ENHANCEMENT OF BACTERIAL MUTAGENICITY OF BIFUNCTIONAL ALKYLATING-AGENTS BY EXPRESSION OF MAMMALIAN GLUTATHIONE-S-TRANSFERASE

Recently, we inserted the plasmid vector pKK233-2 containing rat GSH S -transferase (GST) 5-5 cDNA into Salmonella typhimurium TA1535 and fou nd that these bacteria [GST 5-5(+)] expressed the protein and produced mutations when ethylene or methylene dihalides were added [Thier, R., Taylor, J. B., Pemble, S. E., Ketterer, B., Persmark, M., Humphreys, W. G., and Guengerich, F. P. (1993) Proc. Natl. Acad. Sci. U.S.A. 90, 8576-85803. After exposure to the known GST 5-5 substrate 1,2-epoxy-3- (4'-nitrophenoxy)propane, the GST 5-5(+) strain showed fewer mutants t han the bacteria transfected with the cDNA clone in a reverse orientat ion [GST 5-5(-)], suggesting a protective role of GST 5-5. However, mu tations were considerably enhanced in the GST 5-5(+) strain [as compar ed to GST 5-5(-)] when 1,2,3,4-diepoxybutane (butadiene diepoxide) or 1,2-epoxy-4-bromobutane was added. The GST 5-5(+) and GST 5-5(-) bacte rial stains showed similar responses to 1,2-epoxypropane, 3,4-epoxy-1- butene, and 1,4-dibromobutane. The results suggest that some bifunctio nal activated butanes are transformed to mutagenic products through GS H conjugation. We also found that the GST 5-5(-) strain showed enhance d mutagenicity with 1,4-dibromo-2,3-epoxybutane, 1,2-epoxy-3-bromoprop ane (epibromohydrin), and (+)-1,4-dibromo-2,3-dihydroxybutane. The pos sibility was considered that a 5-membered thialonium ion may be involv ed in the mutagenicity. Model thialonium compounds were rather stable to hydrolysis in aqueous solution at pH 7.4 and slowly alkylated 4-(4- nitrobenzyl)pyridine. The presence of a hydroxyl group beta to the sul fur did not enhance reactivity. Mechanisms involving episulfonium ions are considered more likely. Potential. oxidation products of the toxi c pesticide 1,2-dibromo-3-chloropropane (DBCP) were also considered in this system. DBCP itself gave rather similar results in the two strai ns. Others have reported that oxidation of DBCP is required for mutage nicity, along with GST-catalyzed GSH conjugation [Simula, T. P., Glanc ey, M. J., Soderlund, E. J., Dybing, E., and Wolf, C. R. (1993) Carcin ogenesis 14, 2303-2307]. The putative oxidation product 1,2-dibromopro pional did not show a difference between the two strains. However, 1,3 -dichloroacetone, a model for the putative oxidation product l-bromo-3 -chloroacetone, was considerably more mutagenic in the GST 5-5(+) stra in.