Mutations induced by 1-nitrosopyrene and related compounds during DNA replication in human cells and induction of homologous recombination by these compounds.
The transformation of normal human cells into cancer cells is a multistep process. Evidence suggests that a minimum of five independent steps (changes) are required for the development of certain kinds of human cancer, as well as for malignant transformation of human cells in culture. Mutations are one of the mechanisms involved in bringing about such changes. A single DNA base substitution mutation can activate an oncogene or inactivate a tumor suppressor gene. Because the action of tumor suppressor genes is to prevent cells from becoming malignant, the activity of both copies of such genes must be eliminated before suppression is lifted. Homologous mitotic recombination between a mutant tumor suppressor gene allele and its non-mutant allele is one mechanism for accomplishing this. The present study was designed to investigate the mechanisms by which certain carcinogenic compounds found in diesel exhaust particles and structurally-related N-substituted aryl carcinogens induce such base substitution mutations and homologous recombination events in mammalian cells in culture, including human cells. The system we employed to determine rapidly the kinds of mutations induced by these compounds, as well as the location of the point mutations in the target gene, involved a circular DNA molecule (plasmid) carrying a small target gene, supF. The target gene was exposed in vitro to radiolabeled compounds and then was allowed to replicate in human cells where the mutations were formed. The sites of mutation induction were compared with the sites of stable binding of the carcinogens to the DNA (adducts). The system used to determine whether these agents could induce homologous recombination consisted of a thymidine kinase-deficient mouse L cell line with a recombination substrate stably integrated into the genome. To determine whether or not excision repair was involved in the mechanism by which carcinogens induced recombination, the recombination substrate was introduced into an excision repair-proficient human cell line and two repair-deficient human cell lines. These cell lines were then compared for the frequency of recombination induced by the agents. All four N-substituted aryl compounds tested in the supF mutagenesis assay produced mainly base substitutions involving guanosine-cytosine (G.C)* base pairs, primarily G.C-->thymidine-adenine (T.A) transversions. However, 1,6-dinitropyrene adducts, formed by exposing the plasmids to 1-nitro-6-nitrosopyrene in the presence of a reducing agent, also induced a significant proportion (17%) of single G.C base-pair deletions.(ABSTRACT TRUNCATED AT 400 WORDS)
Maher
V M
VM
Carcinogenesis Laboratory, Michigan State University, East Lansing 48824.
Bhattacharyya
N P
NP
Mah
M C
MC
Boldt
J
J
Yang
J L
JL
McCormick
J J
JJ
D000818:Animals; D001483:Base Sequence; D001665:Binding Sites; D015139:Blotting, Southern; D002460:Cell Line; D004247:DNA; D004261:DNA Replication / Q000187:drug effects*; D004305:Dose-Response Relationship, Drug; D004353:Drug Evaluation, Preclinical; D006801:Humans; D051379:Mice; D008969:Molecular Sequence Data; D016254:Mutagenesis, Insertional; D009154:Mutation; D011721:Pyrenes / Q000633:toxicity*; D011995:Recombination, Genetic / Q000187:drug effects*; D017384:Sequence Deletion; D014162:Transfection
D011721:Pyrenes; C044707:1-nitrosopyrene; D004247:DNA