The RAD4 gene of Saccharomyces cerevisiae is essential for the incision step of UV-induced excision repair. A yeast RAD4 gene has been previously isolated by functional complementation. In order to identify the RAD4 homologous gene from fungus Coprinus cinereus, we have constructed cosmid libraries from electrophoretically separated chromosomes of the C. cinereus. The 13 C. cinereus chromosomes were resolved by pulse-field gel electrophoresis, hybridized with S. cerevisiae RAD4 DNA, and then isolated homologous C. cinereus chromosome. The insert DNA of the RAD4 homolog was contained 3.2 kb. Here, we report the partial cloning and characterization of fungus C. cinereus homolog of yeast RAD4 gene. Southern blot analysis confirmed that C. cinereus contains the sequence homologous DNA to RAD4 gene and this gene exists as a single copy in C. cinereus genome. When total RNA isolated from C. cinereus cells was hybridized with the 1.2 kb PvuII DNA fragment of the S. cerevisiae RAD4 gene, a 2.5 kb of transcript was detected. The level of the transcript did not increase upon UV-irradiation, suggesting that the RAD4 homologous gene in C. cinereus is not UV-inducible.

Variations in adriamycin uptake and cytotoxicity were studied in tumor cells that were grown in different growth states and microenvironments. RIF-1 tumor cells were maintained in an RPMI 1640 medium, and grown in either a monolayer or multicell spheroids. For exponentially growing cells, adriamycin cytotoxicity increased with increased dosage up to 2.5 $\mu$g/ml, and this cytotoxicity was reduced when the cells were grown in a plateau phase or in an acidic microenvironment (pH 6.6). This reduced cytotoxicity was correlated with the uptake of the drug. For multicell spheroids, the cytotoxicity of the drug was reduced dramatically, and this reduction was also correlated with a reduced uptake of the drug and an acidic pH inside of the spheroids. When the drug cytotoxicity was evaluated at different locations within the spheroids, the cells in the inner regions were least affected by the drug, suggesting that both an acidic microenvironment and noncycling plateau phase cells are contributing factors in decreasing the efficacy of the drug in an organized tissue, such as multicell spheroids.

Chromosome rearrangement induced by bleomycin were identified by fluorescence in situ hybridization with probe for chromosome 4. The frequency of color junctions, translocations, dicentric and acenttic fragments increased with bleomycin dose. Different types of balanced translocation and dicentric were scored and compared. The frequency of cells exhibiting multiple aberration was higher compared to that of cells exposed to Gamma radiation suggesting that effect of bleomycin might be similar to that of high LET radiation.

We observed the frequency of chromosome aberrations induced by UVB irradiations, and the suppressing effect of tannic acid on chromosome aberrations induced by UVB irradiations in CHL cells, which is a phenolic compound, a hydrolysate of tannin and a components of green tea. UVB doses used for the frequency of chromosome aberrations were from 0.2 to 1.6 KJ/m$^2$ and tannic acid concentrations were from 1.16 $\mu$g/ml to 37.50 $\mu$g/ml. For the observation of suppressing effect of tannic acid on UVB-induced chromosome aberrations, UVB dose was 1.6 KJ/m$^2$ and tannic acid concentrations were 1.0, 2.0, 4.0 $\mu$g/ml. In our study, tannic acid was treated for 24 hours in CHL, cells after UVB irradiation without S9 mix or for 6 hours with S9 mix. From this study, we obtained the following results : (1) The frequency of chromosome aberrations UVB induced were dose-dependently increased. (2) The tannic acid did not induce chromosome aberrations in cultured Chinese hamster cells. (3) UVB-induced chromosome aberrations were suppressed by tannic acid at every concentration from 1.0 $\mu$g/ml to 4.0 $\mu$g/ml with or without metabolic activation. These results suggest that the tannic acid acts as an inhibitor to UVB-induced clastogenicity of the cultured cell.