EFFECT OF OVEREXPRESSION OF ESCHERICHIA-COLI 3-METHYLADENINE-DNA GLYCOSYLASE-I (TAG) ON SURVIVAL AND MUTATION-INDUCTION IN SALMONELLA-TYPHIMURIUM

Salmonella typhimurium, compared to Escherichia coli, is deficient in an inducible glycosylase activity harbouring only constitutive glycosy lase functions. 3-Methyladenine-DNA glycosylase I encoded by the E. co li tag gene is a constitutively expressed repair enzyme that primarily removes N-3-methyladenine but also N-3-methylguanine from DNA by glyc osylic cleavage in the first step of the base excision repair. In orde r to investigate in vivo effect of the overexpressed glycosylase I act ivity on survival capacity and mutation induction in S. typhimurium, a nd thereby elucidate the significance of both 3-methylpurines in cellu lar sensitivity to methylating agents (e.g., DMS), we transformed four his(-) S. typhimurium strains with the plasmid pCY5 carrying the E. c oli tag gene under the control of the lac promoter. Although the 3-met hyladenine DNA glycosylase activity in cells carrying pCY5 was only 10 -fold higher on exposure to IPTG compared to the TA1535 control strain carrying pUC8, the overexpression of the Tag protein completely suppr essed deficiency in an inducible glycosylase activity, rendering cells resistance to toxic effects of DMS. The suppression was not influence d by the nucleotide excision repair pathway since there was no differe nce in recovered survival among NER-proficient and NER-deficient strai ns. The yield of mutation induction in the reversion assay was decreas ed to the level of spontaneous (his(-) --> his(+)) revertant colonies showing that in the overall population in overexpressed conditions in vivo 3-methylguanine, in addition to 3-methyladenine, must have been r emoved from DNA by the E. coli Tag protein and therefore accounts for the second most important cytotoxic lesion.