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Determination of a comprehensive alternative splicing regulatory network and the combinatorial regulation by key factors during Epithelial-to-Mesenchymal Transition [ESRP KD]

ABSTRACT: Epithelial specific splicing regulatory protein 1 and 2 (ESRP1 and ESRP2) are important regulators of alternative splicing during EMT. To study the alternative splicing events regulated by ESRP1/2 at a genome wide scale, we used lentiviral shRNAs to knockdown ESRP1/2 in H358 cells and performed RNA-seq in biological triplicates. We used lentiviral based shRNAs targeting ESRP1 and ESRP2 to knockdown both regulators in human H358 cells. We harvested total RNA and protein from ESRP1/2 knockdown and control knockdown in biological triplicates. We made cDNA libraries for each replicate and subjected them to RNA-seq.

ORGANISM(S): Homo sapiens

SUBMITTER: Yi Xing

PROVIDER: E-GEOD-75489 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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Determination of a comprehensive alternative splicing regulatory network and the combinatorial regulation by key factors during Epithelial-to-Mesenchymal Transition [ESRP KD]

Project description:Epithelial specific splicing regulatory protein 1 and 2 (ESRP1 and ESRP2) are important regulators of alternative splicing during EMT. To study the alternative splicing events regulated by ESRP1/2 at a genome wide scale, we used lentiviral shRNAs to knockdown ESRP1/2 in H358 cells and performed RNA-seq in biological triplicates.

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Determination of a comprehensive alternative splicing regulatory network and the combinatorial regulation by key factors during Epithelial-to-Mesenchymal Transition [RBM47 KD]

Project description:RBM47 is a RNA binding protein that is known to regulate alternative splicing. To study the genome wide regulatory role of RBM47 in alternative splicing and determine the potential function of RBM47 during EMT, we used lentiviral shRNAs to knockdown RBM47 in H358 cells and performed RNA-seq in biological triplicates. We used lentiviral based shRNAs to knockdown RBM47 in human H358 cells. We harvested total RNA and protein from RBM47 knockdown and control knockdown in biological triplicates. We made cDNA libraries for each replicate and subjected them to RNA-seq.

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Knockout mice reveal an essential role for Epithelial splicing regulatory proteins (Esrps) in mammalian development and epithelial splicing in vivo

Project description:Tissue- and cell-type specific regulators of alternative splicing (AS) are an essential layer of posttranscriptional gene regulation necessary for normal cellular function, patterning, and development. Here we report the Epithelial splicing regulatory proteins (Esrps) are required for patterning of multiple organs, with loss of both paralogs, Esrp1 and Esrp2, resulting in increasingly severe phenotypes. Global profiling of the Esrp splicing regulatory network from total epidermis revealed varied splicing sensitivity of Esrp targets upon loss of Esrp1 or double knockout. This may explain the progressive phenotypes seen in Esrp knockout mice, and these mice provide a unique genetic tool to evaluate functional consequences of epithelial splicing events in vivo. RNA from purified total epidermis (basal keratinocyte layer to cornified layer) of E18.5 mouse embryos were harvested by Trizol extraction. (Esrp1+/+, Esrp2-/- (n=2), Esrp1-/-, Esrp2+/+ (n=3), Esrp1-/-, Esrp2+/- (n=2), and Esrp1-/-, Esrp2-/- (n=2). 1 ug of total RNA was used for for RNA-seq library preparation using the TruSeq™ Stranded mRNA LT Sample Prep Kit (Illumina). 100x2 bp paired-end RNA-seq reads were generated on a HiSeq 2000 sequencer.

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The epithelial splicing factors ESRP1 and ESRP2 positively and negatively regulate alternative splicing events

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