Identification of a Single Nucleotide Polymorphism in the TATA Box of the CYP2A6 Gene: Impairment of Its Promoter Activity

• Published in: Biochemical and biophysical research communications Vol. 284; no. 2; pp. 455 - 460
• Main Authors: Pitarque, Marià, von Richter, Oliver, Oke, Basbug, Berkkan, Hakan, Oscarson, Mikael, Ingelman-Sundberg, Magnus
• Format: Journal Article
• Language: English
• Published: United States Elsevier Inc 08.06.2001
• Subjects: Alleles; Aryl Hydrocarbon Hydroxylases; Asian Continental Ancestry Group - genetics; Cell Line; China - ethnology; Chromatography, High Pressure Liquid; coumarin; CYP2A6 gene; Cytochrome P-450 CYP2A6; Cytochrome P-450 Enzyme System - genetics; DASH; DNA Mutational Analysis; European Continental Ancestry Group - genetics; Gene Expression - physiology; gene transcription; Genes, Reporter; genetic polymorphism; Genetic Testing; Genetic Variation; Genotype; Humans; lung cancer; Mixed Function Oxygenases - genetics; nicotine; Nucleic Acid Denaturation; Nucleic Acid Heteroduplexes - analysis; Nucleic Acid Heteroduplexes - genetics; Nucleic Acid Hybridization - methods; Oxidoreductases - genetics; Polymorphism, Single Nucleotide - genetics; Promoter Regions, Genetic - physiology; Sweden - ethnology; TATA Box - genetics; Transfection; Turkey - ethnology; lung cancer; nicotine; gene transcription; genetic polymorphism; DASH; coumarin
• ISSN: 0006-291X; 1090-2104
• DOI: 10.1006/bbrc.2001.4990

Human cytochrome P450 2A6 (CYP2A6) constitutes the major nicotine oxidase, and large interindividual differences are seen in the levels of this enzyme, to a great extent caused by the distribution of several different polymorphic gene variants mainly located in the open reading frame (ORF). In the present study, we report a common polymorphism located in the 5′ flanking region of CYP2A6 affecting its expression. DHPLC analysis and complete sequence of the open reading frame of the gene from a Turkish individual revealed a −48T > G substitution disrupting the TATA box. Using dynamic allele-specific hybridization (DASH), genotyping of this novel variant (named CYP2A6*9) was carried out in 116 Swedish, 132 Turkish, and 102 Chinese subjects, and the allele frequencies were found to be 5.2, 7.2, and 15.7%, respectively. The significance of the polymorphism was investigated by the construction of luciferase reporter plasmids containing 135 or 500 bp of the 5′-upstream region of the gene transfected into human hepatoma B16A2 cells. The constructs carrying the −48T > G mutation were only expressed at about 50% of the wild-type alleles. It is concluded that the CYP2A6*9 allele might be one of the most common CYP2A6 variants in Caucasians that alters the levels of enzyme expression.