SOS-independent mutagenesis in lacZ induced by methylene blue plus visible light.
In vitro photosensitization by visible light in the presence of methylene blue (MB-light) produces lesions in M13mp18 lacZ phage DNA, the lethal and mutagenic potential of which was analyzed after transfection into various bacterial hosts. Mutagenesis was determined with a forward mutation assay using the lacZ gene of M13mp18 as a target. When, MB-light-treated double-stranded (ds) M13mp18 DNA was used to transfect wild-type cells which were not induced for SOS functions, a fivefold increase in mutation frequency was observed at 10% survival compared to that observed with untreated DNA. Mutation frequency obtained with MB-light-treated ds M13mp18 DNA was greater when transfected into the uvr A fpg-1 double mutant than that seen in uvr A, fpg-1, or umuC single mutants or in the wild-type. Sequence analysis shows that in the wild-type strain, MB-light treatment of ds M13mp18 DNA results mostly in single base substitutions. The most frequent base change is the GC-->TA transversion. MB-light treatment of single-stranded (ss) M13mp18 DNA also results in an increased mutation frequency after transfection into the wild-type strain, yielding mostly G-->T transversions. Our results show that MB-light-induced mutagenesis is at least partially independent of the induction of SOS functions in Escherichia coli. The mutation spectra suggest that 8-oxo-7,8-dihydroguanine is the major promutagenic lesion in DNA.
Tudek
B
B
LA147 CNRS, U140 INSERM, Groupe Réparation des lésions radio et chimioinduites, Institut Gustave Roussy, Villejuif, France.
Laval
J
J
Boiteux
S
S
D017104:Bacteriophage M13 / Q000187:drug effects / Q000235:genetics* / Q000528:radiation effects; D001483:Base Sequence; D004249:DNA Damage; D004260:DNA Repair; D004279:DNA, Viral / Q000187:drug effects / Q000235:genetics / Q000528:radiation effects; D004790:Enzyme Induction; D004926:Escherichia coli / Q000187:drug effects / Q000235:genetics* / Q000528:radiation effects; D016368:Frameshift Mutation; D005609:Free Radicals; D005798:Genes, Bacterial / Q000235:genetics; D006147:Guanine / Q000031:analogs & derivatives; D008027:Light; D008751:Methylene Blue / Q000494:pharmacology; D008969:Molecular Sequence Data; D016296:Mutagenesis; D010787:Photosensitivity Disorders; D017354:Point Mutation; D013014:SOS Response, Genetics; D017422:Sequence Analysis, DNA; D014162:Transfection; D001616:beta-Galactosidase / Q000096:biosynthesis
D004279:DNA, Viral; D005609:Free Radicals; C024829:8-hydroxyguanine; D006147:Guanine; D001616:beta-Galactosidase; D008751:Methylene Blue