Tetsuo Ono

Mutagenicity of metal cations in cultured cells from Chinese hamster.

The mutations at the hypoxanthine-guanine phosphoribosyl transferase (HGPRTase) locus in Chinese hamster V79 cells induced by metal cations were examined by the development of resistance to 8-azaguanine (8AG). The spontaneous frequency of 8AG resistance was 5.8 per 10(6) cells, and the frequency was enhanced to 2-6 times that of the control by treatment of cells with the chlorides of beryllium and manganese. About 75% of 8AG-resistant colonies were sensitive to amethopterin, and 86% of the resistant colonies showed less than 3% of the HGPRTase activity of the wild-type cells. The mutant frequency in cultures treated with cobalt and nickel chlorides were slightly increased, and mutation induction was only detectable at very low rates of cell survival.

Effect of metal cations on misincorporation by E. coli DNA polymerases

Summary The effects of divalent metal cations on DNA polymerase activity of E. coli polymerase I, II or III and on their fidelities during DNA synthesis in vitro have been examined. Carcinogenic or mutagenic metal at the concentration which decreases polymerase activity increased misincorporation, whereas noncarcinogenic metal did not change the fidelity even at a concentration inhibiting polymerase. The change in 3′ → 5′ exonuclease activity of polymerase by metal ions was not responsible for the increase in the misincorporation. The influences of metals on Km for the substrate and template-primer were also observed.

Strand breaks of mammalian mitochondrial DNA induced by carcinogens

Closed circular mitochondrial DNA in mammalian cells was degradated to the open circular form by exposure of the cells to the carcinogens N-methyl-N-nitro-N-nitrosoguanidine (MNNG) and 4-nitroquinoline 1-oxide (4NQO). MNNG caused more strand scission of mitochondrial DNA than 4NQO at the same concentration. The action of the carcinogens on mitochondrial DNA did not parallel that with nuclear DNA which was damaged by 4NQO more markedly than by MNNG. Mitochondrial DNA damaged by carcinogens was not repaired during 4-20 h of post-treatment incubation of the cells. Incorporation of labeled thymidine into the closed circular mitochondrial DNA, decreased by the treatment of cells with carcinogens, recovered during post-treatment incubation.

Chemistry and biological actions of 4-nitroquinoline 1-oxide

1 Introduction.- 2 Chemical Properties.- 3 Biophysics.- 4 Carcinogenicity.- S Metabolism.- 6 Molecular Aspects of the Action.- 7 Anti-tumor Effect.- 8 Microbiology.- References.

UV-induced imbalance of the deoxyribonucleoside triphosphate pool in E. coli

The effect of UV irradiation on the intracellular DNA precursor pool in E. coli was investigated. UV irradiation of E. coli, followed by post-incubation for 1-1.5 h, altered the relative sizes of the deoxyribonucleoside triphosphate (dNTP) pool. The total amount of dNTPs increased: both dATP and dTTP increased several-fold, dCTP about twofold, while dGTP remained almost unchanged. In recA- and umuC- strains, which are defective in UV-induced mutagenesis, the pattern of nucleotide pool alterations was similar to that of wild-type strains.

Susceptibility of skin fibroblasts from patients with retinoblastoma to transformation by murine sarcoma virus.

Cultured skin fibroblasts from patients with retinoblastoma (RB) of different etiology have been studied for their susceptibility to transformation by murine sarcoma virus. The cells from patients with a deletion in chromosome 13 (13q-) and those from sporadic unilateral cases due to somatic mutation were as sensitive as normal cells. However, the cells from familial cases showed an extremely high sensitivity to transformation. Moreover, in familial cases the susceptibility was significantly higher in bilaterally affected patients than in unilateral cases. These findings suggest that the heritable RB gene is different from 13q- and its degree of expression is also manifest at the cellular level.

