Activating mutations in human c-ha-ras-1 gene induced by reactive derivatives of safrole and the glutamic pyrolysis product, Glu-P-3

Ireland, Christine M., Cooper, Colin S. ORCID:, Marshall, Christopher J., Hebert, Eric and Phillips, David H. (1988) Activating mutations in human c-ha-ras-1 gene induced by reactive derivatives of safrole and the glutamic pyrolysis product, Glu-P-3. Mutagenesis, 3 (5). pp. 429-435. ISSN 0267-8357

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Foci of transformed NIH3T3 cells were observed after transfection of plasmids containing the c-Ha-ras-1 protooncogene modified in vitro either with the 3-N,N-acetoxyacetyl derivative (N-AcO-AGlu-P-3) of the mutagenic L-glutamic acid pyrolysis product 3-amino-4, 6-dimethyldipyrido-[1,2-a:3′,2′-d]imidazole (Glu-P-3) or with 1′-acetoxysafrole (AcO-S), a reactive derivative of the carcinogen safrole. DNA isolated from these foci were used in a second round of transfection, and the DNA obtained from the secondary transformants was analysed to determine the nature of mutations responsible for activating the protooncogene. The polymerase chain reaction method was used to amplify sequences of the gene likely to contain activating mutations, and these regions were then subjected to selective hybridization with specific oligonucleotides to locate and identify the point mutations. Five out of six transformants induced by N-AcO-AGlu-P-3 contained mutations at codon 61. Three of the codon 61 mutations were at the first base and the other two were at the third base, all were GC→TA transversions. Two AcO-S-induced transformants contained a GC→TA transversion, in one case at the first base of codon 61, in the other at the first base of codon 12. Another AcO-S-induced transformant, and the sixth transformant induced by N-AcO-AGlu-P-3 were apparently not mutated in codon 12, 61 or 117. Both N-AcO-AGlu-P-3 and AcO-S react predominantly with guanine residues in DNA, and all the mutations identified here were at GC base pairs. Further studies were carried out on transformed foci induced in a previous study by reactive derivatives of benzo[a]pyrene and 2-acetylaminoflu-orene and by depurination. Mutations were detected at codons 12 and 13, in addition to those previously characterized at codon 61.

Item Type: Article
Additional Information: Funding Information: The authors wish to thank Dr C.Farr for advice on the PCR technique and K.Merrifield and Dr M.Osborne for synthesizing oligonucleotides. This work was supported by grants to the Institute of Cancer Research from the Medical Research Council and the Cancer Research Campaign.
Uncontrolled Keywords: genetics,toxicology,genetics(clinical),health, toxicology and mutagenesis ,/dk/atira/pure/subjectarea/asjc/1300/1311
Faculty \ School: Faculty of Medicine and Health Sciences > Norwich Medical School
UEA Research Groups: Faculty of Medicine and Health Sciences > Research Groups > Cancer Studies
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Depositing User: LivePure Connector
Date Deposited: 18 Jul 2022 17:30
Last Modified: 23 Oct 2022 04:02
DOI: 10.1093/mutage/3.5.429

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