Genetic Polymorphism of CYP1A2 in Ethiopians Affecting Induction and Expression: Characterization of Novel Haplotypes with Single-Nucleotide Polymorphisms in Intron 1

Abstract

CYP1A2 polymorphism has been well studied in white persons and Asians but not in Africans. We performed CYP1A2 genotype and phenotype analysis using caffeine in Ethiopians living in Ethiopia (n ϭ 100) or in Sweden (n ϭ 73). We sequenced the CYP1A2 gene using genomic DNA from 12 subjects, which revealed a novel intron 1 single-nucleotide polymorphism (SNP), Ϫ730CϾT. We developed SNP-specific polymerase chain reaction-restriction fragment length polymorphism genotyping and molecular haplotyping methods for the intron 1 SNPs, and four different haplotypes were identified: CYP1A2*1A (wild-type for all SNPs), CYP1A2*1F (Ϫ164A), CYP1A2*1J (Ϫ740G and Ϫ164A), and CYP1A2*1K (Ϫ730T, Ϫ740G, and Ϫ164A), having frequencies of 39.9, 49.6, 7.5, and 3.0%, respectively. The frequency of CYP1A2*1J and CYP1A2*1K among Saudi Arabians (n ϭ 136) was 5.9% and 3.6%, and among Spaniards (n ϭ 117) 1.3% and 0.5%, respectively. Functional significance of the different intron 1 haplotypes was analyzed. Subjects with CYP1A2*1K had significantly decreased CYP1A2 activity in vivo, and reporter constructs with this haplotype had significantly less inducibility with 2,3,7,8-tetrachlorodibenzo-p-dioxin in human B16A2 hepatoma cells. Electrophoretic mobility shift assay using nuclear extracts from B16A2 cells revealed a specific DNA binding protein complex to an Ets element. Efficient competition was obtained using oligonucleotide probes carrying the wt sequence and Ets consensus probe, whereas competition was abolished using probes with the Ϫ730CϾT SNP alone or in combination with Ϫ740TϾG (CYP1A2*1K). The results indicate a novel polymorphism in intron 1 of importance for Ets-dependent CYP1A2 expression in vivo and inducibility of the enzyme, which might be of critical importance for determination of interindividual differences in drug metabolism and sensitivity to carcinogens activated by CYP1A2.

Allele-specific expression and gene methylation in the control of CYP1A2 mRNA level in human livers

“…However, the low frequency of the functionally different variant alleles in the populations and the results of several genotype-phenotype association studies indicate a limited function of CYP1A2 genetic polymorphism as a cause of interindividual variation in CYP1A2 gene expression or activity. 11,12 In line of this research, we have found one allele CYP1A2*1K causing lower expression of the gene but it is present in Ethiopians 13 and almost absent in whites and Asians. 11,12 The interethnic differences in CYP1A2 activity is important and by controlling for the effects of smoking, oral contraceptive use and genotype, we recently reported the presence of significant differences in enzyme activity between Swedish, Korean and Serbian populations, differences not explained by the known CYP1A2 single nucleotide polymorphisms (SNPs)/ haplotypes.…”

“…Previously we identified this SNP (rs2470890) in Ethiopians and reported lack of influence on CYP1A2 enzyme activity using caffeine as a probe. 13 To examine linkage of the marker SNP with other CYP1A2 SNPs, we genotyped livers for known CYP1A2 variant SNPs (À3858G4A, À2467delT, À739T4G, À729C4T, À163C4A) as described previously. 12 PCR primers and minisequencing primers with 5 0 tag sequences covering the marker SNP site were designed using the Autoprimer (http://www.autoprimer.com; Beckman Coulter) software.…”

The cytochrome P450 CYP1A2 genetic polymorphisms *1F and *1D do not affect clozapine clearance in a group of schizophrenic patients

“…À163C4A, used here for Ã1F genotyping, is also part of the Ã1K allele, which has been associated with decreased activity. 6 Some of the subjects designated as Ã1F in this study may thus have the Ã1K allele.…”

An SduI polymorphism at intron 20 of the Chinese Holstein cow STAT4 gene and its effect on milk performance traits

“…Many SNPs in introns are thought to be non-functional (Crawford et al, 2005;Buchanan et al, 2012). Meanwhile, increasing research is revealing that polymorphisms located in introns could influence gene expression by altering splice sites, disrupting a transcription factor-binding site, or a binding site for modified histone proteins (Aklillu et al, 2003;von Ahsen and Oellerich, 2004;Skarratt et al, 2005;Warnecke et al, 2005;Bu et al, 2006;Szafranski et al, 2007;Wang et al, 2011). It is unclear how intronic SNPs in STAT4 cause differences in milk performance traits in Chinese Holstein cows; however, it is possible that they have the potential to influence STAT4 expression or protein activity, and further studies are needed for this mechanism to be uncovered and well understood.…”