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The recA1 mutation was transduced into the tag-2 mutant of E. coli, thus making a strain deficient in the induction of SOS repair as well as in the constitutive repair of 3-alkylated adenines in DNA. The double mutant recA tag is more sensitive to methyl methanesulfonate exposure than either single mutant, indicating that recA and tag mutations block different pathways in repair of alkylation damage. The double mutant is more deficient in host cell reactivation of alkylated phages than the tag single mutant. However, alkylation induction of the double mutant with N-methyl-N'-nitro-N-nitrosoguanidine resulted in killing adaptation of the cells to methyl methanesulfonate and restored the host cell reactivation capacity for alkylated lambda phage to wild-type levels. These adaptive responses can be ascribed to the induction of 3-methyladenine DNA glycosylase II which is shown by enzyme analysis to proceed normally in the recA mutant background. The results imply that the induction of the alkA gene encoding 3-methyladenine DNA glycosylase II is independent of SOS induction.

The ada gene of Escherichia coli K12, the regulatory gene for the adaptive response of bacteria to alkylating agents, was cloned in multicopy plasmids. O6-Methylguanine-DNA methyltransferase and 3-methyladenine-DNA glycosylase II, which are known to be inducible as part of the adaptive response, were produced in ada- cells bearing ada+ plasmids, even without treatment with alkylating agents. When such cells had been treated with methyl methanesulfonate, even higher levels of the enzyme activities were produced. Maxicell experiments revealed that the ada gene codes for a polypeptide with a molecular weight of 38 000. We constructed a hybrid plasmid carrying an ada'-lacZ' fused gene, with the proper control region for ada expression. beta-Galactosidase synthesis from the fused gene was strongly induced only when cells were treated with low doses of methylating agents, but was weakly induced with relatively high doses of ethylating agents. The induction was autogenously regulated by the ada gene product, in a positive manner.

We used alkA'-lacZ' and umuC'-lacZ' fused genes and determined the ability of various alkylating agents to induce adaptive and SOS responses. The degree of induction of expression of these genes was quantitatively measured by a simple colorimetric assay of beta-galactosidase activity. SN1 type methylating agents, such as N-methyl-N'-nitro-N-nitrosoguanidine and N-methyl-N-nitrosourea, were more effective inducers for the alkA than for the umuC system, while SN1 type ethylating agents, such as N-ethyl-N'-nitro-N-nitrosoguanidine and N-ethyl-N-nitrosourea, were more potent inducers for the umuC than for the alkA system. Similar but less striking effects on the two systems were obtained with SN2 type alkylating agents.