title: Role and importance of polymorphisms with respect to DNA methylation for the expression of CYP2E1 enzyme.
abstract: Different individuals possess slightly different genetic information and show genetically-determined differences in several enzyme activities due to genetic variability. Following an integrated approach, we studied the polymorphisms and methylation of sites contained in the 5' flanking region of the metabolizing enzyme CYP2E1 in correlation to its expression in both tumor and non-neoplastic liver cell lines, since to date little is known about the influence of these (epi)genetic elements in basal conditions and under induction by the specific inductor and a demethylating agent. In treated cells, reduced DNA methylation, assessed both at genomic and gene level, was not consistently associated with the increase of enzyme expression. Interestingly, the Rsa/Pst haplotype differentially influenced CYP2E1 enzyme expression. In addition, regarding the Variable Number of Tandem Repeats polymorphism, cells with A4/A4 genotype showed a greater expression inhibition (ranging from 20% to 30%) compared with others carrying the A2/A2 one, while those cells bringing A2/A3 genotype showed an increase of expression (of 25%, about). Finally, we demonstrated for the first time that the A2 and A3 CYP2E1 alleles play a more important role in the expression of the enzyme, compared with other (epi)genetic factors, since they are binding sites for trans-acting proteins.
authors: Dipartimento di Scienze e Tecnologie Biologiche Chimiche e Farmaceutiche (STEBICEF), Sezione di Biologia Cellulare, Edificio 16, Università di Palermo, V.le delle Scienze, 90128 Palermo, Italy.
authors: Flores
authors: F
authors: Naselli
authors: Dipartimento di Scienze e Tecnologie Biologiche Chimiche e Farmaceutiche (STEBICEF), Sezione di Biologia Cellulare, Edificio 16, Università di Palermo, V.le delle Scienze, 90128 Palermo, Italy.
authors: Irene
authors: I
authors: Catanzaro
authors: Dipartimento di Scienze e Tecnologie Biologiche Chimiche e Farmaceutiche (STEBICEF), Sezione di Biologia Cellulare, Edificio 16, Università di Palermo, V.le delle Scienze, 90128 Palermo, Italy; Istituto Ortopedico Rizzoli, c/o Dipartimento di Biopatologia e Biotecnologie Mediche e Forensi (DIBIMEF), Università di Palermo, Via Divisi, 81, 90133 Palermo, Italy.
authors: Daniele
authors: D
authors: Bellavia
authors: Dipartimento di Scienze e Tecnologie Biologiche Chimiche e Farmaceutiche (STEBICEF), Sezione di Biologia Cellulare, Edificio 16, Università di Palermo, V.le delle Scienze, 90128 Palermo, Italy.
authors: Alessandro
authors: A
authors: Perez
authors: Dipartimento di Scienze e Tecnologie Biologiche Chimiche e Farmaceutiche (STEBICEF), Sezione di Biologia Cellulare, Edificio 16, Università di Palermo, V.le delle Scienze, 90128 Palermo, Italy.
authors: Laura
authors: L
authors: Sposito
authors: Dipartimento di Scienze e Tecnologie Biologiche Chimiche e Farmaceutiche (STEBICEF), Sezione di Biologia Cellulare, Edificio 16, Università di Palermo, V.le delle Scienze, 90128 Palermo, Italy. Electronic address: fabio.caradonna@unipa.it.
authors: Fabio
authors: F
authors: Caradonna
mesh_terms: D024506:5' Flanking Region; D001374:Azacitidine / Q000494:pharmacology; D002470:Cell Survival / Q000187:drug effects / Q000235:genetics; D019392:Cytochrome P-450 CYP2E1 / Q000235:genetics*; D019175:DNA Methylation; D015971:Gene Expression Regulation, Enzymologic / Q000187:drug effects; D005838:Genotype; D056945:Hep G2 Cells; D006801:Humans; D018598:Minisatellite Repeats / Q000235:genetics; D011110:Polymorphism, Genetic; D012150:Polymorphism, Restriction Fragment Length; D014407:Tumor Cells, Cultured
chemical_list: D019392:Cytochrome P-450 CYP2E1; D001374:Azacitidine
keywords: 5-Azacytidine; 5-azacytidine; BSA; CYP; CYP2E1; CYP2E1 gene polymorphisms; DMSO; DNA; DNA methylation; DTT; EDTA; EMSA; Electrophoretic Mobility Shift Assay; Ethanol; HCC; HRP; Hepatocellular carcinoma cell lines; MSRE-PCR; MTT; MeSAP-PCR; Methylation Sensitive Restriction Endonuclease-Polymerase Chain Reaction; PBS; PCR; Polymerase Chain Reaction; RFLP; ROS; Reactive Oxygen Species; Restriction Fragment Length Polymorphism; VNTR; Variable Number of Tandem Repeats; ZCyd; [3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide]; bovine serum albumin; cytochrome p450; cytochrome p450 2E1; deoxyribonucleic acid; dimethyl sulfoxyde; dithiothreitol; ethylene diamine tetraacetic acid; hepato-cellular carcinoma; horse radish peroxidase; methylation sensitive arbitrarily primed-polymerase chain reaction; phosphate buffered saline