Practical guide for <scp>circular RNA</scp> analysis: Steps, tips, and resources

Tsitsipatis, Dimitrios; Gorospe, Myriam

doi:10.1002/wrna.1633

Cited by 15 publications

(9 citation statements)

References 81 publications

(124 reference statements)

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“…Moreover, the possible functions of these circRNAs could be validated by visualizing their distribution and cellular localization by circFISH [ 66 ], or by verifying in which compartments are the circRNAs localized by circRNA expression profiling among the subcellular fractions [ 67 ]. In addition, the circRNA–miRNA interactions could be further examined by tagging a novel PRMT1 circRNA for affinity pulldown assays using a biotinylated antisense oligonucleotide that targets the back-splice junction and can be pulled down specifically with streptavidin-coated beads [ 68 ]. However, there lies a limitation of our study.…”

Section: Discussionmentioning

confidence: 99%

Identification of Novel Circular RNAs of the Human Protein Arginine Methyltransferase 1 (PRMT1) Gene, Expressed in Breast Cancer Cells

Papatsirou

Diamantopoulos

Katsaraki

et al. 2022

Genes

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Circular RNAs (circRNAs) constitute a type of RNA formed through back-splicing. In breast cancer, circRNAs are implicated in tumor onset and progression. Although histone methylation by PRMT1 is largely involved in breast cancer development and metastasis, the effect of circular transcripts deriving from this gene has not been examined. In this study, total RNA was extracted from four breast cancer cell lines and reversely transcribed using random hexamer primers. Next, first- and second-round PCRs were performed using gene-specific divergent primers. Sanger sequencing followed for the determination of the sequence of each novel PRMT1 circRNA. Lastly, bioinformatics analysis was conducted to predict the functions of the novel circRNAs. In total, nine novel circRNAs were identified, comprising both complete and truncated exons of the PRMT1 gene. Interestingly, we demonstrated that the back-splice junctions consist of novel splice sites of the PRMT1 exons. Moreover, the circRNA expression pattern differed among these four breast cancer cell lines. All the novel circRNAs are predicted to act as miRNA and/or protein sponges, while five circRNAs also possess an open reading frame. In summary, we described the complete sequence of nine novel circRNAs of the PRMT1 gene, comprising distinct back-splice junctions and probably having different molecular properties.

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Section: Discussionmentioning

confidence: 99%

Identification of Novel Circular RNAs of the Human Protein Arginine Methyltransferase 1 (PRMT1) Gene, Expressed in Breast Cancer Cells

Papatsirou

Diamantopoulos

Katsaraki

et al. 2022

Genes

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“…CircRNA is a type of non-coding RNA widely distributed in eukaryotes. CircRNA is highly stable as it is resistant to exonuclease- or ribonuclease-mediated degradation ( 13 ). It has been reported that circRNA is a key regulator of tumor gene expression and serves key roles in the signaling networks in oral squamous cell carcinoma, including the Wnt/β-catenin and PI3K/AKT pathways ( 14 ).…”

Section: Discussionmentioning

confidence: 99%

Suppression of circ_0008932 inhibits tumor growth and metastasis in osteosarcoma by targeting miR‑145‑5p

