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In the absence of nucleotide excision repair, the additional deficiency of the DNA alkyltransferase (ATase) encoded by the constitutive ogt gene of Escherichia coli caused a marked increase in mutation induction by N-butyl-N-nitrosourea (BNU). Irrespective of the presence or absence of the Ogt ATase, little mutagenic response was detected in Uvr+ bacteria in the concentration range 0-8 mM BNU, indicating that most premutagenic DNA lesions induced at these concentrations are efficiently recognized and repaired by the nucleotide excision repair system. Increased susceptibility to mutagenesis by BNU was detected in Uvr- Ogt+ bacteria, but the Uvr- Ogt- double mutant exhibited much higher sensitivity. These data suggest that the Ogt ATase can replace to a great extent the repair capacity of the (A)BC excinuclease. Forward mutations induced by 6 mM BNU within the initial part of the lacI gene of E.coli were recovered from Uvr+ Ogt-, Uvr- Ogt+ and Uvr- Ogt- bacteria. A total of 454 independent mutations were characterized by DNA sequence analysis. The BNU-induced spectra were dominated by G:C--&gt;A:T transitions, consistent with the major role of the O6-alkylguanine miscoding lesion in mutagenesis by alkylating agents. Specific sites for G:C--&gt;A:T transitions were recovered more or less frequently in one genetic background versus the others, giving statistically significant differences among the spectra (P &lt; 10(-6)). We examined the influence of DNA repair by (A)BC excinuclease and Ogt ATase on the 5'-flanking base associated with the BNU-induced G:C--&gt;A:T transitions; preferences different from those previously reported for other alkylnitrosoureas were detected. We discuss how these differences might be caused by BNU producing branched chain derivatives, in addition to the expected linear chain adducts, and by possible preferences with respect to both the initial distribution of O6-butylguanine lesions and their repairability.

mutagenesis

Role of DNA repair by (A)BC excinuclease and Ogt alkyltransferase in the final distribution of LacI−d mutations induced by N-butyl-N-nitrosourea in Escherichia coli