Use of shuttle vectors to study the molecular processing of defined carcinogen-induced DNA damage: mutagenicity of single O4-ethylthymine adducts in HeLa cells.
We developed a simian virus 40 based shuttle vector system to study the molecular consequences of distinct carcinogen-induced DNA lesions in human cells. To establish the mutagenicity of O4-ethylthymine adducts, oligonucleotides carrying a single O4-ethylthymine adduct at a unique position were ligated into the vector molecules. Following replication in HeLa cells on average 23% of the progeny molecules carried a mutation in the region of modification. The vast majority of these mutations represented single T----C transitions at the position of the modified base, most probably as a consequence of mispairing of the O4-ethylthymine residues during replication. To a minor extent the O4-ethylthymine adduct may also induce T----A transversions or double point mutations. The in vivo mutation frequency of the adduct was found to be comparable to that of a C-A mismatch at the same position, but was lower than that expected from in vitro experiments with adducted DNA templates and purified DNA polymerases.
Klein
J C
JC
Division of Chemical Carcinogenesis, The Netherlands Cancer Institute, Amsterdam.
Bleeker
M J
MJ
Lutgerink
J T
JT
van Dijk
W J
WJ
Brugghe
H F
HF
van den Elst
H
H
van der Marel
G A
GA
van Boom
J H
JH
Westra
J G
JG
Berns
A J
AJ
D001483:Base Sequence; D002273:Carcinogens / Q000633:toxicity*; D002460:Cell Line; D004249:DNA Damage; D004261:DNA Replication; D005822:Genetic Vectors; D006367:HeLa Cells; D006801:Humans; D007700:Kinetics; D008969:Molecular Sequence Data; D009152:Mutagenicity Tests; D009154:Mutation; D009838:Oligodeoxyribonucleotides / Q000302:isolation & purification; D013941:Thymine / Q000031:analogs & derivatives* / Q000633:toxicity; D014162:Transfection
D002273:Carcinogens; D009838:Oligodeoxyribonucleotides; C040384:4-ethylthymine; D013941:Thymine