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Exon Junction Complexes Suppress Spurious Splice Sites to Safeguard Transcriptome Integrity.

Molecular cell | 2018

Productive splicing of human precursor messenger RNAs (pre-mRNAs) requires the correct selection of authentic splice sites (SS) from the large pool of potential SS. Although SS consensus sequence and splicing regulatory proteins are known to influence SS usage, the mechanisms ensuring the effective suppression of cryptic SS are insufficiently explored. Here, we find that many aberrant exonic SS are efficiently silenced by the exon junction complex (EJC), a multi-protein complex that is deposited on spliced mRNA near the exon-exon junction. Upon depletion of EJC proteins, cryptic SS are de-repressed, leading to the mis-splicing of a broad set of mRNAs. Mechanistically, the EJC-mediated recruitment of the splicing regulator RNPS1 inhibits cryptic 5'SS usage, while the deposition of the EJC core directly masks reconstituted 3'SS, thereby precluding transcript disintegration. Thus, the EJC protects the transcriptome of mammalian cells from inadvertent loss of exonic sequences and safeguards the expression of intact, full-length mRNAs.

Pubmed ID: 30388410 RIS Download

Associated grants

None

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This is a list of tools and resources that we have found mentioned in this publication.


Human Splicing Finder (tool)

RRID:SCR_005181

Software tool to help study pre-mRNA splicing and to better understand intronic and exonic mutations leading to splicing defects. To calculate the consensus values of potential splice sites and search for branch points, new algorithms were developed. Furthermore, they have integrated all available matrices to identify exonic and intronic motifs, as well as new matrices to identify hnRNP A1, Tra2-? and 9G8.

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CummeRbund (tool)

RRID:SCR_014568

Software R package used for simplifying and analyzing Cufflink RNA-Seq output. This program takes various output files from a cuffdiff run and creates a SQLite database of the results that will describe the appropriate relationships between the genes, transcripts, transcription start sites and CDS regions.

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MAJIQ (tool)

RRID:SCR_016706

Software package to detect and quantify local splicing variations (LSV) from RNA-Seq data. Used to automatically detect and downweight outliers in RNA-Seq datasets with replicates for differential splicing (SD) analysis.

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STAR (tool)

RRID:SCR_004463

Software performing alignment of high-throughput RNA-seq data. Aligns RNA-seq reads to reference genome using uncompressed suffix arrays.

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SAP18 antibody (antibody)

RRID:AB_777829

This polyclonal targets Human SAP18

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Monoclonal Anti-alpha-Tubulin antibody produced in mouse (antibody)

RRID:AB_477582

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Anti-β-Actin Antibody (antibody)

RRID:AB_476744

This monoclonal targets slightly modified β-cytoplasmic actin N-terminal peptide, Ac-Asp-Asp-Asp-Ile-Ala-Ala-Leu-Val-Ile-Asp-Asn-Gly-Ser-Gly-Lys, conjugated to KLH

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GFP-avictoria (antibody)

RRID:AB_303395

This polyclonal targets GFP

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Flp-In-T-REx-293 (cell line)

RRID:CVCL_U427

Cell line Flp-In-T-REx-293 is a Transformed cell line with a species of origin Homo sapiens (Human)

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HeLa Tet-Off (cell line)

RRID:CVCL_V352

Cell line HeLa Tet-Off is a Cancer cell line with a species of origin Homo sapiens (Human)

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Integrative Genomics Viewer (software resource)

RRID:SCR_011793

A high-performance visualization tool for interactive exploration of large, integrated genomic datasets.

View all literature mentions

Image Lab Software (software resource)

RRID:SCR_014210

Imaging software used to acquire and analyze images from specific Bio-Rad imaging systems. Users can analyze gel or blot features, capture optimized image data, and generate a report of the data. Image Lab software exclusively runs on the Gel Doc EZ imager, Gel Doc XR+ imaging system, ChemiDo MP, ChemiDoc XRS+ imaging systems, Criterion Stain Free imager, and the GS-900calibrated densitometer.

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GraphPad Prism (software resource)

RRID:SCR_002798

Statistical analysis software that combines scientific graphing, comprehensive curve fitting (nonlinear regression), understandable statistics, and data organization. Designed for biological research applications in pharmacology, physiology, and other biological fields for data analysis, hypothesis testing, and modeling.

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Cufflinks (software resource)

RRID:SCR_014597

Software tool for transcriptome assembly and differential expression analysis for RNA-Seq. Includes script called cuffmerge that can be used to merge together several Cufflinks assemblies. It also handles running Cuffcompare as well as automatically filtering a number of transfrags that are likely to be artifacts. If the researcher has a reference GTF file, the researcher can provide it to the script to more effectively merge novel isoforms and maximize overall assembly quality.

