The relationship between dNTP pool levels and mutagenesis in an Escherichia coli NDP kinase mutant.
Loss of nucleoside diphosphate kinase (Ndk) function in Escherichia coli results in an increased frequency of spontaneous mutation and an imbalance in dNTP pool levels. It is presumed that the imbalance in dNTP pool levels is responsible for the mutator phenotype of an E. coli ndk mutant. A human homologue of Ndk and potential suppressor of tumor metastasis, nm23-H2, can complement the mutagenic phenotype of an E. coli ndk mutant. Here, we show that the antimutagenic property of nm23-H2 in E. coli is independent of dNTP pool levels, indicating that dNTP pool imbalance is not responsible for the mutator phenotype associated with the loss of ndk function. We have identified multiple genetic interactions between ndk and genes involved in the metabolism of dUTP, a potentially mutagenic precursor of thymidine biosynthesis. We show that loss of ndk function is synergistic with a dut-1 mutation and synthetically lethal with the loss of thymidine kinase function. Our results suggest that Ndk prevents the accumulation of dUTP in vivo. Based on these results and biochemical studies of Ndk, we propose that the mutagenic phenotype of an ndk mutant is caused by excess misincorporation of uracil in place of thymidine combined with a defect in the uracil base excision pathway.
Nordman
Jared
J
Department of Molecular Biology and Microbiology, Tufts University School of Medicine, 136 Harrison Avenue, Boston, MA 02111, USA.
Wright
Andrew
A
D004261:DNA Replication; D003854:Deoxyribonucleotides / Q000378:metabolism*; D004926:Escherichia coli / Q000201:enzymology / Q000235:genetics* / Q000378:metabolism*; D005798:Genes, Bacterial; D005816:Genetic Complementation Test; D006801:Humans; D016296:Mutagenesis; D009154:Mutation; D054778:NM23 Nucleoside Diphosphate Kinases / Q000235:genetics / Q000378:metabolism; D009701:Nucleoside-Diphosphate Kinase / Q000235:genetics* / Q000378:metabolism*; D010641:Phenotype; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D013045:Species Specificity; D013936:Thymidine / Q000378:metabolism; D013937:Thymidine Kinase / Q000235:genetics / Q000378:metabolism; D014498:Uracil / Q000378:metabolism
D003854:Deoxyribonucleotides; D054778:NM23 Nucleoside Diphosphate Kinases; D011994:Recombinant Proteins; D014498:Uracil; D013937:Thymidine Kinase; C517938:NME1 protein, human; D009701:Nucleoside-Diphosphate Kinase; D013936:Thymidine