Full text in pdf format

Cytotoxic activation of the pyrrolizidine alkaloid integerrimine in the yeast Saccharomyces cerevisiae is caused by repairable DNA damage

A.L.L. Paula-Ramos^I; C.B. Querol^I; E.K. Marques^II; J.A.P. Henriques^I

^IDepartamento de Biofisica e Centro de Biotecnologia, Instituto de Biociências - UFRGS, Rua Saimento Leite, 500, 90049 Porto Alegre, RS, Brasil. Send correspondence to A.L.L.P.-R.
^IIDepartamento de Genética, Instituto de Biociências - UFRGS, Av. Bento Gonçalves, 9500, Bloco 3-E2, 91501 Porto Alegre, RS, Brasil

ABSTRACT

The cytostatic and cytotoxic effects of integerrimine (ITR), a cyclic diester type pyrrolizidine alkaloid from Senecio brasiliensis Less. var. tripartitus, on Saccharomyces cerevisiae depend on the DNA repair capacity of the strains and on alkaloid concentration. The cytotoxic effect is also dependent on the growth phase of the cells. The high sensitivity of exponentially growing cells to the cytotoxic effect of ITR may be due to the metabolic activation of the alkaloid. The induction of SOS-function by ITR in the SOS-chromotest is also metabolic activation dependent. The sensitivity to ITR of the mutants defective in excision-resynthesis (rad3-e5, rad2-6), mutagenic (rad6-1, rad18-1) and DNA strand break (rad52-1) repair pathways indicates that the lesions induced by this alkaloid are probably bulky adducts, single and double strand breaks or DNA-DNA cross-links. The analysis of cell survival after ITR treatment showed that there are epistatic and non-epistatic interactions of selected DNA repair genes, therefore: (a) a linear sequence of excision and mutagenic repair steps must be generally necessary, and (b) there probably is a co-operation between RAD52 and RAD3 or RAD6 gene products for repair of ITR lesions.

Keywords: Cytotoxic; Pyrrolizidine alkaloid; Saccharomyces cerevisiae; Repairable DNA damage.

REFERENCES

Brendel, M. and Haynes, R.H. (1973). Interactions among genes controlling sensitivity to radiation and alkylation in yeast. Mol. Gen. Genet. 125: 197-216.

Brendel, M. and Ruhiand, A. (1984). Relationships between functionality and genetic toxicology of selected DNA-damaging agents. Mutat. Res. 133: 51-85.

Brendel, M., Khan, N.A. and Haynes, R.H. (1970). Common steps in the repair of alkylation and radiation damage in yeast. Mol. Gen. Genet. 106: 289-295.

Chies, J.M. and Marques, E.K. (1979). Ação mutagênica da integerrimina no sistema de pelos estaminais de Tradescantia. Cienc. Cult. 31: 577.

Chies, J.M. and Marques, E.K. (1980). Ação mutagênica da integerrimina sobre os índices mitóticos dos meristemas radiculares deAllium cepa. Cienc. Cult. 32: 682.

Chlebowicz, E. and Jachymczyk, W.J. (1979). Repair of MMS-induced DNA double-strand breaks in haploid cells of Saccharomyces cerevisiae, which requires the presence of a duplicate genome. Mol. Gen. Genet. 167: 279-286.

Cox, B.S. and Parry, J.M. (1968). The isolation genetics and survival characteristics of ultraviolet-light sensitive mutants in yeast. Mutat. Res. 6: 37-55.

Del Carratore, R., Bronzetti, G., Bauer, C., Corsi, C., Nieri, R., Paolini, M. and Giagoni, P. (1983). Cytochrome P-450 factors determining synthesis in strain D7 Saccharomyces cerevisiae. Mutas. Res. 121: 117-123.

Frayssinet, C. and Moulé, Y. (1969). Effect of lasiocarpine on transcription in the liver cells. Nature 223:1269-1270.

Friedberg, E.C. (1988). Deoxyribonucleic acid repair in the yeast Saccharomyces cerevisiae. Micro b. Reviews 52: 70-102.

