Role of DNA methylation in expression control of the IKZF3-GSDMA region in human epithelial cells

Chromosomal region 17q12-q21 is associated with asthma and harbors regulatory polymorphisms that influence expression levels of all five protein-coding genes in the region: IKAROS family zinc finger 3 (Aiolos) (IKZF3), zona pellucida binding protein 2 (ZPBP2), ORMDL sphingolipid biosynthesis regulator 3 (ORMDL3), and gasdermins A and B (GSDMA, GSDMB). Furthermore, DNA methylation in this region has been implicated as a potential modifier of the genetic risk of asthma development. To further characterize the effect of DNA methylation, we examined the impact of treatment with DNA methyltransferase inhibitor 5-aza-2'-deoxycytidine (5-aza-dC) that causes DNA demethylation, on expression and promoter methylation of the five 17q12-q21 genes in the human airway epithelium cell line NuLi-1, embryonic kidney epithelium cell line 293T and human adenocarcinoma cell line MCF-7. 5-aza-dC treatment led to upregulation of expression of GSDMA in all three cell lines. ZPBP2 was upregulated in NuLi-1, but remained repressed in 293T and MCF-7 cells, whereas ORMDL3 was upregulated in 293T and MCF-7 cells, but not NuLi-1. Upregulation of ZPBP2 and GSDMA was accompanied by a decrease in promoter methylation. Moreover, 5-aza-dC treatment modified allelic expression of ZPBP2 and ORMDL3 suggesting that different alleles may respond differently to treatment. We also identified a polymorphic CTCF-binding site in intron 1 of ORMDL3 carrying a CG SNP rs4065275 and determined its methylation level. The site's methylation was unaffected by 5-aza-dC treatment in NuLi-1 cells. We conclude that modest changes (8-13%) in promoter methylation levels of ZPBP2 and GSDMA may cause substantial changes in RNA levels and that allelic expression of ZPBP2 and ORMDL3 is mediated by DNA methylation.