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Global evidence for programmed switching of gene expression from astrocytes to neurons in response to PTB depletion

ABSTRACT: Global evidence for programmed switching of gene expression from astrocytes to neurons in response to PTB depletion

PROVIDER: PRJNA596394 | ENA |

REPOSITORIES: ENA

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			Action	DRS
	SRR10729700.fastq.gz	Fastqsanger.gz
	SRR10729701.fastq.gz	Fastqsanger.gz
	SRR10729702.fastq.gz	Fastqsanger.gz
	SRR10729703.fastq.gz	Fastqsanger.gz
	SRR10729704.fastq.gz	Fastqsanger.gz

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Global evidence for programmed switching of gene expression from astrocytes to neurons in response to PTB depletion

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Project description:Recent transcriptome analysis indicates that >90% of human genes undergoes alternative splicing, underscoring the contribution of differential RNA processing to diverse proteomes in higher eukaryotic cells. The polypyrimidine tract binding protein PTB is a well-characterized splicing repressor, but PTB knockdown causes both exon inclusion and skipping. Genome-wide mapping of PTB-RNA interactions and construction of a functional RNA map now revealed that dominant PTB binding near a competing constitutive splice site generally induces exon inclusion whereas prevalent binding close to an alternative site often causes exon skipping. This positional effect was further demonstrated by disrupting or creating a PTB binding site on minigene constructs and testing their responses to PTB knockdown or overexpression. These findings suggest a mechanism for PTB to modulate splice site competition to produce opposite functional consequences, which may be generally applicable to RNA binding splicing factors to positively or negatively regulate alternative splicing in mammalian cells. Examination of PTB-RNA binding in Hela cells using CLIP-seq (Cross-Linking ImmunoPrecipitation coupled with high-throughput sequencing) method. Peaks: The four alignment files (linked as supplementary files on Sample records) were combined together for peak finding, as we found that most of the monomeric and dimeric tags are similarly distributed in the genome with high pearson correlation coefficient. The method to detect the peaks above gene-specific randomized background was similar to (Yeo et al., 2009) and described in the paper (Xue et al., 2009).

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Project description:The purpose of the study was to determine proteins that specially interact cytoplasmic PTB. PTB is an RNA binding protein that shuttles between the nucleus and cytoplasm. To identified proteins interacting with cytoplasmic PTB, nucleus localization signal was deleted (PTB ΔNLS-GFP) and overexpressed followed by IP and MS.

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Project description:MBNL1 regulates programmed postnatal switching between regenerative and differentiated cardiac states [scRNA-seq]

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Position-dependent alternative splicing activity revealed by global profiling of alternative splicing events regulated by PTB (HJAY)

Project description:To gain global insights into the role of the well-known repressive splicing regulator PTB we analyzed the consequences of PTB knockdown in HeLa cells using high-density oliogonucleotide splice-sensitive microarrays. The major class of identified PTB-regulated splicing event was PTB-repressed cassette exons, but there was also a substantial number of PTB-activated splicing events. PTB repressed and activated exons showed a distinct arrangement of motifs with pyrimidine-rich motif enrichment within and upstream of repressed exons, but downstream of activated exons. The N-terminal half of PTB was sufficient to activate splicing when recruited downstream of a PTB-activated exon. Moreover, insertion of an upstream pyrimidine tract was sufficient to convert a PTBactivated to a PTB-repressed exon. Our results demonstrate that PTB, an archetypal splicing repressor, has variable splicing activity that predictably depends upon its binding location with respect to target exons.

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