Demethylation of a specific hypersensitive site in the Th2 locus control region

A growing body of literature has examined and implicated DNA methylation as a critical epigenetic modification in T helper (Th) cell differentiation. The absence of DNA methyltransferases or methyl-binding proteins derepresses many cytokine loci, allowing their ectopic expression, while methylation of specific CpG residues is sufficient to prevent expression. Here, we characterize demethylation events of the Th2 cytokine locus control region (LCR). rad50 hypersensitive site 7 (RHS7), a hypersensitive site within this LCR, becomes demethylated in a STAT6-dependent manner and only in cells stimulated under type 2 conditions. Robust demethylation appears to require signaling contributions from both IL-4 receptor, via STAT6, and CD28, but it cannot be effected by GATA3. Finally, RHS7 is demethylated independently of cell division, consistent with an "active," rather than passive, mechanism. Taken together, these findings firmly connect RHS7 demethylation and Th2 LCR activation in the type 2 differentiation program.