Novel single nucleotide polymorphisms in the promoter and intron 1 of human pregnane X receptor/NR1I2 and their association with CYP3A4 expression.
The hypothesis was tested that sequence diversity in pregnane X receptor (PXR) cis-regulatory regions is a significant determinant of variation in inducible and constitutive CYP3A4 expression. A combination of comparative genomics and computational algorithms was used to select regions of the human PXR promoter and intron 1 that were resequenced in the polymorphism discovery resource 24 DNA subset. PXR single nucleotide polymorphisms (SNP) were then genotyped in donor human livers phenotyped for CYP3A4 and multidrug resistance protein 1 mRNA and primary human hepatocytes phenotyped for basal and rifampin-inducible CYP3A4 activity. The human PXR promoter [16.9 kilobase (kb)] was significantly larger than in rodents (2.9 kb). Eighty-nine SNPs were identified in the promoter and intron 1 of PXR. The SNPs most consistently associated with CYP3A4 phenotypic measures were a 44477T>C(-1359) promoter SNP (in linkage disequilibrium with SNP 463970, a 6-base pair deletion in intron 1a, and SNP 46551, a C nucleotide insertion in intron 1b); SNP 63396C>T in intron 1 (in linkage disequilibrium with SNP 63704A>G, a 63813(CAAA)(CA) variable repeat, and SNP 65104T>C); and SNP 56348C>A, SNP 69789A>G, and SNP 66034T>C. Donor livers with the variant PXR alleles had altered hepatic expression of PXR targets compared with livers with PXR wild-type alleles. These results identified PXR promoter and intron 1 SNPs associated with PXR target gene expression (CYP3A4) in donor livers and cultured hepatocytes and that a striking number of the linked intron 1 SNPs will affect putative binding sites for hepatic nuclear factor 3beta (FOXA2), a transcription factor linked with PXR expression.
Lamba
Jatinder
J
Department of Pharmaceutical Sciences, St. Jude Children's Research Hospital, Memphis, Tennessee 38105, USA.
Lamba
Vishal
V
Strom
Stephen
S
Venkataramanan
Raman
R
Schuetz
Erin
E
D051544:Cytochrome P-450 CYP3A; D003577:Cytochrome P-450 Enzyme System / Q000096:biosynthesis* / Q000378:metabolism; D005260:Female; D022781:Hepatocytes / Q000187:drug effects / Q000201:enzymology; D006801:Humans; D007438:Introns / Q000235:genetics*; D008099:Liver / Q000187:drug effects / Q000201:enzymology*; D008297:Male; D020641:Polymorphism, Single Nucleotide; D000077297:Pregnane X Receptor; D011401:Promoter Regions, Genetic; D011987:Receptors, Steroid / Q000235:genetics*; D012293:Rifampin / Q000494:pharmacology; D012727:Sex Characteristics
C000625852:NR1I2 protein, human; D000077297:Pregnane X Receptor; D011987:Receptors, Steroid; D003577:Cytochrome P-450 Enzyme System; D051544:Cytochrome P-450 CYP3A; C510163:CYP3A4 protein, human; D012293:Rifampin