title: Genetic analysis of the anti-mutagenic effect of genistein in Escherichia coli.
abstract: Genistein, the main isoflavone in soy, has received considerable attention for its potential anti-carcinogenic properties. In a previous report, we investigated the possible role of genistein in anti-mutagenesis, using an Escherichia coli reversion assay system. Genistein reduced ENU-induced mutagenesis in a dose-dependent manner and the reduction of mutation frequency was differential among several categories of mutation. Most notable was a loss of transversion mutations, which require SOS functions. In this report, we further investigated the anti-mutagenic effect of genistein using a genetic approach. E. coli strains having alterations in genes involved in SOS-mutagenesis were examined, as were strains having defects in proteins that might serve as potential targets for genistein. The results showed that ENU-induced mutations produced in recA730 and lexA(Def) strains, both expressing a constitutive SOS response, were reduced by genistein to a lesser extent than in the wild-type strain. The effect of genistein was not entirely abolished, however. ENU mutagenesis in a umuC derivative, which reflects predominantly transition mutations, was unaffected by genistein. ENU-induced mutations in strains having defects in topA, gyrA, typA or uspA were not different than the wild-type, suggesting that these gene products were not involved in genistein's anti-mutagenic effect. In addition, we determined the distribution of genistein in various cellular fractions using HPLC. These studies revealed that genistein could be recovered from E. coli cells grown on agar media containing genistein; the intracellular concentration was similar to that in the agar plates. Further, most of the genistein recovered was associated with proteins in the cytosolic fraction and little partitioned in the membrane fraction. In vitro studies showed that genistein could be precipitated from a protein (BSA) containing solution. Finally, we examined the effect of genistein on formation of the RecA filament on ssDNA in vitro and observed an inhibition at high concentrations of genistein. In total, these results suggested that genistein may reduce SOS-dependent mutagenesis by reducing the interaction of RecA protein with ssDNA. As a consequence, genistein could cause a reduction in (1) the overall SOS response (confirmed using beta-galactosidase assays) and (2) trans-lesion DNA synthesis by DNA polymerase V.
authors.affiliation: Department of Microbiology, Southern Illinois University, Carbondale, IL 62901, United States.
authors.forename: Yixin
authors.initials: Y
authors.lastname: Yang
authors.forename: Douglas
authors.initials: D
authors.lastname: Fix
mesh_terms: D016588:Anticarcinogenic Agents / Q000494:pharmacology*; D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism; D002851:Chromatography, High Pressure Liquid; D004277:DNA, Single-Stranded / Q000378:metabolism; D004259:DNA-Directed DNA Polymerase; D024202:Electrophoretic Mobility Shift Assay; D004926:Escherichia coli / Q000235:genetics*; D029968:Escherichia coli Proteins / Q000235:genetics / Q000378:metabolism; D005038:Ethylnitrosourea / Q000378:metabolism / Q000494:pharmacology; D005798:Genes, Bacterial / Q000187:drug effects; D019833:Genistein / Q000494:pharmacology*; D016296:Mutagenesis / Q000187:drug effects; D009154:Mutation / Q000187:drug effects; D011938:Rec A Recombinases / Q000235:genetics / Q000378:metabolism; D013014:SOS Response, Genetics; D012697:Serine Endopeptidases / Q000235:genetics / Q000378:metabolism; D013997:Time Factors; D000099160:LexA Repressor Protein
chemical_list: D016588:Anticarcinogenic Agents; D001426:Bacterial Proteins; D004277:DNA, Single-Stranded; D004259:DNA-Directed DNA Polymerase; D029968:Escherichia coli Proteins; D005038:Ethylnitrosourea; D019833:Genistein; D011938:Rec A Recombinases; D012697:Serine Endopeptidases; D000099160:LexA Repressor Protein; C078202:UmuC protein, E coli