hsa-miR-20b-5p and hsa-miR-363-3p Affect Expression of PTEN and BIM Tumor Suppressor Genes and Modulate Survival of T-ALL Cells In Vitro

Drobna, Monika; Szarzyńska, Bronisława; Jaksik, Roman; Sędek, Łukasz; Kuchmiy, Anna; Taghon, Tom; Vlierberghe, Pieter Van; Szczepański, Tomasz; Witt, Michał; Dawidowska, Małgorzata

doi:10.3390/cells9051137

Cited by 25 publications

(22 citation statements)

References 59 publications

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“…Regarding miRNAs upregulated following EBOV infection ( Table 1 and Table 2 ), it is interesting to note the existence of strain-specific miRNAs that can be detected during the early stage of infection and that may serve as precocious biomarkers, and potentially contribute either positively or negatively to the virulence and pathogenicity of the EBOV strains. For example, Mayinga seemed to correlate in a specific manner with miR-363-3p (belonging to miR-106-363 cluster), a regulator of apoptosis and proliferation in various cell lines, including those derived from the liver [ 97 , 98 , 99 ]. The Makona strain was associated with miR-132-3p which is known to promote H1N1 Influenza A Virus replication by suppressing type I interferon response through targeting Interferon Regulatory Factor 1 (IRF1) [ 100 ].…”

Section: Discussionmentioning

confidence: 99%

Altered microRNA Transcriptome in Cultured Human Liver Cells upon Infection with Ebola Virus

Diallo

Laffont

et al. 2021

IJMS

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Ebola virus (EBOV) is a virulent pathogen, notorious for inducing life-threatening hemorrhagic fever, that has been responsible for several outbreaks in Africa and remains a public health threat. Yet, its pathogenesis is still not completely understood. Although there have been numerous studies on host transcriptional response to EBOV, with an emphasis on the clinical features, the impact of EBOV infection on post-transcriptional regulatory elements, such as microRNAs (miRNAs), remains largely unexplored. MiRNAs are involved in inflammation and immunity and are believed to be important modulators of the host response to viral infection. Here, we have used small RNA sequencing (sRNA-Seq), qPCR and functional analyses to obtain the first comparative miRNA transcriptome (miRNome) of a human liver cell line (Huh7) infected with one of the following three EBOV strains: Mayinga (responsible for the first Zaire outbreak in 1976), Makona (responsible for the West Africa outbreak in 2013–2016) and the epizootic Reston (presumably innocuous to humans). Our results highlight specific miRNA-based immunity pathways and substantial differences between the strains beyond their clinical manifestation and pathogenicity. These analyses shed new light into the molecular signature of liver cells upon EBOV infection and reveal new insights into miRNA-based virus attack and host defense strategy.

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Section: Discussionmentioning

confidence: 99%

Altered microRNA Transcriptome in Cultured Human Liver Cells upon Infection with Ebola Virus

Diallo

Laffont

et al. 2021

IJMS

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“…Mohamed et al reported that miR ‐ 363 ‐ 3p increased chemoresistance of an ovarian cancer line to taxane 16 . Dorbna et al 17 demonstrated that upregulated miR ‐ 363 ‐ 3p prevented apoptosis and promoted growth of leukemic cells in vitro . In contrast, miR ‐ 363 was shown to be a tumour suppressor in lung cancer, osteosarcoma, and colorectal cancer 18‐20 .…”

Section: Introductionmentioning

confidence: 99%

miR‐363‐3p induces EMT via the Wnt/β‐catenin pathway in glioma cells by targeting CELF2

