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. 2022 Feb 2;13(2):95. doi: 10.1038/s41419-022-04536-y

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© The Author(s) 2022

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. 2 — A RBM5-AS1 levels in MCF-7 and MDA-MB-231 cells under hypoxic condition for 24 and 48 h. B Transcriptional activity of the RBM5-AS1 promoter region. The pGL3-RBM5-AS1-promoter were transfected into MCF-7 and MDA-MB-231 cells, then luciferase activity was checked. C Binding sequences of top-10 ranked transcription factors on RBM5-AS1 promoter predicted in JASPAR website. D RUNX2 levels in 1104 breast cancer tissues (left) and 113 normal breast tissues (right) from TCGA database extracted using StarBase 3.0. E Spearman correlation analysis of the RUNX2 mRNA and RBM5-AS1 levels in 1104 human breast cancer tissues from TCGA project extracted using StarBase 3.0. F RUNX2 levels in 40 paired breast cancer tissues and paracancerous normal tissues. G Spearman correlation analysis of the correlation between RUNX2 and RBM5-AS1 levels in 40 paired breast cancer tissues. (H) RBM5-AS1 levels in control or RUNX2-overexpressing MCF-7 and MDA-MB-231 cells. (I) RUNX2 levels in MCF-7 and MDA-MB-231 cells under hypoxic condition for 24 and 48 h. J Putative RUNX2 binding sites on the promoter region of RBM5-AS1 (upper). Enrichment of RUNX2 on RBM5-AS1 promoter in MCF-7 and MDA-MB-231 cells detected by ChIP assays (lower left) and PCR analysis (lower right). Approximately one −42 to −28 bp fragment of the RBM5-AS1 promoter is responsible for RUNX2 binding. K Mutagenesis of the putative RUNX2 binding site (−50 to −10 bp) on the RBM5-AS1 promoter abolished the transcriptional activation of RBM5-AS1 by RUNX2 in MCF-7 cells. L Enrichment of RUNX2 on RBM5-AS1 promoter region in MCF-7 and MDA-MB-231 cells under hypoxic condition for 48 h detected by ChIP-qPCR assays (left) and PCR analysis (right). M RBM5-AS1 levels in si-NC or si-RUNX2 transfected MCF-7 and MDA-MB-231 cells under hypoxic condition for 0 or 48 h. N The relationship between RUNX2 expression and OS of breast cancer patients from GEPIA website. All data are shown as the mean ± SEM. *P < 0.05 and **P < 0.01 by two-tailed Student’s t-test.