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. 2019 Dec 10;8(12):73. doi: 10.1038/s41389-019-0182-7

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© The Author(s) 2019

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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Fig. 6 — a Western blots showing the expression of PTBP1 in BT549 cells after transfection with PTBP1 siRNA or scrambled siRNA for 48 h. b qRT-PCR indicating the relative levels of MACC1-AS1 in PTBP1 knockdown and control cells. Data are presented as means ± SD from three independent experiments. **P < 0.01 as determined by one-way ANOVA assay. c Levels of MACC1-AS1 in BT549 and BT549/PTBP1-siRNA cells were analyzed after treatment with actinomycin D for 12 h. Relative MACC1-AS1 levels were determined by qRT-PCR. Data are presented as means ± SD from three independent experiments. **P < 0.01 as determined by Student’s t test. d MDA-MB-231/MACC1-AS1-MS2 cells were transfected with PTBP1 siRNA for 48 h. RNA agarose gel electrophoresis and western blots indicated downregulation of PTBP1 mRNA and protein. GAPDH was used as a control. e Cells transfected as in (d) were subjected qRT-PCR to detect the levels of individual miRNAs associated with MACC1-AS1 in pulldown precipitates. **P < 0.01 and *P < 0.05 as determined by Student’s t test. f MDA-MB-231 cells were transfected with constructs expressing MACC1-AS1-MS2 and MACC1-AS1(m)-MS2 for 72 h. RNA agarose gel electrophoresis showed the precipitated MACC1-AS1-MS2 and MACC1-AS1(m)-MS2 in cell extracts. g Cells transfected as in (f) were subjected qRT-PCR, showing that levels of MACC1-AS1-associated miRNAs were greatly reduced when the biding site for PTBP1 was mutated. **P < 0.01 as determined by Student’s t test.