CRISPR-Assisted Detection of RNA-Protein Interactions in Living Cells
Ontology highlight
ABSTRACT:
ORGANISM(S): Mus musculus musculus x Mus musculus castaneus Homo sapiens
PROVIDER: GSE137556 | GEO | 2020/06/21
REPOSITORIES: GEO
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Project description:We develop a CRISPR-Assisted RNA-Protein Interaction Detection method (CARPID), which leverages CRISPR/CasRx-based RNA targeting and proximity labeling to identify binding proteins of specific lncRNA in the native cellular context.
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Project description:Sustained hepatic inflammation can contribute to cancer initiation and progression via tumor-initiating cell expansion, but underlying mechanisms remains unknown. Expression profiles of lncRNAs/mRNAs were measured in normal, chronic hepatitis, cirrhotic and cancerous livers. We found abundant disease-related lncRNAs/mRNAs deregulated across different stages of inflammation-triggered liver disease and refined a transformation gene signature to distinguish pathological liver tissues. Amongst this signature, a conserved lncRNA DANCR was silenced in normal adult liver, but overexpressed in fetal and cancerous livers. Remarkably, increased DANCR significantly correlates with poor prognosis in multiple-center cohorts and is directly induced by inflammatory pathways including NF-κB and STAT3. DANCR could suppress cell differentiation and drive expansion of tumor-initiating cells, leading to chemoresistance. Moreover, in vitro and in vivo inhibition confirms the significance of DANCR as a therapeutic target when combined with other chemotherapy. We illustrate the role of DANCR relies on the regulation of CTNNB1 in a novel miRNA-blocking manner. Our studies reveal the expression of lncRNAs/mRNAs in normal and pathological livers and suggest the importance of oncofetal lncRNA DANCR in inflammation-induced malignant transformation, offering a potential prognostic marker and a therapeutic target for HCC.
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Project description:We report that long noncoding RNAs contribute to transcription and developmental process. Thousands of lncRNAs have been identified in the whole genome, and tend to located closely to protein-coding genes. To study position relationship between lncRNA and protein-coding genes, we classified all of lncRNA to several subgroups based on the genome position with their coding neighbors. XH, the head to head subgroup is associated with transcription and development in GO analysis. Here, we knockdown serveral XH lncRNA by shRNA in embryonic stem cells and induce nondirectional differnetiation by removing LIF or neural differnetiation by RA. Knockdown of XH lncRNAs led to uniform downregulation of nearby coding genes, and form regulatory circuits with its nearby coding genes to fine-tune embryonic lineage development. In addition, we also knockout one lncRNA-Evx1as and its nearby protein-coding gene-EVX1 by CRISPR, and get similar results as knockdown.We propose that XH lncRNA may function primarily as 'cis-regulators' of the expression of nearby protein-coding genes, and tend to participate in transcriptional or development regulations as their coding neighbors. All RNA-seq(s) were designed to reveal the differentially expressed genes between wild-type and XH lncRNA knockdown/knockout ESCs during differentiation.
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