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. 2020 Oct 7;10:16755. doi: 10.1038/s41598-020-73785-8

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© The Author(s) 2020

Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons licence, and indicate if changes were made. The images or other third party material in this article are included in the article's Creative Commons licence, unless indicated otherwise in a credit line to the material. If material is not included in the article's Creative Commons licence and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this licence, visit http://creativecommons.org/licenses/by/4.0/.

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E2F2 is a direct target of miR-522-3p and its expression is upregulated in paclitaxel-resistant cells. (A) The list of downregulated genes in the SKOV3ip1-PR and HeyA8-PR cells transfected with miR-522-3p, relative to the cells transfected with control miRNA (red indicates the downregulated genes captured by a biotinylated miR-522-3p mimic based on Ref.¹⁶). (B) The quantitative reverse transcription polymerase chain reaction findings on the relative expression of E2F2 and p27 (vs. control miRNA) in paclitaxel-resistant ovarian cancer cell lines transfected with miR-522-3p. (C) Schematic illustration of the predicted E2F2 3′-untranslated region (UTR)-binding site of miR-522-3p. (D) Paclitaxel-resistant cells were co-transfected with miRNA precursor (miR-522-3p or control-miR), a luciferase reporter vector containing the wildtype or mutant 3′-UTR of E2F2, and the Renilla luciferase control vector. After 24 h of incubation, the luciferase activity level was measured (normalized to Renilla activity). Data are represented as mean ± standard deviation and were obtained from three independent experiments. (E) Western blot results for E2F2 expression in the parental cells and paclitaxel-resistant cells. Lamin B1 was used as a loading control (upper). Densitometric ratio of the expression of E2F2 / lamin B1 (lower). (F) Western blot results for E2F2 expression in paclitaxel-resistant cells transfected with miR-522-3p or control-miR (left). Densitometric ratio of the expression of E2F2 / lamin B1 (right). (G) Correlation plot extracted from seven ovarian cancer cell lines deposited in the NCI60 microarray data sets. x axis; the average transcript intensity z score of miR-522-3p, y axis; that of miR-E2F2. *P < 0.05, ***P < 0.001, n.s. not significant.