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Disruption of CTCF at the miR-125b1 locus in gynecological cancers
Ernesto Soto-Reyes1, Rodrigo González-Barrios, Fernanda Cisneros-Soberanis
1Unidad de Investigación Biomédica en Cáncer, Instituto Nacional de Cancerología (INCan), Universidad Nacional Autónoma de México (UNAM), México, DF, México.
Background:
In cancer cells, transcriptional gene silencing has been associated with genetic and epigenetic defects. The disruption of DNA methylation patterns and covalent histone marks has been associated with cancer development. Until recently, microRNA (miRNA) gene silencing was not well understood. In particular, miR-125b1 has been suggested to be an miRNA with tumor suppressor activity, and it has been shown to be deregulated in various human cancers. In the present study, we evaluated the DNA methylation at the CpG island proximal to the transcription start site of miR-125b1 in cancer cell lines as well as in normal tissues and gynecological tumor samples. In addition, we analyzed the association of CTCF and covalent histone modifications at the miR-125b1 locus.
Methods:
To assess the DNA methylation status of the miR-125b1, genomic DNA was transformed with sodium bisulfite, and then PCR-amplified with modified primers and sequenced. The miR-125b1 gene expression was analyzed by qRT-PCR using U6 as a control for constitutive gene expression. CTCF repressive histone marks abundance was evaluated by chromatin immunoprecipitation assays.
Results:
The disruption of CTCF in breast cancer cells correlated with the incorporation of repressive histone marks such H3K9me3 and H3K27me3 as well as with aberrant DNA methylation patterns. To determine the effect of DNA methylation at the CpG island of miR-125b1 on the expression of this gene, we performed a qRT-PCR assay. We observed a significant reduction on the expression of miR-125b1 in cancer cells in comparison with controls, suggesting that DNA methylation at the CpG island might reduce miR-125b1 expression. These effects were observed in other gynecological cancers, including ovarian and cervical tumors.
Conclusions:
A reduction of miR-125b1 expression in cancers, correlated with methylation, repressive histone marks and loss of CTCF binding at the promoter region.
Insights
microRNA-125b1 (miR-125b1) expression is reduced in various cancers due to DNA methylation, repressive histone marks, and loss of CTCF binding. This epigenetic silencing impacts tumor suppressor activity.
Area of Science:
- Molecular Biology
- Epigenetics
- Cancer Research
Background:
- Transcriptional gene silencing in cancer involves genetic and epigenetic defects, including altered DNA methylation and histone modifications.
- MicroRNA (miRNA) gene silencing, particularly of tumor suppressor miR-125b1, is increasingly recognized as a factor in cancer development.
- miR-125b1 is deregulated in various human cancers, suggesting its role in tumorigenesis.
Purpose of the Study:
- To investigate DNA methylation patterns at the miR-125b1 CpG island in cancer cell lines and gynecological tumors.
- To analyze the association between CTCF binding, histone modifications, and miR-125b1 expression in cancer.
- To understand the epigenetic mechanisms underlying miR-125b1 deregulation in cancer.
Main Methods:
- DNA methylation analysis using bisulfite sequencing of the miR-125b1 CpG island.
- Quantitative reverse transcription PCR (qRT-PCR) to measure miR-125b1 gene expression.
- Chromatin immunoprecipitation assays to evaluate CTCF and repressive histone mark (H3K9me3, H3K27me3) abundance.
Main Results:
- Aberrant DNA methylation patterns and incorporation of repressive histone marks (H3K9me3, H3K27me3) were observed in cancer cells, correlating with CTCF disruption.
- A significant reduction in miR-125b1 expression was detected in cancer cells compared to controls.
- These epigenetic alterations and reduced expression were also found in ovarian and cervical tumors.
Conclusions:
- Reduced miR-125b1 expression in cancers is linked to promoter DNA methylation, repressive histone marks, and loss of CTCF binding.
- Epigenetic silencing of miR-125b1 contributes to its reduced expression in various gynecological cancers.
- These findings highlight the role of epigenetic dysregulation in the silencing of tumor suppressor miRNAs like miR-125b1.
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