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Efficient, specific and combinatorial control of endogenous exon splicing with dCasRx-RBM25

ABSTRACT: Efficient, specific and combinatorial control of endogenous exon splicing with dCasRx-RBM25

PROVIDER: PRJNA1020835 | ENA |

REPOSITORIES: ENA

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			Action	DRS
	SRR26170180_1.fastq.gz	Fastqsanger.gz
	SRR26170180_2.fastq.gz	Fastqsanger.gz
	SRR26170181_1.fastq.gz	Fastqsanger.gz
	SRR26170181_2.fastq.gz	Fastqsanger.gz
	SRR26170182_1.fastq.gz	Fastqsanger.gz

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Efficient, specific and combinatorial control of endogenous exon splicing with dCasRx-RBM25

Project description:Efficient targeted control of exon splicing is a major goal of functional genomic and therapeutic applications. Guide RNA-directed, deactivated (d)Cas CRISPR enzymes fused to splicing effectors represent a promising strategy due to the flexibility and presumed specificity of these systems. However, efficient, selective, and generalizable activation of targeted endogenous exons using this approach has not been reported. Here, we identify dCasRx-RBM25 as a potent activator of exons by screening over 300 dCasRx splicing factor fusions tethered to splicing reporters. dCasRx-RBM25 also strongly activates splicing of endogenous alternative exons, when recruited to downstream intron sequences using single guide RNAs. In transcriptome wide analyses we observe a high degree of specificity of dCasRx-RBM25 for endogenous exon targeting. We further leverage the guide array-processing activity of dCasRx to simultaneously target multiple endogenous exons for activation and repression by dCasRx-RBM25. Our results pave the way for versatile exon-resolution functional assays and splicing-directed therapeutic applications.

2024-09-19 | GSE243979 | GEO

Identification of Rbm25-binding RNAs in bone-marrow-derived macrophages by RIP-seq

Project description:To identify the potential mRNAs that bound by Rbm25, which might be the direct target of Rbm25 for alternative splicing in murine macrophages.

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Project description:Cancer sequencing studies have implicated regulators of pre-mRNA splicing as important disease determinants in Acute Myeloid Leukemia (AML), but the underlying mechanisms have remained elusive. We hypothesized that “non-mutated” splicing regulators may also play a role in AML biology and therefore conducted an in vivo shRNA screen in a mouse model of CEBPA mutant AML. This led to the identification of the splicing regulator RBM25 as a novel tumor suppressor, and down-regulation of RBM25 increased proliferation and decreased apoptosis in human leukemic cell lines. Mechanistically, we could show that RBM25 controlled the splicing of key genes, including those encoding the apoptotic regulator BCL-x and the MYC inhibitor BIN1. Specifically, we demonstrated that RBM25 acts as a regulator of MYC activity and sensitizes cells to increased MYC levels. This mechanism also appears to be operative in human AML patients where RBM25 levels correlative inversely with MYC activity and clinical outcome.

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