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ShRNA-based miRNA overexpression results in significant off-target effects caused by isomiR production [miRNA-seq PC-3]

ABSTRACT: shRNA-based miRNA overexpression results in significant off-target effects caused by isomiR production [miRNA-seq PC-3]

PROVIDER: PRJNA1101081 | ENA |

REPOSITORIES: ENA

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			Action	DRS
	SRR28709477.fastq.gz	Fastqsanger.gz
	SRR28709478.fastq.gz	Fastqsanger.gz

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shRNA-based miRNA overexpression results in significant off-target effects caused by isomiR production [miRNA-seq]

Project description:shRNA-based miRNA overexpression results in significant off-target effects caused by isomiR production [miRNA-seq]

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shRNA-based miRNA overexpression results in significant off-target effects caused by isomiR production

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shRNA-based miRNA overexpression results in significant off-target effects caused by isomiR production [mRNA-seq]

Project description:shRNA-based miRNA overexpression results in significant off-target effects caused by isomiR production [mRNA-seq]

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shRNA-based miRNA overexpression results in significant off-target effects caused by isomiR production [miRNA-seq PC-3]

Project description:miRNA overexpression strategy based on RNA Polymerase (Pol) III mediated expression of short hairpin RNA (shRNA) coding particular miRNA is widely used for miRNA functional studies. For some miRNAs, e.g., encoded in the genome as a part of a polycistronic miRNA cluster, it is most likely the only way for their individual stable overexpression. Here we have revealed that extra \texttt{U} residues (up to five) added Pol III at the 3$'$-end of the transcribed shRNA associated with a shift in the Dicer cleavage position of the shRNA. This results in the undesirable formation of 5$'$-end miRNA isoforms (5$'$-isomiRs), which have a significantly altered seed region compared to the initially encoded canonical hsa-miR-93-5p. The predominant expression of 5$'$-isomiRs without three or four first nucleotides instead of the canonical isoform could be disclosed based on miRNA-Seq analysis. Moreover, mRNA sequencing data showed that the 5$'$-isomiRs of hsa-miR-93-5p presumably regulate their own mRNA targets. Thus, omitting miRNA-Seq analysis may lead to erroneous conclusions regarding revealed mRNA targets and possible molecular mechanisms in which studied miRNA is involved.

2024-04-17 | GSE264166 | GEO

shRNA-based miRNA overexpression results in significant off-target effects caused by isomiR production [DU-145 cells]

Project description:shRNA-based miRNA overexpression results in significant off-target effects caused by isomiR production [DU-145 cells]

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shRNA-based miRNA overexpression results in significant off-target effects caused by isomiR production [miRNA-seq]

2023-08-22 | GSE241301 | GEO

shRNA-based miRNA overexpression results in significant off-target effects caused by isomiR production [mRNA-seq]

2023-08-22 | GSE241300 | GEO

shRNA-based miRNA overexpression results in significant off-target effects caused by isomiR production [DU-145 cells]

2023-11-17 | GSE247987 | GEO

shRNA-based miRNA overexpression results in significant off-target effects caused by isomiR production

Project description:This SuperSeries is composed of the SubSeries listed below.

2023-08-21 | GSE241303 | GEO

Protein profiles following miRNA-23c and -4328 overexpression in prostate cancer cell lines, PC-3 and 22Rv1

Project description:The 22Rv1 and PC-3 cells were transduced with shMIMIC human lentiviral vectors (Horizon Discovery, Cambridge, UK) to stably overexpress miRNA-23c or -4328. The SMARTvector Non-Targeting Control (NTC) was expressed to serve as a negative control. The vectors contained a turbo green fluorescent protein (turboGFP) and a puromycin resistance gene cassette. After transduction, cells were cultured in a medium supplemented with 5 µg/mL puromycin (Takara Bio, Tokyo, Japan) for antibiotic selection. Overexpression was confirmed by monitoring the turboGFP by fluorescence microscopy and by RT-qPCR analysis of miRNA-23c and -4328 levels. Relative protein quantification was performed to compare protein expression in 22Rv1 and PC-3 single cell clones overexpressing miRNA-23c and -4328, compared to corresponding NTC cells. Single cell clones overexpressing either miRNA-23c (n = 3), -4328 (n = 3) or NTC (n = 3) were analyzed in triplicate.

2023-04-24 | PXD041683 | Pride

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