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Expression data HEK293T cells transfected with either Ago2, Ago2 + miR-1, Ago2 + miR-124, mock

ABSTRACT: HEK293T cell expression data from four conditions: mock transfection, Ago2 transfection, Ago2 & miR-1 transfection, and Ago2 & miR-124 transfection. RNA was isolated directly from 10cm dishes using Trizol reagent and amplified. Set of arrays that are part of repeated experiments Compound Based Treatment: FLAG-Ago2 transfection Keywords: Biological Replicate

ORGANISM(S): Homo sapiens

PROVIDER: GSE11080 | GEO | 2008/04/08

SECONDARY ACCESSION(S): PRJNA108829

REPOSITORIES: GEO

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Expression data HEK293T cells transfected with either Ago2, Ago2 + miR-1, Ago2 + miR-124, mock

Project description:HEK293T cell expression data from four conditions: mock transfection, Ago2 transfection, Ago2 & miR-1 transfection, and Ago2 & miR-124 transfection. RNA was isolated directly from 10cm dishes using Trizol reagent and amplified. Set of arrays that are part of repeated experiments Compound Based Treatment: FLAG-Ago2 transfection Keywords: Biological Replicate Biological Replicate Using regression correlation

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Project description:Targetome of miR-124/miR-132 was enriched in HEK293T cells, a cell line which lacks the expression of these miRNA. The targetome was captured using miR-CLIP, a transfection-based double-purification crosslinking and immunoprecipitation (CLIP) method. Therefore HEK293T cells were transfected with miR-124/miR-132 miR-CLIP probes, which are trioxsalen, biotin bis-modified pre-miRNAs. Unspecific RNA-protein and trioxsalen mediated RNA-RNA crosslinking were induced by UV-irradiation before cell lysis. Total RNA (input samples) was enriched for AGO2-bound RNA using AGO2 immunoprecipitation, by depleting non-RISC associated RNAs. A second enrichment for specific targets of the transfected miRNA is performed using magnetic streptavidin-beads. This step relies on the interaction of the biotinylated probe, on which the RNA target is crosslinked, with the beads. The method was shown to deliver high-quality miRNA targetomes of both 5p and 3p miRNAs.

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mRNA abundance in HEK293T cells mock or miR-124 transfected

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Project description:Increasing evidence suggests that global downregulation of miRNA expression is a hallmark of human cancer, potentially due to defects in the miRNA processing machinery. In this study, we found that the protein expression of Argonaute 2 (AGO2), a key regulator of miRNA processing, was downregulated in colorectal cancer (CRC) tissues, which was also consistent with the findings of the Clinical Proteomic Tumor Analysis Consortium (CPTAC). Furthermore, the correlation between the levels of AGO2 and epithelial-mesenchymal transition (EMT) markers (E-cadherin and vimentin) indicated that reduced levels of AGO2 promoted EMT in CRC. Low expression of AGO2 was an indicator of a poor prognosis among CRC patients. Knockdown of AGO2 in CRC cells promoted migration, invasion and metastasis formation in vitro and in vivo but had no influence on proliferation. To provide detailed insight into the regulatory roles of AGO2, we performed integrated transcriptomic, quantitative proteomic and microRNA sequencing (miRNA-seq) analyses of AGO2 knockdown cells and the corresponding wild-type cells and identified neuropilin 1 (NRP1) as a new substrate of AGO2 via miR-185-3p. Our data provided evidence that knockdown of AGO2 resulted in a reduction of miR-185-3p expression, leading to the upregulation of the expression of NRP1, which is a direct target of miR-185-3p, and elevated CRC cell metastatic capacity. Inhibition of NRP1 or treatment with a miR-185-3p mimic successfully rescued the phenotypes of impaired AGO2, which suggested that therapeutically targeting the AGO2/miR-185-3p/NRP1 axis may be a potential treatment approach for CRC.

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Genome-wide hsa-miR-191 target profile [chip]

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