Upstream stimulatory factor is required for human angiotensinogen expression and differential regulation by the A-20C polymorphism. Circ Res 2008 Oct 24;103(9):940-7
Date
09/20/2008Pubmed ID
18802024Pubmed Central ID
PMC2678906DOI
10.1161/CIRCRESAHA.108.180653Scopus ID
2-s2.0-55449090345 (requires institutional sign-in at Scopus site) 17 CitationsAbstract
Among naturally occurring polymorphisms in the 5' flanking region of the human angiotensinogen (AGT) gene, the -20 and -217 polymorphisms have the strongest effects on AGT regulation in AGT-expressing cells derived from liver, kidney, brain, and fat. These polymorphisms may affect allele-specific transcription factor binding, and the high-expressing alleles are both relatively common. We show herein that the -20C allele has higher transcriptional activity than -20A, and the -20A allele confers no additional transactivation potential beyond that of a mutated vector. Gel-shift assays show that upstream stimulatory factor (USF)1 and USF2 preferentially bind the -20C allele, whereas the -20A allele retains a low affinity USF binding site. Plasmid immunoprecipitation assays confirmed preferential association of USF1 with the -20C allele in transfected HepG2 cells. Chromatin immunoprecipitation confirmed that USF1 binds to the endogenous AGT -20C allele in CCF cells, the only cell line tested that carries the -20C allele, and to the human AGT promoter in liver and adipose tissue from transgenic mice. Transduction of AGT-expressing cells with short hairpin RNAs specifically targeting USF1 or USF2, resulted in cell- and allele-specific attenuation of AGT promoter activity. In vivo, knockdown of USF expression in the liver of transgenic mice expressing the -20C allele of AGT resulted in lower AGT expression, a decrease in circulating human AGT protein but no change in expression of GAPDH or hepatocyte nuclear factor-4alpha. We conclude that USF1 functionally and differentially regulates AGT expression via the -20 polymorphism and that the differential expression exhibited by -20 can be accounted for by differential association with USF1.
Author List
Dickson ME, Tian X, Liu X, Davis DR, Sigmund CDAuthor
Curt Sigmund PhD Chair, Professor in the Physiology department at Medical College of WisconsinMESH terms used to index this publication - Major topics in bold
3T3-L1 Cells5' Flanking Region
Adipose Tissue
Angiotensinogen
Animals
Cell Line, Tumor
Chromatin Immunoprecipitation
Electrophoretic Mobility Shift Assay
Humans
Liver
Mice
Mice, Transgenic
Polymorphism, Genetic
Promoter Regions, Genetic
RNA Interference
RNA, Small Interfering
Transcription, Genetic
Transcriptional Activation
Transfection
Upstream Stimulatory Factors
