CpG methylation at the USF-binding site mediates cell-specific transcription of human ascorbate transporter SVCT2 exon 1a

SVCT2 (sodium vitamin C co-transporter 2) is the major transporter mediating vitamin C uptake in most organs. Its expression is driven by two promoters (CpG-poor exon 1a promoter and CpG-rich exon 1b promoter). In the present study, we mapped discrete elements within the proximal CpG-poor promoter responsible for exon 1a transcription. We identified two E boxes for USF (upstream stimulating factor) binding and one Y box for NF-Y (nuclear factor Y) binding. We show further that NF-Y and USF bind to the exon la promoter in a co-operative manner, amplifying the bindimg of each to the promoter, and is absolutely required for the full activity of the exon la promoter. The analysis of the CpG site located at the upstream USF-binding site in the promoter showed a strong correlation between expression and demethylation. It was also shown that exon I a transcription was induced in cell culture treated with the demethylating agent decitabine. The specific methylation of this CpG site impaired both the binding of USE and the formation of the functional NF-Y USF complex as well as promoter activity, suggesting its importance for cell-specific transcription. Thus CpG methylation at the upstream USF-binding site functions in establishing and maintaining cell-specific transcription from the CpG-poor SVCT2 ex on 1a promoter.