Misincorporation in DNA synthesis after modification of template or polymerase by MNNG, MMS and UV radiation

The synthetic DNA polymers, poly(dG-dC), poly(dC), poly(dA-dT), poly(dA) and poly(dT), were treated with N-methyl-N-nitro-N-nitrosoguanidine (MNNG), methyl methanesulfonate (MMS) and UV irradiation. The modified polymers were used as templates to examine the incorporation of non-complementary nucleotides by E. coli DNA polymerase I. Methylation of poly(dG-dC) by MNNG predominantly induced the misincorporation of dTMP, whereas methylation by MMS induced that of dAMP. Treatment of poly(dT) with MNNG caused the misincorporation of dGMP to a considerable extent, but MMS did not enhance the error on poly(dT). The misincorporation of dAMP on poly(dC) and that of dGMP on poly(dA) were also increased by these chemicals. UV irradiation of poly(dT) and poly(dC) induced the error of dGMP and dAMP, respectively. These data on MNNG and MMS in vitro were in fair agreement with the directions of mutation in vivo. But the predominant induction of transitions by UV in vitro did not agree with the UV-induced transversions in E. coli. This inconsistency suggested the participation of other factors than direct mispairing in UV-induced transversion. Modification of DNA polymerase I by MNNG changed the ratio of polymerase to 3 leads to 5 exonuclease activity altering the fidelity of this enzyme, whereas MMS and UV-irradiation did not alter the fidelity of the enzyme.

Enhancement of DNA polymerase II activity in E. coli after treatment with N-methyl-N′-nitro-N-nitrosoguanidine

Abstract Treatment of E. coli P3478 (polAl, thy−) with N-methyl-N′-nitro-N-nitrosoguanidine followed by incubation of cells in growth medium enhanced DNA polymerase II activity up to 5 times as high as that of control cells, in contrast to slight change in polymerase III activity. This enhancement was inhibited by chloramphenicol added to the post-treatment incubation medium.

Effect of medium change on poly(ADP-ribose) synthesis in friend erythroleukemic cells

Abstract In Friend leukemic cells cultured in the presence of 5 mM hexamethylene bisacetamide, a potent differentiation-inducer, poly(ADP-ribose) synthesis was reduced to about one-third of that in control cells. Replacing the original culture medium with fresh medium resulted in a decrease of poly(ADP-ribose) synthesis in confluent control cultures, while cells induced to differentiate were not affected by the medium change. This is not attributable to the difference of the level of poly(ADP-ribose) synthesis in different cell cycle stages, since DNA synthesis and cell growth in differentiating cells were maintained at the same level with those of control cells. In control cultures, a medium change during the log-phase effected a prolongation in the rise of poly(ADP-ribose) synthesis. When conditioned medium was substituted during log-phase growth, poly(ADP-ribose) synthesis was stimulated in control cells. This stimulating effect was not lost by dialysis but was lost by heat-treatment or trypsin-digestion. Results suggest that poly(ADP-ribose) synthesis is regulated by some factor(s) released into the culture medium.

Mechanism of increased susceptibility to 4-nitroquinoline-1-oxide in cultured skin fibroblasts from patients with familial polyposis coli

About 50% of the strains of cultured fibroblasts from patients with familial polyposis coli (FPC) exhibited increased susceptibility to cytotoxicity of 4-nitroquinoline-1-oxide (4NQO) compared with cells from normal individuals. The FPC cells that showed hyper-sensitivity to 4NQO were also hyper-sensitive to mitomycin C (MMC), but susceptibilities of these cells to UV radiation, methyl methanesulfonate (MMS) and N-methyl-N-nitro-N-nitrosoguanidine (MNNG) were within the normal range. The extent of single-strand scission of DNA in the 4NQO-sensitive FPC cells was greater than in normal cells, and the amount of [14C]4NQO bound to DNA in the FPC cells was twice as high as in normal cells. The rate of release of [14C]4NQO from DNA by the post-culture was the same as in both FPC and normal cells. The 4NQO-sensitive FPC cells exhibited increased 4NQO-reductase activity; the level of this activity was consistent with the extent of the decrease in colony formation by 4NQO. These results suggest that the enhanced ability to activate 4NQO might be an important factor in the mechanism of susceptibility of FPC cells to 4NQO rather than the reduced ability to repair DNA.