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RNA-Seq of control and EJC factor (EIF4A3, RBM8A, MAGOH) knockdowns in part with EIF4A3 rescue in HeLa Tet-Off cells

ABSTRACT: RNA-Seq of control and EJC factor (EIF4A3, RBM8A, MAGOH) knockdowns in part with EIF4A3 rescue in HeLa Tet-Off cells

PROVIDER: PRJEB46520 | ENA |

REPOSITORIES: ENA

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			Action	DRS
	ERR6349300_1.fastq.gz	Fastqsanger.gz
	ERR6349300_2.fastq.gz	Fastqsanger.gz
	ERR6349310_1.fastq.gz	Fastqsanger.gz
	ERR6349310_2.fastq.gz	Fastqsanger.gz
	ERR6349320_1.fastq.gz	Fastqsanger.gz

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RNA-Seq of control and EJC factor (EIF4A3, RBM8A, MAGOH) knockdowns in part with EIF4A3 rescue in HeLa Tet-Off cells

Project description:The exon junction complex is involved in gene expression regulation on multiple co- and post-transcriptional levels. We aimed to investigate in HeLa Tet-Off cells which gene expression alterations can be observed upon knockdown of core EJC factors (EIF4A3, RBM8A and MAGOH) or upon EIF4A3 knockdown and rescue with EIF4A3 wild type construct by using RNA-Seq analyses. The rescue construct was stably integrated into the genome using the PiggyBac transposon system. As control Luciferase (Luc) siRNA was used.

2022-05-06 | E-MTAB-10771 | biostudies-arrayexpress

Transcriptome analyses of EJC haploinsufficient neocortices

Project description:RNA sequencing of E10.5 neocortices from control, Magoh, Rbm8a, and Eif4a3 haploinsufficient embryos (generated with Emx1-Cre)

2016-09-01 | GSE85576 | GEO

Role of the Exon Junction Complex in Danio rerio

Project description:The exon junction complex (EJC) is composed of three core proteins Rbm8a, Magoh and Eif4a3 and is thought to play a role in several post-transcriptional processes. In this study we focus on understanding the role of EJC in zebrafish development. We identified transcriptome-wide binding sites of EJC in zebrafish via RNA:protein immunoprecipitation followed by deep sequencing (RIP-Seq). We find that, as in human cells, zebrafish EJC is deposited about 24 nts upstream of exon-exon junctions. We also identify transcripts regulated by Rbm8a and Magoh in zebrafish embryos using whole embryo RNA-seq from rbm8a mutant, magoh mutant and wild-type sibling embryos. This study shows that nonsense mediated mRNA decay is dysregulated in zebrafish EJC mutants.

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eIF4A3 interactome by IP-MS in mouse ESCs

Project description:RNA helicases are involved in multiple steps of RNA metabolism to direct their roles in gene expression, yet their functions in pluripotency control remain largely unexplored. Starting from an RNAi screen of RNA helicases, we identified that eIF4A3, a DEAD-box (Ddx) helicase component of the exon junction complex (EJC), is essential for the maintenance of embryonic stem cells (ESCs). We mapped the eIF4A3 interactomes in mouse ESCs, revealing that eIF4A3 is widely involved in the post-transcriptional regulation of gene expression. Mechanistically, we show that eIF4A3 post-transcriptionally controls the pluripotency-related cell cycle regulators and that its depletion causes cellular differentiation via cell cycle dysregulation. Specifically, eIF4A3 is required for the efficient nuclear export of Ccnb1 mRNA, which encodes Cyclin B1, a key component of the pluripotency-promoting pathway during cell cycle progression of ESCs. Our results reveal a previously unappreciated role of eIF4A3 and its associated EJC in the post-transcriptional cell cycle control in maintaining stem cell pluripotency.

2023-03-11 | PXD031362 | Pride

Transcriptome-wide modulation of splicing by the exon junction complex

Project description:We report that knockdown of EJC core proteins, eIF4A3, Y14, Magoh, causes a transcript-wide changes in alternative splicing, as well as some transcriptional changes. These changes are specific to EJC core proteins, and KD of UPF1 protein caused different sets of alterantive splicing changes. These changes are linked to the rate of transcription. Examination of 4 different knockdown, as well as GFP knockdown in HeLa cells, 2 replicates each condition.

