Regulation of the Human Intrauterine Renin Angiotensin System by DNA Methylation (#361)
Several RAS components have CpG islands near their transcription start sites. During pregnancy the RAS has potential roles in decidualisation and placentation. Therefore, DNA methylation of key RAS genes may participate in these processes.
To explore this, we measured the methylation status and mRNA expression of 3 RAS genes with CpG islands near their promoter (angiotensin converting enzyme; ACE, angiotensin II type 1 receptor AGTR1, prorenin receptor; ATP6AP2) and 2 proteases that activate prorenin (cathepsin D; CTSD and kallikrein 1; KLK1) in term amnion, decidua and placenta using the Methyl-Profiler assay and qPCR, respectively.
ACE, AGTR1 and CTSD promoters were predominantly unmethylated (>90%) and the methylation status did not differ between tissues. ATP6AP2 methylation was also low overall, but was significantly higher in decidua compared to placenta (P<0.01). KLK1 hypermethylation was less in placenta (3.3%) than in amnion and decidua (33%; P=0.001). When examining the corresponding changes in mRNA abundance, ACE expression was found to be significantly higher in decidua compared to amnion (P=0.015), whilst CTSD expression was significantly higher in placenta compared to amnion (P=0.049). Placental expression of AGTR1 was significantly higher than either amnion (P=0.01) or decidua (P=0.004).
The methylation status of ACE, AGTR1 and CTSD was unchanged and yet there were significant differences in their mRNA expression. By contrast, changes in the methylation status of KLK1 and ATP6AP2 did not result in corresponding changes in their gene expression. Thus it is unlikely that methylation of CpG islands near the promoter regions of these genes controls gene expression. This finding is reinforced by studies in which we showed that treatment of a trophoblast cell line (HTR-8/SVneo) with the demethylating agent, 5’aza-2’-deoxycytidine had no effect on the expression of ACE, ATP6AP2 and AGTR1. Thus other factors may be responsible for regulating their expression in intrauterine tissues.