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Bowtie 2 (software resource)

RRID:SCR_016368

Ultrafast and memory efficient tool for aligning sequencing reads to long reference sequences. Supports gapped, local, and paired end alignment modes. More suited to finding longer, gapped alignments in comparison with original Bowtie method.

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Ensembl (software resource)

RRID:SCR_002344

Collection of genome databases for vertebrates and other eukaryotic species with DNA and protein sequence search capabilities. Used to automatically annotate genome, integrate this annotation with other available biological data and make data publicly available via web. Ensembl tools include BLAST, BLAT, BioMart and the Variant Effect Predictor (VEP) for all supported species.

View all literature mentions

STAR (software resource)

RRID:SCR_015899

Software performing alignment of high-throughput RNA-seq data. Aligns RNA-seq reads to reference genome using uncompressed suffix arrays.

View all literature mentions

Image Lab Software (software resource)

RRID:SCR_014210

Imaging software used to acquire and analyze images from specific Bio-Rad imaging systems. Users can analyze gel or blot features, capture optimized image data, and generate a report of the data. Image Lab software exclusively runs on the Gel Doc EZ imager, Gel Doc XR+ imaging system, ChemiDo MP, ChemiDoc XRS+ imaging systems, Criterion Stain Free imager, and the GS-900calibrated densitometer.

View all literature mentions

STAR (software resource)

RRID:SCR_015899

Software performing alignment of high-throughput RNA-seq data. Aligns RNA-seq reads to reference genome using uncompressed suffix arrays.

View all literature mentions

Cufflinks (software resource)

RRID:SCR_014597

Software tool for transcriptome assembly and differential expression analysis for RNA-Seq. Includes script called cuffmerge that can be used to merge together several Cufflinks assemblies. It also handles running Cuffcompare as well as automatically filtering a number of transfrags that are likely to be artifacts. If the researcher has a reference GTF file, the researcher can provide it to the script to more effectively merge novel isoforms and maximize overall assembly quality.

View all literature mentions

Bowtie 2 (software resource)

RRID:SCR_016368

Ultrafast and memory efficient tool for aligning sequencing reads to long reference sequences. Supports gapped, local, and paired end alignment modes. More suited to finding longer, gapped alignments in comparison with original Bowtie method.

View all literature mentions

GraphPad Prism (software resource)

RRID:SCR_002798

Statistical analysis software that combines scientific graphing, comprehensive curve fitting (nonlinear regression), understandable statistics, and data organization. Designed for biological research applications in pharmacology, physiology, and other biological fields for data analysis, hypothesis testing, and modeling.

View all literature mentions

Integrative Genomics Viewer (software resource)

RRID:SCR_011793

A high-performance visualization tool for interactive exploration of large, integrated genomic datasets.

View all literature mentions

SAP18 antibody (antibody)

RRID:AB_777829

This polyclonal targets Human SAP18

View all literature mentions

GFP-avictoria (antibody)

RRID:AB_303395

This polyclonal targets GFP

View all literature mentions

HeLa Tet-Off (cell line)

RRID:CVCL_V352

Cell line HeLa Tet-Off is a Cancer cell line with a species of origin Homo sapiens (Human)

View all literature mentions

Flp-In-T-REx-293 (cell line)

RRID:CVCL_U427

Cell line Flp-In-T-REx-293 is a Transformed cell line with a species of origin Homo sapiens (Human)

View all literature mentions

Ensembl (software resource)

RRID:SCR_002344

Collection of genome databases for vertebrates and other eukaryotic species with DNA and protein sequence search capabilities. Used to automatically annotate genome, integrate this annotation with other available biological data and make data publicly available via web. Ensembl tools include BLAST, BLAT, BioMart and the Variant Effect Predictor (VEP) for all supported species.

View all literature mentions

Monoclonal Anti-alpha-Tubulin antibody produced in mouse (antibody)

RRID:AB_477582

This monoclonal targets alpha-Tubulin antibody produced in mouse

View all literature mentions

Anti-β-Actin Antibody (antibody)

RRID:AB_476744

This monoclonal targets slightly modified β-cytoplasmic actin N-terminal peptide, Ac-Asp-Asp-Asp-Ile-Ala-Ala-Leu-Val-Ile-Asp-Asn-Gly-Ser-Gly-Lys, conjugated to KLH

View all literature mentions