Mutator phenotype in a mutant of S49 mouse T-lymphoma cells with abnormal sensitivity to thymidine.
We have selected and characterized a thymidine-sensitive S49 mutant line, MC-3-3. MC-3-3 cells are 35-fold more sensitive to the cytotoxic effects of thymidine and 15-fold more sensitive to the cytotoxic effects of 5-bromodeoxyuridine than wild type S49 cells. In contrast, the MC-3-3 mutant line does not exhibit increased sensitivity to the cytotoxic action of 5-fluorodeoxyuridine. The MC-3-3 mutant line possesses levels of thymidylate synthetase and thymidine kinase activity which are equivalent to the levels in wild type S49 cells, but the ribonucleotide reductase activity in MC-3-3 cells, using CDP as a substrate, is only 10-30% of that in wild type cells. Using ADP as a substrate, the ribonucleotide reductase activity in permeabilized MC-3-3 cells is slightly higher than that in wild type S49 cells. The deoxyribonucleotide pools in exponentially growing MC-3-3 cells are approximately 40-50% of those in wild type S49 cells. By hybrid analysis, we determined that the thymidine sensitivity of the MC-3-3 cells is recessive. The MC-3-3 mutant line displays a rate of spontaneous mutation which is 15-30-fold higher than that of wild type S49 cells. The MC-3-3 mutant cells are also 5-10-fold more sensitive than wild type cells to the cytotoxic effects of tunicamycin and compactin. These results suggest that the MC-3-3 mutant line possesses a mutation in the dTTP binding site in ribonucleotide reductase; abnormal regulation of this enzyme results in an increase in the rate of spontaneous mutation.
D000818:Animals; D001973:Bromodeoxyuridine / Q000633:toxicity; D002460:Cell Line; D002470:Cell Survival / Q000187:drug effects; D002999:Clone Cells; D003853:Deoxyribonucleosides / Q000633:toxicity; D003854:Deoxyribonucleotides / Q000378:metabolism; D006822:Hybrid Cells / Q000502:physiology; D007700:Kinetics; D008223:Lymphoma / Q000235:genetics* / Q000503:physiopathology; D051379:Mice; D009154:Mutation; D013936:Thymidine / Q000633:toxicity*
D003853:Deoxyribonucleosides; D003854:Deoxyribonucleotides; D001973:Bromodeoxyuridine; D013936:Thymidine