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SEQUENCE ANALYSIS OF MUTATIONS INDUCED BY N-ETHYL-N-NITROSOUREA IN THE TK AND HPRT GENES OF MOUSE LYMPHOMA CELLS.
Citation:
Chen, T., K H. Brock, AND M M. Moore. SEQUENCE ANALYSIS OF MUTATIONS INDUCED BY N-ETHYL-N-NITROSOUREA IN THE TK AND HPRT GENES OF MOUSE LYMPHOMA CELLS. Presented at Environmental Mutagen Society Meeting, New Orleans, LA, April 9-14, 2000.
Description:
The mouse lymphoma assay is widely used to identify chemicals that are capable of inducing mutational damages. The Tk+/- gene located on an autosome in mouse lymphoma cells may recover a wider range of mutational events than the X-linked Hprt locus. However, chemical-induced mutation spectra in the Tk gene of mouse lymphoma cells have not been characterized. In this study, we have identified more than 50 ENU induced mutations in the Tk gene of mouse lymphoma cells and compared them with those induced in the Hprt gene and the spontaneous Tk mutations. Treatment of mouse lymphoma cells with 100 ug/ml N-ethyl-N-nitrosourea (ENU) resulted in both Tk and Hprt mutation induction. The mutant frequencies induced by ENU were 756 x 10-6 in Tk and 311 x 10-6 in Hprt. Sequence analysis of Tk and Hprt mutant cDNA produced similar overall mutation patterns in the two genes. ENU induced primarily base pair substitutions (72% in Tk and 75% in Hprt) and exon splice mutations (29% in Tk and 25% in Hprt). The most common base pair substitution was G:C > T:A transversion (28% of independent mutations in both of Tk and Hprt genes). In contrast to ENU induced mutations, the spontaneous mutations in the Tk gene showed a different pattern. About 30% of mutations were in-frame deletions in the control, whereas no frameshifts were found in ENU-induced Tk mutants. Seventy five percent of ENU-induced Tk base pair substitutions occurred at the mutated guanine and thymidine (possible targets of ENU binding) on the non-transcribed DNA strand, while no such strand-related bias was found in the spontaneous Tk mutants. The results suggest that the Tk locus in mouse lymphoma cells is a useful reporter of mutation spectra induced by chemicals.