title: Spontaneous and ethylnitrosourea-induced mutation fixation and molecular spectra at the lacI transgene in the Big Blue rat-2 embryo cell line.
abstract: Big Blue Rat-2 cells were evaluated for mutagenesis and mutational spectra (spontaneous and ethylnitrosourea [ENU]-induced). Survival, mutant frequency, population doubling time, and kinetics of mutant increase (to 120 hr) were determined. Exposures were 100, 200, 400, 600, and 1,000 micrograms ENU/ml. The spontaneous mutant frequency was similar to that previously reported in vivo, i.e., 5 X 10(5). Dose-related increases in mutant frequency were observed following ENU treatment. Kinetics (time course) of mutant frequency increase, population doubling, and mutational spectra were investigated following treatment at 1,000 micrograms ENU/ml. Among 39 spontaneous mutants, 26 independent mutations were found as follows: nine (34.6%) G:C-->A:T transitions (five at CpG sites), six (23%) G:C-->T:A transversions, three (11.5%) G:C-->C:G transversions (two at CpG sites), two (7.7%) frameshifts, five (19%) deletions or insertions, and one (3.8%) complex (deletion+insertion) mutation. Among 46 ENU-induced mutants, 37 independent mutations (all base substitutions) were found as follows: 15 (40.5%) G:C-->A:T transitions (four at CpG sites), five (13.5%) A:T-->G:C transitions, four (10.8%) G:C-->T:A transversions, 11 (30%) A:T-->T:A transversions, and two (5.4%) A:T-->C:G transversions. Nearly 50% of the base substitutions in the ENU-treated cells were at A:T base pairs, in contrast to the spontaneous mutants where none was found. Both the spontaneous and the ENU-induced mutational spectra were similar to that reported in vivo and for other cells. An important aspect of the experiment is that all mutations sequenced following ENU treatment (1,000 micrograms/ml) occurred under conditions which our experiments show corresponded to very little mitotic activity.
authors.affiliation: Pharmacia & Upjohn, Inc., Kalamazoo, Michigan 49007, USA.
authors.forename: D M
authors.initials: DM
authors.lastname: Zimmer
authors.forename: X B
authors.initials: XB
authors.lastname: Zhang
authors.forename: P R
authors.initials: PR
authors.lastname: Harbach
authors.forename: J K
authors.initials: JK
authors.lastname: Mayo
authors.forename: C S
authors.initials: CS
authors.lastname: Aaron
mesh_terms: D000818:Animals; D030801:Animals, Genetically Modified / Q000235:genetics; D001426:Bacterial Proteins / Q000187:drug effects / Q000235:genetics*; D016923:Cell Death / Q000187:drug effects / Q000235:genetics; D002455:Cell Division / Q000187:drug effects / Q000235:genetics; D002460:Cell Line; D004305:Dose-Response Relationship, Drug; D004622:Embryo, Mammalian / Q000166:cytology* / Q000187:drug effects / Q000502:physiology; D029968:Escherichia coli Proteins; D005038:Ethylnitrosourea / Q000633:toxicity*; D056968:Lac Repressors; D009152:Mutagenicity Tests / Q000379:methods; D009153:Mutagens / Q000633:toxicity; D009154:Mutation; D051381:Rats; D012097:Repressor Proteins / Q000187:drug effects / Q000235:genetics*; D017422:Sequence Analysis, DNA; D019076:Transgenes
chemical_list: D001426:Bacterial Proteins; D029968:Escherichia coli Proteins; D056968:Lac Repressors; D009153:Mutagens; D012097:Repressor Proteins; D005038:Ethylnitrosourea