2014-11-14 | E-GEOD-63091 | biostudies-arrayexpress

Transcriptome-wide modulation of splicing by the exon junction complex

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Mammalian UPF3A and UPF3B activate NMD independently of their EJC binding

Project description:Nonsense-mediated mRNA decay (NMD) can occur independently of some NMD factors such as UPF3B. We have performed total RNA-Seq in HCT116 cells under NMD factor knockout (with CRISPR-Cas9) and/or knockdown (with siRNA transfection) conditions to identify mRNA substrates regulated by different NMD factors. We have also performed RIPiT-Seq to identify the footprints of three compositionally distinct EJCs that contain the following pairs of proteins: MAGOH-EIF4A3, UPF3B-EIF4A3 or CASC3-EIF4A3 in WT HCT116 cells.

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mRNAseq of Drosophila S2 cells after knockdown of EJC components

Project description:This study used Drosophila melanogaster S2 cells treated with siRNA against the EJC components mago or eIF4A3, or a non-targeting control siRNA (three samples per condition). mRNA-seq libraries were prepared from the samples and the data was used to examine 'RS-exons' - exons which reconstitute a 5' splice site when they are spliced to the upstream exon. In contrast to mammalian cells, RS-exons are not recursively spliced out from Drosophila transcripts after perturbation of the EJC.

2018-10-15 | E-MTAB-7271 | biostudies-arrayexpress

PRPF8 iCLIP on HeLa cells in response to eIF4A3 Knockdown

Project description:This experiment uses iCLIP to identify the binding pattern of the spliceosomal protein PRPF8 on RNA. The data shows that PRPF8 binds strongly and specifically in the region 12 to 14nt upstream of 5' splice sites (5ss). Due to PRPF8's role in the formation of the catalytically active spliceosome, this data can be used as a readout of 5ss selection. Here, we performed iCLIP on HeLa cells treated with control or EIF4A3 siRNA, with 4 replicate samples per condition and eIF4A3 protein levels reduced ~50% in knockdown. We investigated the role of the exon junction complex (EJC) in suppressing 5ss that are reconstituted at the junction of two canonical exons (RS-5ss) - selection of these splice sites would result in recursive splicing of canonical exons. We plotted the crosslink sites of reads that span an exon-exon junction, seperating reads that span RS-5ss from those that do not. We found that reads that span an RS-5ss are enriched at the 12-14nt window associated with 5ss selection, while reads that span other exon-exon junctions are not enriched. This effect is magnified greatly by knockdown of eIF4A3. The results indicate that RS-5ss can be used by the spliceosome, but that this process is usually repressed by the EJC. This data is evidence of recursive splicing of canonical exons and the role of the EJC in repressing recursive splicing.

2018-10-14 | E-MTAB-7269 | biostudies-arrayexpress

Magoh, Y14, eIF4AIII interactomes by TurboID proximity labelling provide no evidence for a canonical EJC in (almost) intron-less trypanosomatids

Project description:In metazoans, mRNA quality is tightly monitored from transcription to translation. A key role lies with the exon junction complex (EJC) that is placed upstream of the exon-exon junction after splicing. The EJC inner core is composed of Magoh, Y14, eIF4AIII and BTZ and the outer core of proteins involved in mRNA splicing (CWC22), export (Yra1), translation (PYM) and non-sense mediated decay (NMD, UPF1/2/3). The protozoan parasite Trypanosoma brucei encodes only two genes with introns, but all mRNAs are processed by trans-splicing. The presence of the three core EJC proteins and a potential BTZ homologue (Rbp25) in trypanosomes has been suggested as an adaptation of the EJC function to mark trans-spliced mRNAs. Here we explore the interactome of Magoh, Y14, eIF4AIII in T. brucei by TurboID proximity labelling.

2024-03-25 | PXD050946 | Pride

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