Abstract
PurposeDownregulation of miR-125b-1 is associated with poor prognosis in breast cancer patients. In this work we investigated the effect of histone modifications on the regulation of this gene promoter.Methods and resultsWe evaluated the enrichment of two histone modifications involved in gene repression, H3K9me3 and H3K27me3, on the miR-125b-1 promoter in two breast cancer cell lines, MCF7 (luminal A subtype) and MDA-MB-231 (triple-negative subtype), compared to the non-transformed breast cell line MCF10A. H3K27me3 and H3K9me3 were enriched in MCF7 and MDA-MB-231 cells, respectively. Next, we used an EZH2 inhibitor to examine the reactivation of miR-125b-1 in MCF7 cells and evaluated the transcriptional levels of pri-miR-125b-1 and mature miR-125b by qRT-PCR. pri-miRNA and mature miRNA transcripts were both increased after treatment of MCF7 cells with the EZH2 inhibitor, whereas no effect on miR-125b-1 expression levels was observed in MDA-MB-231 and MCF10A cells. We subsequently evaluated the effect of miR-125b-1 reactivation on the expression and protein levels of BAK1, a target of miR-125b. We observed 60 and 70 % decreases in the expression and protein levels of BAK1, respectively, compared to cells that were not treated with the EZH2 inhibitor. We over-expressed KDM4B/JMJD2B to reactivate this miRNA, resulting in a three-fold increase in miR-125b expression compared with the same cell line without KDM4B/JMJD2B over-expression.ConclusionThe miR-125b-1 is repressed by different epigenetic mechanisms depending on the breast cancer subtype and that miR-125b-1 reactivation specifically eliminates the effect of repressive histone modifications on the expression of an pro-apoptotic target.Electronic supplementary materialThe online version of this article (doi:10.1186/s40064-016-2475-z) contains supplementary material, which is available to authorized users.
Highlights
MicroRNAs are short single-stranded RNAs that could regulate gene expression at the post-transcriptional level. miRNAs are typically transcribed as primary transcripts that are subsequently matured in a multi-step biogenesis process to generate the functional form or mature miRNA (Lee and Dutta 2009)
In most breast cancer tumors, miR-125b-1 downregulation is associated with a poor prognosis. miR-125b-1 can be repressed by DNA methylation in the promoter region (Zhang et al 2011; Soto-Reyes et al 2012)
To determine miR-125b-1 expression levels, we evaluated pri-miRNA and mature miRNA levels in the breast cancer cell lines MCF7 and MDA-MB-231 by qRTPCR compared with the non-transformed breast cell line MCF10A. pri-miR-125b-1 levels in MCF7 and MDAMB-231 cells were reduced by 99 and 72 %, respectively, compared with MCF10A cells
Summary
MicroRNAs (miRNAs) are short single-stranded RNAs that could regulate gene expression at the post-transcriptional level. miRNAs are typically transcribed as primary transcripts (pri-miRNAs) that are subsequently matured in a multi-step biogenesis process to generate the functional form or mature miRNA (Lee and Dutta 2009). MicroRNAs (miRNAs) are short single-stranded RNAs that could regulate gene expression at the post-transcriptional level. Some miRNAs are associated with tumor suppressor or. MiR-125b-1 exhibits tumor suppressor activity in some types of cancer, including breast, ovarian and bladder cancer (Banzhaf-Strathmann 2014). Mir125b-1 may be involved in biological processes such as apoptosis, cell proliferation and cell migration because it regulates genes such as BAK1 (Zhou et al 2010), ERBB2 (Scott et al 2007) and ETS1 (Zhang et al 2011), respectively. In most breast cancer tumors, miR-125b-1 downregulation is associated with a poor prognosis. MiR-125b-1 can be repressed by DNA methylation in the promoter region (Zhang et al 2011; Soto-Reyes et al 2012). Promoters with intermediate CpG content are typically regulated by DNA methylation and histone modifications
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