Genetic polymorphism of CYP1A2 in Ethiopians affecting induction and expression: characterization of novel haplotypes with single-nucleotide polymorphisms in intron 1.
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 = 100) or in Sweden (n = 73). We sequenced the CYP1A2 gene using genomic DNA from 12 subjects, which revealed a novel intron 1 single-nucleotide polymorphism (SNP), -730C>T. 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 (-164A), CYP1A2*1J (-740G and -164A), and CYP1A2*1K (-730T, -740G, and -164A), 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 = 136) was 5.9% and 3.6%, and among Spaniards (n = 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 -730C>T SNP alone or in combination with -740T>G (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.
Aklillu
Eleni
E
Division of Molecular Toxicology, Institute of Environmental Medicine, Karolinska Institutet, SE-171 77 Stockholm, Sweden. magnus.ingelman-sundberg@imm.ki.se
Carrillo
Juan Antonio
JA
Makonnen
Eyasu
E
Hellman
Karin
K
Pitarque
Marià
M
Bertilsson
Leif
L
Ingelman-Sundberg
Magnus
M
D000328:Adult; D000704:Analysis of Variance; D001483:Base Sequence; D019388:Cytochrome P-450 CYP1A2 / Q000096:biosynthesis* / Q000235:genetics*; D004790:Enzyme Induction / Q000235:genetics; D005002:Ethiopia; D015971:Gene Expression Regulation, Enzymologic / Q000235:genetics*; D008040:Genetic Linkage / Q000235:genetics; D006239:Haplotypes / Q000235:genetics*; D006801:Humans; D007438:Introns / Q000235:genetics*; D008969:Molecular Sequence Data; D011110:Polymorphism, Genetic / Q000235:genetics; D020641:Polymorphism, Single Nucleotide / Q000235:genetics*; D013548:Sweden; D014407:Tumor Cells, Cultured
D019388:Cytochrome P-450 CYP1A2