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Epigenetic silencing involving the aberrant DNA methylation of promoter-associated CpG islands is one mechanism leading to the inactivation of tumor suppressor genes in human cancers. However, the molecular mechanisms underlying this event remains poorly understood. TMS1/ASC is a novel proapoptotic signaling factor that is subject to epigenetic silencing in human breast and other cancers. The TMS1 promoter is embedded within a CpG island that is unmethylated in normal cells and is spanned by three DNase I-hypersensitive sites (HS). Silencing of TMS1 in cancer cells is accompanied by local alterations in histone modification, remodeling of the HS, and hypermethylation of DNA. In this study, we probed the functional significance of the CpG island-specific HS. We identified a methylation-sensitive complex that bound a 55-bp intronic element corresponding to HS2. Affinity chromatography and mass spectrometry identified a component of this complex to be the GA-binding protein (GABP) ␣. Supershift analysis indicated that the GABP␣ binding partner, GABP␤1, was also present in the complex. The HS2 element conferred a 3-fold enhancement in TMS1 promoter activity, which was dependent on both intact tandem ets binding sites and the presence of GABP␣/␤1 in trans. GABP␣ was selectively enriched at HS2 in human cells, and its occupancy was inversely correlated with CpG island methylation. Down-regulation of GABP␣ led to a concomitant decrease in TMS1 expression. These data indicate that the intronic HS2 element acts in cis to maintain transcriptional competency at the TMS1 locus and that this activity is mediated by the ets transcription factor, GABP␣.Methylation of DNA in the human genome is tightly controlled during development by the action of DNA methyltransferases. DNA methyltransferases catalyze the transfer of a methyl group from S-adenosylmethionine to the carbon-5 position of cytosine in the dinucleotide 5Ј-CpG-3Ј. This epigenetic mark is copied after DNA synthesis, providing a heritable memory of transcriptional status. Overall, CpG dinucleotides are depleted in the human genome except in CpG islands (1). CpG islands are ϳ200 bp to several kb in length and are found mainly in the 5Ј-regions of 70% of human genes (2). Most CpG dinucleotides in the genome are heavily methylated whereas, in contrast, the CpG sites in the CpG islands, especially those associated with gene promoters, are usually unmethylated. Aberrant methylation of such CpG islands is associated with inappropriate gene silencing and has been implicated in the inactivation of tumor suppressor genes in human cancers.At present, the mechanisms underlying cancer-associated CpG island methylation are unknown. There are at least two mechanisms in which aberrant DNA methylation is thought to contribute to stable gene repression. DNA methylation can affect gene expression through its affects on local chromatin structure. Methylated DNA is recognized by methyl-CpGbinding domain proteins, which are components of repressor complexes that contain histone deacetylas...

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