Changes in the template activity of chromatin isolated from sarcoma-180 ascites cells treated with mitomycin C and gamma irradiation in vivo.

The murine ascites sarcoma 180 cells were used to test the in vivo effectiveness of mitomycin C (MMC) and gamma-radiation applied in combination. The action of intraperitoneal administration of MMC and/or whole-body gamma irradiation on sarcoma 180 tumor bearing Swiss albino mice was investigated by studying the template activity of isolated tumor chromatin. The Km value for transcription of 10 Gy-irradiated chromatin was found to decrease with time implying an increase in the template efficiency with respect to that of the unirradiated control. Maximum decrease in Km was observed after 24 h of irradiation. MMC treatment (7 mg/kg body weight of mouse) for 18 h resulted in an inhibition of the transcription rate. Severe inhibition in the template activity was found when cells were subjected to MMC treatment 18 h prior to irradiation with 10 Gy. Susceptibility of tumor chromatin to DNase II followed the same pattern as observed in the case of transcription indicating structural alteration of the treated chromatin. The data showed that DNA damage and its consequences produced in the ascites cells by prior treatment of MMC were not repaired during the 18 h period after which the application of radiation enhanced cytotoxicity.

Test of models for the sequence specificity of UV-induced mutations in mammalian cells.

We have used mathematical modeling and statistical analysis to examine the correlation between UV-induced DNA damage and resulting base-substitution mutations in mammalian cells. The frequency and site specificity of UV-induced photoproducts in the supF gene of the pZ189 shuttle vector plasmid were compared with the frequency and site specificity of base-substitution mutations induced upon passage of the UV-irradiated vector in monkey cells. The hypothesis that the observed mutational spectrum is due to a preferential insertion of adenosine opposite UV photoproducts in the DNA template was found to best explain the mutational data. Models in which it was postulated that only (6-4) photoproducts, and not cyclobutane dimers, are mutagenic, or that the relative frequency of photoproduct formation does not influence mutation frequencies, fit the data much less well. This analysis demonstrates that molecular mechanisms of mutagenesis in mammalian cells can be deduced from mutational data obtained with a shuttle vector system.

Induction and repair of DNA strand breaks in cultured mammalian cells following fast neutron irradiation.

Induction and repair of DNA breaks following irradiation with NIRS cyclotron neutrons were studied in cultured mammalian cells (L5178Y) in comparison to those following gamma-rays. The yield of the total single-strand breaks, 3'OH terminals and sites susceptible to S1 endonuclease following fast neutrons was found to be approximately 50 per cent of that following gamma-irradiation. On the other hand, the yield of double-strand breaks was slightly higher after fast neutrons than after gamma-rays. The percentage of the total single-strand breaks remaining unrejoined at 3 hours after post-irradiation incubation was found to be distinctly higher after the fast neutrons than after gamma-rays. The neutron-induced damage appears to carry a higher proportion of alkali-labile lesions compared to gamma-rays. It was concluded that the increase in the yield of double-strand breaks and of unrejoinable breaks is responsible for a high r.b.e. of the cyclotron neutrons.

Effects of irradiation on RNA synthesis primed by calf thymus deoxyribonucleoprotein treated with salt and salt-urea.

1. The effects of ionizing radiation on the activity of calf thymus templates were examined in a Escherichia coli RNA polymerase system. 2. The template activity of native and 2 M NaCl-5M urea-treated deoxyribonucleoproteins was enhanced by relatively low doses of irradiation, while that of 2 M NaCl-treated deoxyribonucleoprotein was not enhanced by irradiation. 3. The template activity of purified DNA was markedly decreased by irradiation, while that of native deoxyribonucleoprotein, 2 M NaCl-treated, and 2 M NaCl-5 M urea-treated ones were slightly decreased at a higher dose range. The doses for 50% inactivation of these templates were 1.3, 210, 140, and approximately 200 krad, respectively.