Isolation and characterization of alkylation-induced vermilion mutants in Drosophila melanogaster

278

typical telomere. Our analysis revealed a tandem duplication of parts of chromosome 22, namely part A, composed of the segments 22pll.2-q22.1, and part B, 22q13.1-q13.31. The formula of this marker, which we called tandem-duplicated Philadelphia (mtPh), is as follows: 22pter -A : A : B : A : A : B- -22qter. While the first complex A : A : B has always been observed to form the proximal half of the marker's long arm the second has been seen as well as the A : A : A : B combination. There are always 2 copies of the mtPh in the analysed K562 cells, one having twice the A : A : B combination to form the long arm and the other being variable. There is no cytogenetic evidence for the presence of any parts of chromosome 9, namely 9q34-qter, that are usually involved in the standard 9;22 translocation which generates the classical Ph chromosome. It is likely that the original marker in the K562 ceils derives from chromosome 22 but has formed atypical Ph. The suggested model is in agreement with the data for multiple copies of c-X and c-abl genes (Seldon et al., 1983) and is a new chromosomal manifestation of gene amplification.

24 Nikolov, I.G. ~, I.N. Chernozemsky ~, I.S. Stoyanov ~, M. Castegnaro 2, V. Maru 2, j. Idle 3 and T. Petkova-Bocharova 4, ~ Cancer Cell and Molecular Biology Group, Bulgarian Medical Academy, Sofia (Bulgaria), 2 International Agency for Research on Cancer, Lyon (France), 3 St. Mary's Hospital for Children, London (U.K.) and 4 National Oncological Centre, Sofia (Bulgaria)

Debrisoquine phenotyping and urinary ochratoxin A in patients with Balkan endemic nephropathy (BEN) and urinary tract tumours (uTr) The ability of patients with BEN a n d / o r UTT to metabolise xenobiotics was tested by debrisoquine phenotyping. Over 800 patients and controls were tested. It was found that among BEN patients the percentage of extensive debrisoquine metabolisers was higher than in the control persons. Urinary ochratoxin A and its metabolite 4-hydroxy-ochratoxin A were measured in the same persons. Possible relations be-

tween the metabolism mycotoxin are discussed.

of

debrisoquine

and

25 Nivard, M., E.W. Vogel, A. Pastink and M. Sierra, Department of Radiation Genetics and Chemical Mutagenesis, State University of Leiden, Sylvius Laboratories, Wassenaarseweg 72, 2300 RA Leiden (The Netherlands)

Isolation and characterization of aikylation-induced vermilion mutants in Drosophila melanogaster The major aim of our study was to contribute to an understanding of the molecular action mechanisms on DNA of alkylating carcinogens in higher eukaryotic organisms. Mutants were induced by small alkylating agents at the vermifion locus of Drosophila, isolated by genetic techniques and then their nature was characterized at the DNA level. Due to the fact that the vermilion gene is relatively small (2 kb), and because it codes for an RNA of only about 1.3 kb, vermilion is very suitable for molecular characterization of (chemically) induced mutations. The mutants were analyzed by Southern blotting, which enables the detection of insertions or deletions larger than 50-100 bp, and by DNA-sequencing techniques. This Drosophila system provides us with the possibility of studying the significance of a number of variables in the spectrum of mutations finally recovered. This could be an important tool for understanding at the cellular level some of the action principles of alkylating carcinogens. These variables are: (1) the type of initial DNA adducts: O- versus N-alkylation; (2) the effect of dose: comparison of high exposure levels with low-dose treatment; (3) the repair condition: isolation of mutants in excision repair-proficient or -deficient background; and (4) the time parameter: the significance of the time interval between termination of treatment and first DNA replication. Data will be presented showing results from the analysis of vermilion mutants induced by ENU, DEN, DES and MMS. The most striking observation was that the ethylating carcinogens gave 70% or more transitions while MMS predominantly produced transversions. Clearly, the mutational

279 pattern of the efficient N-alkylator MMS differed drastically from that of the other 3 agents which produce significant levels of O-ethylation adducts in DNA.

tests resulting in greater rates of biotransformation (as measured by increased Asp phase-I/Asp phase-II ratio), D N A binding and genotoxic response.

26 Paolini, M., P. Hrelia and G. Cantelli-Forti, Istituto di Farmacologia dell'UniversitS, Via Irnerio 48, 1-40126 Bologna (Italy)

27 Pasquini, R., G. Scassellati Sforzolini 1, A. Savino 1, S. Monarca 2 and G. Angeli 1, i Department of Hygiene, University of Perugia (Italy) and 2 Chair of Environmental Health, University of Brescia (Italy)

Further evidence on the optical pH for the liver microsomai assay

The aim of this investigation was to optimize the pH in the liver microsomal assay (LMA) as a function of the relative activities and stabilities of the liver microsomal cytochrome P450- and FAD-containing monooxygenase-dependent biotransformation enzymes present in the LMA mixtures. Ethoxyresorufin O-deethylase, dinemorphan N-demethylase, aminopyrine N-demethylase, p-nitroanisole O-demethylase and thiobenzamide S-oxidase were examined in terms of their exact incubation conditions for the LMA during a period of pre-incubation (1 h) over the pH range 0-9. For comparison, the behaviors of glutathione S-transferase (GST) and epoxide hydrase (EH) activities were studied. Lipid peroxidation (LP) was also determined. Experiments were carried out on liver $9 fractions derived from Na-phenobarbital- and fl-naphthoflavone-induced mice. The maximal value of the mean specific activity (Asp) was found at pH 7.8 for the phase-I enzymes (30-45% increase). By contrast, only a small increase in Asp for E H and GST (about 14%) was observed between p H 7.4 and 7.8. LP was not changed appreciably by varying pH. In vitro binding of [1 4 C]dimethylnitrosamine (14C-DMNA) forward calf thymus DNA showed a significant increase in specific activity at p H 7.8 (2.8-fold) compared to pH 7.4. Additional support for the above results has come from mutagenesis experiments using D M N A on the D7 strain of S. cerevisiae. In fact, a significant enhancement of mitotic gene conversion (1.7-fold), mitotic cross-over (2.6-fold) and reverse point mutation (2.3-fold) frequencies was observed at pH 7.8 compared to p H 7.4. These data indicate that pH 7.8 provides a more favorable condition for in vitro mutagenesis

Enzymatic activities of human lung tissue: relationship with smoking habits

Twenty-two $12 preparations of surgical lung specimens obtained from smoker and non-smoker cancer patients were assayed to detect aryl hydrocarbon hydroxylase (AHH), dimethylnitrosalnine demethylase (DMND), and glutathione S-transferase (GST) activities, in both normal and neoplastic lung tissue from the same patients. Pulmonary homogenates were also tested for their ability to activate some precarcinogens into mutagenic metabolites in the Ames test. Statistically significant differences were found for A H H and D M N D activities between normal and neoplastic tissue of smoker patients. In addition, higher A H H activity in the neoplastic tissue of the smoker group was observed compared to that found in the non-smoker group. No differences were found for GST activity. All the lung $12 preparations were able to metabolise water-soluble bases and water-insoluble bases, derived from mainstream cigarette smoke condensate, into mutagenic agents in the Salmonella test system. However, microsomal preparations from smoker neoplastic tissues were more effective. 28 Radoslavova, E., S. Petkova and S. Chankova, Higher Pedagogical Institute, Shumen (Bulgaria), Higher Institute of Agriculture, Plovdiv (Bulgaria) and Institute of Genetics, Sofia (Bulgaria)

Chlorella vulgaris used as a test system to assess the biological and genetic effects of Perocyne and Cuprocyne super