, Py-tract
2022-06-01 | GSE195669 | GEO
Project description:Spliceosome recognition of pre-mRNA junctions requires binding of splicing factors to the 3' splice site. Most splicing occurs co-transcriptionally, but how splicing factors bind expressed gene is not understood. We show that the SF3B1, U2AF1, and U2AF2 splicing factors bind active promoters of both intron-containing and intronless genes. RNase A treatment reduces binding at promoters, implicating general interactions with nascent transcripts in promoter localization. Strikingly, U2AF2 also accumulates on the bodies of some genes and is not eliminated by RNase treatment. Neither RNA polymerase II nor the active gene body chromatin mark H3K36me3 is required to maintain U2AF2 on chromatin. We found that most exons in U2AF2-bound have phased nucleosomes at their exon-intron boundaries. However, some exons are poorly phased, and these require U2AF2 for inclusion in mature mRNA. We propose that nucleosome phasing and U2AF2 binding ensure exon inclusion.
2025-04-10 | GSE270327 | GEO
Project description:Changes in the amino acid sequences of proteins cause thousands of human genetic diseases. However, only a subset of variants in any protein is typically pathogenic, with variants having a diversity of molecular consequences. Determining which of the thousands of possible variants in any protein have similar molecular effects is very challenging, but crucial for identifying pathogenic variants, determining disease mechanisms, understanding clinical phenotypic variation, and developing targeted therapeutics. Here we present a general method to classify variants by their molecular effects that we term intramolecular genetic interaction profiling. The approach relies on the principle that variants with similar molecular consequences have similar genetic interactions with other variants in the same protein. These intramolecular genetic interactions are straightforward to quantify for any protein with a selectable function. We apply intramolecular genetic interaction profiling to amyloid beta, the protein that aggregates in Alzheimer’s disease (AD) and is mutated in familial AD (fAD). Genetic interactions identify two classes of gain-of-function variants, with all known familial Alzheimer’s disease variants having very similar genetic interaction profiles, consistent with a common gain-of-function mechanism leading to pathology. We believe that intramolecular genetic interaction profiling is a powerful approach for classifying variants in disease genes that will empower rare variant association studies and the discovery of disease mechanisms.
2023-11-15 | GSE247583 | GEO
Project description:Analysis of U2AF2-TurboID-HA interactions by immunoprecipitation for comparison with proximity labeling. Cells expressing U2AF2 (iU2) fused with TurboID in either the undifferentiated state (D0) or after mesoderm differentiation (D1) were analyzed to identify the interactome and detect changes during the differentiation process.
2025-04-13 | PXD062890 | Pride
Project description:T cell activation leads to dramatic changes in cellular phenotype. We used activated human CD4 T cells to study how RNA binding proteins define the post-transcriptional landscape. Using RIPseq, we identified the RNA interactome of U2AF2 and show that U2AF2 binds the majority of transcripts that are differentially expressed and/or alternatively spliced during activation. Using RIP mass spectrometry, a unique protein interactome centered on U2AF2 is assembled by activation and comprised of both directly bound central members (RNAse-resistant) and indirectly bound peripheral members (RNAse-sensitive). Knocking down specific U2AF2 interactome members (U2AF1, SYNCRIP, SRRM2, ILF2) selectively affects cytokine secretion and activation markers. The expression and/or alternative splicing of transcripts important for immune cell function are also affected by knocking down these interactome members, both peripheral and central. Furthermore, we show that knockdown of interactome members can affect the proteins and transcripts bound to U2AF2, altering the transcriptome of activated T cells. Our work highlights the importance of understanding the assembly of RNA binding protein complexes as regulators of T cell activation and function.
2016-04-27 | PXD001843 | Pride
Project description:Alternative splicing drives transcriptome and proteome diversification. While splicing regulatory proteins govern this process, their global mechanisms remain enigmatic. We generated high resolution transcriptome-wide protein-RNA interaction maps to determine how the splicing repressor hnRNPA1 influences the global association of spliceosome assembly factor U2AF2 and SRSF1 with pre-mRNA. We observed changes in the distribution of U2AF2 crosslinking sites relative to the 3’ splice sites of cassette exons but not constitutive exons upon hnRNPA1 overexpression. By contrast, SRSF1 crosslinking patterns relative to splice sites are independent of hnRNPA1 expression levels. We also observed an hnRNPA1-dependent increase in U2AF2 but not SRSF1 crosslinking to exon proximal antisense Alu elements. Thus Alu elements can serve as splicing factor-responsive sinks for U2AF2. These results not only demonstrate a novel mechanism for alternative splicing regulation but also implicate retrotransposon-derived sequences in the evolution of species-specific alternative splicing.
2017-06-01 | GSE83923 | GEO