Cao

Shu

2021

Exp Ther Med

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Osteosarcoma (OS) is a common type of primary malignant tumor. Although the pathogenesis of OS has been extensively studied, the underlying molecular mechanisms have remained to be fully elucidated. Accumulating evidence has revealed that dysregulation of various circular RNAs (circRNAs) is associated with tumorigenesis and recent studies have indicated that circRNA circ_0008932 is aberrantly expressed in tumors. In the present study, the expression and detailed function of circ_0008932 in OS were elucidated. The levels of circ_0008932 in OS samples and cell lines were examined using reverse transcription-quantitative PCR. A cell model with circ_0008932 knockdown was generated using specific small interfering RNA (si-circ_0008932). Cell viability was determined by a Cell Counting Kit-8 assay, the cell migratory/invasive capacity was evaluated using Transwell assays and cell apoptosis was assessed by flow cytometry. The results suggested that circ_0008932 was upregulated in most primary OS tumors, suggesting that circ_0008932 is associated with the development of OS. In the in vitro assays, si-circ_0008932 inhibited the proliferation, migration and invasion of OS cells, while apoptosis was promoted. A luciferase reporter assay revealed that circ_0008932 may downregulate microRNA (miR)-145-5p through direct binding. Furthermore, the expression of miR-145-5p was negatively correlated with circ_0008932 levels in OS specimens. In addition, further functional studies indicated that miR-145-5p inhibitors eliminated the effects caused by si-circ_0008932 in OS cells. In comparison, the changes in the biological behavior of OS cells transfected with si-circ_0008932 were enhanced by miR-145-5p. In summary, circ_0008932 may be a novel oncogenic factor during the progression and development of OS by targeting miR-145-5p; more importantly, circ_0008932 may be a potential therapeutic target for OS.

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“…Circular RNAs (circRNAs) are a novel class of discovered ncRNAs with structural stability, formed by reverse splicing of the 3′-end and 5′-end of a chain-like pre-mRNA [ 43 , 44 ]. In general, circRNAs are formed via exon or intron cyclization, has no 3′-end or 5′-end of linear RNA, their structure is very stable and cannot be hydrolyzed by ordinary RNase; [ 45 – 47 ]. Circular RNA homeodomain-interacting protein kinase three (circHIPK3) is produced from exon two of HIPK3 and have been studied in many recent studies.…”

Section: Introductionmentioning

confidence: 99%

Mesenchymal stem cell-derived extracellular vesicles prevent the development of osteoarthritis via the circHIPK3/miR-124-3p/MYH9 axis

Liu

et al. 2021

J Nanobiotechnol

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Background Extracellular vesicles (EVs) secreted by mesenchymal stem cells (MSCs) may play a vital role in a variety of biological processes, including cartilage regeneration. However, few studies reported their potential in the development of osteoarthritis (OA) previously. In this study, we explored the biological roles and underlying mechanism of MSCs-EVs in OA. Results Co-culture experiments revealed that MSCs-EVs could promote the expression of collagen type II alpha 1 chain (COL2A1), SRY-box transcription factor 9 (SOX9) and Aggrecan while negatively regulate the expression of chondrocyte hypertrophy markers matrix metallopeptidase 13 (MMP-13) and RUNX family transcription factor 2 (Runx2) in mouse chondrocytes in the OA model. Besides, the results of cell experiments indicated that MSCs-EVs could notably weaken the suppression of chondrocyte proliferation, migration and the promotion of chondrocyte apoptosis via interleukin1β (IL-1β) induction. In addition, MSCs-circHIPK3-EVs (EVs derived from MSCs overexpressing circHIPK3) considerably improved IL-1β-induced chondrocyte injury. Mechanistically, we elucidated that circHIPK3 could directly bind to miR-124-3p and subsequently elevate the expression of the target gene MYH9. Conclusion The findings in our study demonstrated that EVs-circHIPK3 participated in MSCs-EVs-mediated chondrocyte proliferation and migration induction and in chondrocyte apoptosis inhibition via the miR-124-3p/MYH9 axis. This offers a promising novel cell-free therapy for treating OA. Graphic abstract

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Practical guide for circular RNA analysis: Steps, tips, and resources

Cited by 15 publications

References 81 publications

Identification of Novel Circular RNAs of the Human Protein Arginine Methyltransferase 1 (PRMT1) Gene, Expressed in Breast Cancer Cells

Identification of Novel Circular RNAs of the Human Protein Arginine Methyltransferase 1 (PRMT1) Gene, Expressed in Breast Cancer Cells

Suppression of circ_0008932 inhibits tumor growth and metastasis in osteosarcoma by targeting miR‑145‑5p

Mesenchymal stem cell-derived extracellular vesicles prevent the development of osteoarthritis via the circHIPK3/miR-124-3p/MYH9 axis

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