Green, M.H.L. and Muriel, W.J. (1975). Use of repair-deficient strains of E. coli and liver microsomes to detect and characterise DNA damage caused by the pyrrolizidine alkaloids heliotrine and monocrataline. Mutai. Res. 28: 331-336.

Guengerich, F.P. and Mitchel, M.R. (1980). Metabolic activation of model pyrroles by cytochrome P-450. Drug Meta bol. Dispos. 8: 34-38.

Haynes, R.H. and Kunz, A.B. (1981). DNA repair and mutagenesis in yeast. In: Molecular biology of the yeast Saccharomyces cerevisiae: life cycle and inheritance. Cold Spring Harbor Laboratory. New York, p. 371-414.

Henriques, J.A.P. and Moustacchi, E. (1981). Interaction between mutation for sensitivity to psoralen photoaddition (pso) and to radiation (rad) in Saccharomyces cerevisiae. J. Bacteriol. 148: 248-256.

Ho, K. (1975). Induction of DNA double-strand breaks by X-rays in a radiosensitive strain of the yeast Saccharomyces cerevisiae. Mutas. Res. 30: 327-334.

Jachymczyk, W.J., Chlebowicz, E., Swietlinska, Z. and Zuk, J. (1977). Alkaline sucrose sedimentation studies of MMS-induced DNA single-strand breakage and rejoining in the wild-type and in UV-sensitive mutants of Saccharomyces cerevisiae. Mutas. Res. 43: 1-10.

Koekemoer, M.J. and Warren, F.L. (1951). The Senecio alkaloids. Part VIII. The occurrence and preparation of the N-oxides. An improved method of extraction of the Senecio alkaloids. J. Chem. Soc. 66-68.

Maron, D.M. and Ames, B.N. (1983). Revised methods for the Salmonella mutagenicity test. Mutas. Res. 133: 173-215.

Mattocks, A.R. (1968). Toxicity of pyrrolizidine alkaloids. Nature 217: 723-728.

Mattocks, A.R. (1972). Toxicity and metabolism of Senecio alkaloids. In: Phytochemical Ecology (Harbone, J.B., ed.). Academic Press, London and New York, p. 179-200.

Melo, A.A., Querol, C.B., Henriques, A.T. and Henriques, J.A.P. (1986). Cytostatic, cytotoxic and mutagenic effects of the voacristine and indole alkaloid in wild type and repair deficient yeasts. Mutas. Res. 171: 17-24.

Moore, C.W. (1978). Responses of radiation-sensitive mutants of Saccharomyces cerevisiae to lethal effects of bleomycin. Mutas. Res. 51: 165-180.

Motidome, M. and Ferreira, P.C. (1966). Alcalóides do Senecio brasiliensis Less. Rev. Fac. Farm. Bioquím. (USP) 4: 13-44.

Moustacchi, E., Favaudon, V. and Bisagni, E. (1983). Likehood of the new antitumoral drug 10-[g-diethylaminopropylamino]-6-methyl-5 H-pyrido [3', 4': 4,5] pyrrolo [2,3 - q] isoquinoline (BD-40), a pyridopyrroloisoquinoline derivative, to induce DNA strand breaks in vivo and its non mutagenicity in yeast. Cancer Res. 43: 3700-3706.

Mowat, M. and Hastings, P.J. (1979). Repair of y-ray induced DNA strand breaks in radiation sensitive (rad) mutants of Saccharomyces cerevisiae. Can. J. Genet. Cytol. 21: 574.

Parry, I.M., Davies, P.J. and Evans, W.E. (1976). The effects of "Cell age" upon the lethal effects of physical and chemical mutagens in the yeast Saccharomyces cerevisiae. Mol. Gen. Genet. 146: 27-35.

Paula-Ramos, A.L.L. de. (1986). Efeitos genotóxico, mutagênico e recombinogênico de alcalóides pirrolizidinicos do gênero Senecio em células procarióticas e eucarióticas. Doctoral Thesis, Universidade Federal do Rio Grande do Sul, Porto Alegre, RS, Brasil.

