Abstract
Angiotensinogen (AGT) is the precursor of one of the most potent vasoconstrictors, peptide angiotensin II. Genome-wide association studies 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 associated with hypertension. Polymorphisms of the AGT gene result in two main haplotypes. Hap-I contains the variants -217A, -6A, +507G, and +1164A and is pro-hypertensive, whereas Hap-II contains the variants -217G, -6G, +507A, and +1164G and does not affect blood pressure. The nucleotide sequence of intron I of the hAGT gene containing the +1164A variant has a stronger homology with the hepatocyte nuclear factor 3 (HNF3)-binding site than +1164G. Here we found that an oligonucleotide containing +1164A binds HNF3β more strongly than +1164G and that Hap-I-containing reporter gene constructs have increased basal and HNF3- and glucocorticoid-induced promoter activity in transiently transfected liver and kidney cells. Using a knock-in approach at the hypoxanthine-guanine phosphoribosyltransferase locus, we generated a transgenic mouse model containing the human renin (hREN) gene and either Hap-I or Hap-II. We show that transgenic animals containing Hap-I have increased blood pressure compared with those containing Hap-II. Moreover, the transcription factors glucocorticoid receptor, CCAAT enhancer-binding protein β, and HNF3β bound more strongly to chromatin obtained from the liver of transgenic animals containing Hap-I than to liver chromatin from Hap-II-containing animals. These findings suggest that, unlike Hap-II variants, Hap-I variants of the hAGT gene have increased transcription rates, resulting in elevated blood pressure.
Highlights
Angiotensinogen (AGT) is the precursor of one of the most potent vasoconstrictors, peptide angiotensin II
It was shown that polymorphism M235T of the human AGT (hAGT) gene is associated with essential hypertension, and homozygotes with variant T235 contain 15%– 40% higher plasma AGT levels [3]
We show the following: reporter constructs containing ϩ1164A have increased basal and glucocorticoid-induced promoter activity on transient transfection in liver and kidney cells compared with ϩ1164G; an oligonucleotide containing ϩ1164A binds more strongly to hepatocyte nuclear factor 3 (HNF3) compared with the same oligonucleotide containing ϩ1164G; double TG animals containing human renin (hREN) and Hap-I of the hAGT gene have increased blood pressure compared with double TG animals containing hREN and Hap-II of the hAGT gene; and the transcription factors GR, C/EBP, and HNF3 bind more strongly to chromatin obtained from livers of TG animals containing Hap-I compared with Hap-II
Summary
Proteolytic cleavage of a larger precursor molecule, angiotensinogen (AGT), which is primarily synthesized in the liver and, to a lesser extent, in fat, kidney, brain, and heart and adrenal and vascular walls [2]. Two other GWAS have shown that an A/G SNP (rs2004776) located at the ϩ1164 position in intron I in the hAGT gene is associated with blood pressure [12, 13]. Ated haplotypes across the AGT promoter in 256 cases and 126 controls from the PATH-SCOR study group They concluded that the hAGT haplotype that contains variants Ϫ217A, Ϫ6A, and ϩ1164A of intron I (Hap-I) predisposes to hypertension and that the haplotype containing Ϫ217G, Ϫ6G, and ϩ1164G of intron I (Hap-II) does not have an appreciable effect on blood pressure (Fig. 2A). We have shown previously that the nucleotide sequence of the hAGT gene containing variant Ϫ217A has stronger homology with C/EBP and GR binding sites compared with Ϫ217G and that reporter constructs containing Ϫ217A have increased promoter activity on transient transfection in human liver and kidney cells compared with Ϫ217G. We show the following: reporter constructs containing ϩ1164A have increased basal and glucocorticoid-induced promoter activity on transient transfection in liver and kidney cells compared with ϩ1164G; an oligonucleotide containing ϩ1164A binds more strongly to HNF3 compared with the same oligonucleotide containing ϩ1164G; double TG animals containing hREN and Hap-I (containing Ϫ6A, Ϫ217A, ϩ507G, and ϩ 1164A) of the hAGT gene have increased blood pressure compared with double TG animals containing hREN and Hap-II (containing Ϫ6G, Ϫ217G, ϩ507A, and ϩ1164G) of the hAGT gene; and the transcription factors GR, C/EBP, and HNF3 bind more strongly to chromatin obtained from livers of TG animals containing Hap-I compared with Hap-II
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