Fan

Su²,

Song

et al. 2021

J Cellular Molecular Medi

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In our previous study, we reported that CELF2 has a tumour‐suppressive function in glioma. Here, we performed additional experiments to elucidate better its role in cancer. The expression profile of CELF2 was analysed by the GEPIA database, and Kaplan–Meier curves were used to evaluate the overall survival rates. Four different online databases were used to predict miRNAs targeting CELF2, and the luciferase assay was performed to identify the binding site. The biological effects of miR‐363‐3p and CELF2 were also investigated in vitro using MTT, Transwell, and flow cytometry assays. Western blotting, qPCR, and TOP/FOP flash dual‐luciferase assays were performed to investigate the impact of miR‐363‐3p and CELF2 on epithelial‐to‐mesenchymal transition (EMT) and the Wnt/β‐catenin pathway. The effect of miR‐363‐3p was also tested in vivo using a xenograft mouse model. We observed an abnormal expression pattern of CELF2 in glioma cells, and higher CELF2 expression correlated with better prognosis. We identified miR‐363‐3p as an upstream regulator of CELF2 and confirmed its direct binding to the 3′‐untranslated region of CELF2. Cell function experiments showed that miR‐363‐3p affected multiple aspects of glioma cells. Suppressing miR‐363‐3p expression inhibited glioma cell proliferation and invasion, as well as promoted cell death via attenuating EMT and blocking the Wnt/β‐catenin pathway. These effects could be abolished by the downregulation of CELF2. Treatment with ASO‐miR‐363‐3p decreased tumour size and weight in nude mice. In conclusion, miR‐363‐3p induced the EMT, which resulted in increased migration and invasion and reduced apoptosis in glioma cell lines, via the Wnt/β‐catenin pathway by targeting CELF2.

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“…To further assess the effectiveness of CRISPRi-mediated repression of miRNAs, we evaluated the level of selected target genes for miRNAs of interest. We chose BCL2L11 (also known as BIM ), which we have previously shown as a common target for miR-20b-5p and miR-363-3p in T-ALL cells in vitro 38 . Since BCL2L11 can potentially be targeted by miRNAs from miR-17, miR-19 and miR-92 seed families, we analyzed its expression upon CRISPRi-mediated silencing of mir-106a-363 cluster (Supplementary Figs.…”

Section: Resultsmentioning

confidence: 99%

“…Inhibitors were used in the final concentration of 100 nM. Cells were electroporated with the use of the Neon Electroporation System (Thermo Fisher Scientific) as described previously 38 .…”

Section: Methodsmentioning

confidence: 99%

CRISPRi for specific inhibition of miRNA clusters and miRNAs with high sequence homology

Drobna-Śledzińska

Maćkowska-Maślak

Jaksik

et al. 2022

Sci Rep

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AbstractmiRNAs form a class of noncoding RNAs, involved in post-transcriptional regulation of gene expression, broadly studied for their involvement in physiological and pathological context. Inhibition of mature miRNA transcripts, commonly used in miRNA loss-of-function experiments, may not be specific in case of miRNAs with high sequence homology, e.g. miRNAs from the same seed family. Phenotypic effects of miRNA repression might be biased by the repression of highly similar miRNAs. Another challenge is simultaneous inhibition of multiple miRNAs encoded within policistronic clusters, potentially co-regulating common biological processes. To elucidate roles of miRNA clusters and miRNAs with high sequence homology, it is of key importance to selectively repress only the miRNAs of interest. Targeting miRNAs on genomic level with CRISPR/dCas9-based methods is an attractive alternative to blocking mature miRNAs. Yet, so far no clear guidelines on the design of CRISPR inhibition (CRISPRi) experiments, specifically for miRNA repression, have been proposed. To address this need, here we propose a strategy for effective inhibition of miRNAs and miRNA clusters using CRISPRi. We provide clues on how to approach the challenges in using CRISPR/dCas in miRNA studies, which include prediction of miRNA transcription start sites (TSSs) and the design of single guide RNAs (sgRNAs). The strategy implements three TSS prediction online tools, dedicated specifically for miRNAs: miRStart, FANTOM 5 miRNA atlas, DIANA-miRGen, and CRISPOR tool for sgRNAs design; it includes testing and selection of optimal sgRNAs. We demonstrate that compared to siRNA/shRNA-based miRNA silencing, CRISPRi improves the repression specificity for miRNAs with highly similar sequence and contribute to higher uniformity of the effects of silencing the whole miRNA clusters. This strategy may be adapted for CRISPR-mediated activation (CRISPRa) of miRNA expression.

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hsa-miR-20b-5p and hsa-miR-363-3p Affect Expression of PTEN and BIM Tumor Suppressor Genes and Modulate Survival of T-ALL Cells In Vitro

Cited by 25 publications

References 59 publications

Altered microRNA Transcriptome in Cultured Human Liver Cells upon Infection with Ebola Virus

Altered microRNA Transcriptome in Cultured Human Liver Cells upon Infection with Ebola Virus

miR‐363‐3p induces EMT via the Wnt/β‐catenin pathway in glioma cells by targeting CELF2

CRISPRi for specific inhibition of miRNA clusters and miRNAs with high sequence homology

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