Paula-Ramos, A.L.L. de and Marques, E.K. (1978). Mutagenic action of integerrimine alkaloid of the Senecio brasiliensis (Sprengel) Less., in Drosophila. Rev. Brasil. Genet. 1: 279-287.

Petry, T.W., Bowden, G.T., Huxtable, R.J. and Sipes, I.G. (1984). Characterization of hepatic DNA damage induced in rats by the pyrrolizidine alkaloid monocrotaline. Cancer Res. 44: 1505-1509.

Prakash, L. (1974). Lack of chemically induced mutation in repair-deficient mutants of yeasts. Genetics 78:1101-1118.

Prakash, L. (1981). Characterization of postreplication repair in Saccharomyces cerevisiae and effects of radio, radl8, rev3 and rad52 mutations. Mol. Gen. Genet. 184: 471-478.

Prakash, S., Prakash, L., Burke, W. and Montelone, B.A. (1980). Effects of the RAD52 gene on recombination in Saccharomyces cerevisiae. Genetics 94: 31-50.

Quillardet, P., Huisman, O., D'An, R. and Hofnung, M. (1982). SOS chromotest, a direct assay of induction of an SOS function in Escherichia coli K-12 to measure genotoxicity. Proc. Natl. Acad. Sci. (USA) 79: 5971-5975.

Resnick, M.A. (1969). Genetic contro1 of radiation sensitivity in Saccharomyces cerevisiae. Genetics 62: 519-531.

Resnick, M.A. and Martin, P. (1976). The repair of double-strand breaks in the nuclear DNA of Saccharomyces cerevisiae and its genetic control. Mol. Gen. Genet. 143: 119-129.

Reynolds, R.J. and Friedberg, E.C. (1981). Molecular mechanism of pyrimidine dimer excision in Saccharomyces cerevisiae: incision of ultraviolet irradiated deosyribonucleic acid in vivo. J. Bacteriol. 146: 692.

Rocha, C.L.M.S.C. and Azevedo, J.L. (1986). Use of the meth GI reversion system of Aspergillus nidulans for the detection of mutagenicity of the pyrrolizidine alkaloid integerrimine. Rev. Brasil. Genet. 9: 393-394.

Rose, E.F. (1972). Senecio species. Toxic plants used as food and medicine in Transkei. South Africa Med. J. 46: 1039-1043.

Ruhland, A. and Brendel, M. (1979). Mutagenesis by cytostatic alkylating agents in yeast strains of differing repair capacities. Genetics 92: 83-97.

Ruhland, A., Haase, E., Siede, W. and Brendel, M. (1981). Isolation of yeast mutants sensitive to the bifunctional alkylating agent nitrogen mustard. Mol. Gen. Genet. 181: 346-351.

Saeki, T., Machida, I. and Nakai, S. (1980). Genetic control of diploid recovery after - irradiation in the yeast. Saccharomyces cerevisiae. Mutas. Res. 73: 251-265.

Singer, B. and Kusmierek, J.T. (1982). Chemical mutagenesis. Ann. Rev. biochem. 52: 655.

Smith, L.W. and Culvenor, C.C.J. (1981). Plant sources of hepatotoxic pyrrolizidine alkaloids. J. Nat. Prod. (LLOYDIA) 44: 129-152.

Wilcox, D.R. and Prakash, L. (1981). Incision and post-incision steps of pyrimidine dimer removal in excision-defective mutants of Saccharomyces cerevisiae. J. Bacteriol 148: 618.

Zimmermann, F.K., Borstel, R.C. von, Halle, E.S. von; Parry, J.M., Siebert, D., Zeiterberg, G., Barale, R. and Loprieno, N. (1984). Testing of chemicals for genetic activity with Saccharomyces cerevisiae: a report of the U.S. Environmental Protection Agency Gene-Tox Program. Mutat. Res. 133: 199-244.