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. 2017 Nov 27;7:16422. doi: 10.1038/s41598-017-16616-7

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© The Author(s) 2017

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 license, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons license 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 license, visit http://creativecommons.org/licenses/by/4.0/.

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EV71 promotes pri-miR-16-1 processing. (a) Schematic of the miR-16-1 gene and its promoter regions. Promoter 1 A and promoter 1B are two previously reported alternative promoters. The promoter (−1000–200) was from 1000 bp upstream to 200 bp downstream of oriented transcriptional start sites. (b,c) The EV71infection does not influence the transcription of pri-miR-16-1. Promoters 1A and 1B and −1000–100 regions were cloned to pGL3 vectors. Transcription experiments were performed with indicated reports or empty vectors together with an internal control, pRL-TK. Transfected 293 T cells were subjected to EV71 or mock infection for another 12 h. Then, reporter activity was determined by dual luciferase reporter assays. (d,e) Expression analysis of pri-miR-16-1 and DLEU2 in EV71-infected RD (d) and CCF-STTG1 cells (e). After EV71 virus infection for the indicated time, the expression levels of pri-miR-16-1 and DLEU2 were quantified by using qRT-PCR and the expression level of GAPDH as internal control. The relative expression levels of the target genes were calculated according to the 2^−∆∆Ct method. Data are representative of a minimum of three independent experiments, with each determination performed in triplicate.