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ChIP of PolII from cells with Mago knockdown, control knockdown, and RnpS1 knockdwon [Promoter-proximal pausing mediated by the exon junction complex regulates splicing]

ABSTRACT: ChIP of PolII from cells with Mago knockdown, control knockdown, and RnpS1 knockdwon [Promoter-proximal pausing mediated by the exon junction complex regulates splicing]

PROVIDER: PRJNA418419 | ENA |

REPOSITORIES: ENA

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			Action	DRS
	SRR6292564_1.fastq.gz	Fastqsanger.gz
	SRR6292564_2.fastq.gz	Fastqsanger.gz
	SRR6292565_1.fastq.gz	Fastqsanger.gz
	SRR6292565_2.fastq.gz	Fastqsanger.gz
	SRR6292566_1.fastq.gz	Fastqsanger.gz

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RNA-Seq of control and RNPS1 knockdown with or without RNPS1 wildtype or RNPS1-RRM domain rescue in Flp-In T-REx 293 cells

Project description:RNPS1 is a splicing regulatory protein and a component of the ASAP/PSAP complex, which is associated with the exon junction complex and modulates alternative splicing. It was previously postulated that the isolated RRM domain of RNPS1 in complex with ASAP/PSAP is able to regulate certain alternative splicing events. We aimed to investigate in Flp-In T-REx 293 cells which alternative splicing events are rescued by the expression of the wild type RNPS1 or the isolated RRM domain of RNPS1 in a RNPS1 knockdown background by using RNA-Seq analyses. The rescue construct was stably integrated into the genome using the PiggyBac transposon system. As controls, either Luciferase (Luc) siRNA was used or RNPS1 was knocked down without rescue.

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ChIP of Ser2P PolII from cells with Mago knockdown and control knockdown [Promoter-proximal pausing mediated by the exon junction complex regulates splicing]

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RNA-Seq of control, RNPS1 knockdown and rescue in HeLa cells

Project description:We report the knockdown and rescue of RNPS1, a component of the ASAP and PSAP complexes and, hence, an auxiliary component of the exon junction complex. The studied conditions show an extensive change in alternative splicing and the cell's transcriptome. For the knockdown, HeLa cells were transfected with siRNAs targeting RNPS1 or the Luciferase control. Total RNA was extracted with peqGOLD TriFast, underwent ribosomal depletion and strand-specific library preparation (TruSeq R Stranded Total RNA LT). Sequencing occurred in an Illumina HiSeq4000 sequencer with 2×75bp, producing roughly 35 million read-pair per sample.

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ChIP of PolII from cells with Mago knockdown, control knockdown, and RnpS1 knockdwon [Promoter-proximal pausing mediated by the exon junction complex regulates splicing]

Project description:Promoter-proximal pausing of RNA polymerase II (Pol II) is a widespread transcriptional regulatory step across metazoans. Here we find that the nuclear exon junction complex (pre-EJC) is a critical and conserved regulator of this process. Depletion of pre-EJC subunits leads to a global decrease in Pol II pausing and to premature entry into elongation. This effect occurs, at least in part, via non-canonical recruitment of pre-EJC components at promoters. Failure to recruit the pre-EJC at promoters results in increased binding of the positive transcription elongation complex (P-TEFb) and in enhanced Pol II release. Notably, restoring pausing is sufficient to rescue exon skipping and the photoreceptor differentiation defect associated with depletion of pre-EJC components in vivo. We propose that the pre-EJC serves as an early transcriptional checkpoint to prevent premature entry into elongation, ensuring proper recruitment of RNA processing components that are necessary for exon definition.

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RNA-Seq of control and RNPS1 knockdown with or without RNPS1 wildtype or RNPS1-RRM domain rescue in Flp-In T-REx 293 cells

Project description:RNA-Seq of control and RNPS1 knockdown with or without RNPS1 wildtype or RNPS1-RRM domain rescue in Flp-In T-REx 293 cells

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Effect of mago nashi depletion on gene expression in Drosophila S2R+ cells

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Insights into transcriptome surveillance and alternative splicing regulation by the EJC-associated splicing factor RNPS1

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