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miR-125b-1 is repressed by histone modifications in breast cancer cell lines.

Cisneros-Soberanis F, Andonegui MA, Herrera LA - Springerplus (2016)

Bottom Line: In this work we investigated the effect of histone modifications on the regulation of this gene promoter.H3K27me3 and H3K9me3 were enriched in MCF7 and MDA-MB-231 cells, respectively.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.

View Article: PubMed Central - PubMed

Affiliation: Unidad de Investigación Biomédica en Cáncer, Instituto Nacional de Cancerología, Instituto de Investigaciones Biomédicas, Universidad Nacional Autónoma de México, Avenida San Fernando 22, Mexico City, 14080 Mexico.

ABSTRACT

Purpose: Downregulation 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 results: We 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.

Conclusion: The 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.

No MeSH data available.


Related in: MedlinePlus

Elevated levels of miR-125b-1 affect the expression levels of BAK1. The target BAK1 was selected based on an evaluation of H3K27me3 enrichment in MCF7 cells by ENCODE (a). Next, we evaluated BAK1 expression levels by qRT-PCR (b). Finally, we determined protein levels by Western blotting (c, d). n = 3 *p > 0.05
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Fig5: Elevated levels of miR-125b-1 affect the expression levels of BAK1. The target BAK1 was selected based on an evaluation of H3K27me3 enrichment in MCF7 cells by ENCODE (a). Next, we evaluated BAK1 expression levels by qRT-PCR (b). Finally, we determined protein levels by Western blotting (c, d). n = 3 *p > 0.05

Mentions: mir-125b is involved in biological processes such as apoptosis, cell proliferation and cell migration-regulating genes such as BAK1 (Zhou et al. 2010), ERBB2 (Scott et al. 2007) and ETS1 (Zhang et al. 2011), respectively. We therefore sought to evaluate the effects of miR-125b-1 reactivation on the expression and protein levels of its targets. Using the ENCODE database, we evaluated the presence of H3K27me3 on some miR-125b target gene promoters in the MCF7 cell line (Additional file 3: Fig. 3). This histone modification was absent only in BAK1 (Fig. 5a). This analysis is important because GSK126 treatment of MCF7 cells may alter the expression and protein levels of target genes via an miR-125b-independent mechanism.Fig. 5


miR-125b-1 is repressed by histone modifications in breast cancer cell lines.

Cisneros-Soberanis F, Andonegui MA, Herrera LA - Springerplus (2016)

Elevated levels of miR-125b-1 affect the expression levels of BAK1. The target BAK1 was selected based on an evaluation of H3K27me3 enrichment in MCF7 cells by ENCODE (a). Next, we evaluated BAK1 expression levels by qRT-PCR (b). Finally, we determined protein levels by Western blotting (c, d). n = 3 *p > 0.05
© Copyright Policy - OpenAccess
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4930440&req=5

Fig5: Elevated levels of miR-125b-1 affect the expression levels of BAK1. The target BAK1 was selected based on an evaluation of H3K27me3 enrichment in MCF7 cells by ENCODE (a). Next, we evaluated BAK1 expression levels by qRT-PCR (b). Finally, we determined protein levels by Western blotting (c, d). n = 3 *p > 0.05
Mentions: mir-125b is involved in biological processes such as apoptosis, cell proliferation and cell migration-regulating genes such as BAK1 (Zhou et al. 2010), ERBB2 (Scott et al. 2007) and ETS1 (Zhang et al. 2011), respectively. We therefore sought to evaluate the effects of miR-125b-1 reactivation on the expression and protein levels of its targets. Using the ENCODE database, we evaluated the presence of H3K27me3 on some miR-125b target gene promoters in the MCF7 cell line (Additional file 3: Fig. 3). This histone modification was absent only in BAK1 (Fig. 5a). This analysis is important because GSK126 treatment of MCF7 cells may alter the expression and protein levels of target genes via an miR-125b-independent mechanism.Fig. 5

Bottom Line: In this work we investigated the effect of histone modifications on the regulation of this gene promoter.H3K27me3 and H3K9me3 were enriched in MCF7 and MDA-MB-231 cells, respectively.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.

View Article: PubMed Central - PubMed

Affiliation: Unidad de Investigación Biomédica en Cáncer, Instituto Nacional de Cancerología, Instituto de Investigaciones Biomédicas, Universidad Nacional Autónoma de México, Avenida San Fernando 22, Mexico City, 14080 Mexico.

ABSTRACT

Purpose: Downregulation 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 results: We 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.

Conclusion: The 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.

No MeSH data available.


Related in: MedlinePlus