Abstract
XAF1 is a tumour suppressor gene that compromises cell viability by modulating different cellular events such as mitosis, cell cycle progression and apoptosis. In cancer, the XAF1 gene is commonly silenced by CpG-dinucleotide hypermethylation of its promoter. DNA demethylating agents induce transcriptional reactivation of XAF1, sensitizing cancer cells to therapy. The molecular mechanisms that mediate promoter CpG methylation have not been previously studied. Here, we demonstrate that CTCF interacts with the XAF1 promoter in vivo in a methylation-sensitive manner. By transgene assays, we demonstrate that CTCF mediates the open-chromatin configuration of the XAF1 promoter, inhibiting both CpG-dinucleotide methylation and repressive histone posttranslational modifications. In addition, the absence of CTCF in the XAF1 promoter inhibits transcriptional activation induced by well-known apoptosis activators. We report for the first time that epigenetic silencing of the XAF1 gene is a consequence of the loss of CTCF binding.
Highlights
We demonstrated that CTCF directly regulates XAF1 expression by binding to a methylation-sensitive CTCF-binding site in its promoter
As expected based on previous reports showing that XAF1 promoter is hypermethylated in cancer[9,12,17], here, pre-exposure to demethylating agents increased the transcriptional activation of XAF1 in basal conditions (Supplementary Fig.1a)
CpG methylation in the XAF1 promoter represents the main epigenetic mechanism involved in XAF1 silencing and, in resistance against apoptosis
Summary
We demonstrated that CTCF directly regulates XAF1 expression by binding to a methylation-sensitive CTCF-binding site in its promoter. The absence of CTCF promotes epigenetic silencing of the XAF1 promoter by both accelerated CpG-dinucleotide methylation and the transition from active to repressive HPMs. Importantly, in cancer cell lines, the lack of CTCF regulation on the XAF1 promoter via methylation on its cognate binding site partially blocks its transcriptional responsiveness to two well-known transcriptional activators, TNF-α or IFN-α. In cancer cell lines, the lack of CTCF regulation on the XAF1 promoter via methylation on its cognate binding site partially blocks its transcriptional responsiveness to two well-known transcriptional activators, TNF-α or IFN-α These findings uncover for the first time an epigenetic mechanism involved in establishing the repressive configuration of the XAF1 promoter and, transcriptional unresponsiveness
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