Effect of Dietary Salt excess on DNA Methylation and Transcriptional Regulation of human Angiotensinogen (hAGT) Gene Expression.

Background: Hypertension is caused by a combination of genetic and environmental factors. Angiotensinogen (AGT) is a component of renin-angiotensin-aldosterone system (RAAS), that regulates blood pressure. Genome-wide association studies (GWAS) have shown that two A/G polymorphisms (rs2493134 and rs2004776), located at +507 and +1164 in intron I of the human AGT (hAGT) gene, are linked to hypertension. AGT polymorphisms result in two haplotypes, Hap-I a pro-hypertensive, whereas Hap-II is normotensive. Previous studies support the role of epigenetics in blood pressure regulation. In this study we generated transgenic mice (TG) with hAGT containing Hap-I and Hap-II variants to investigate the effect of high salt diet (HSD) on epigenetics and transcriptional regulation.

Methods: We treated Hap-I and Hap-II TG mice with 4% HSD and identified DNA methylation patterns. We measured hAGT mRNA and protein by qPCR and immunoblot respectively. ChIP assay and RNA sequencing were performed.

Results: hAGT gene expression is increased by HSD in both Hap-I and Hap-II TG mice. In the liver and kidney, we observed significantly higher DNA demethylation (less CpG's) and stronger binding of transcription factors in the promoter of Hap-I TG mice as compared to Hap-II post HSD. RNA-Seq identified differentially expressed genes, novel target genes, canonical pathways, and upstream regulators associated with hypertension.

Conclusions: Our findings identified a novel high salt-sensitive risk haplotype, novel CpG sites and DNA methylation patterns, potential gene targets, and pathways implicated in hypertension. Combining epigenetic and transcriptional analysis allows for a more holistic understanding of the regulatory mechanisms that govern the hAGT gene.