Ferrezuelo F, Prieto-Alamo M J, Jurado J, Pueyo C
Departamento de Bioquímica y Biología Molecular, Universidad de Córdoba, Spain.
Mutagenesis. 1998 Sep;13(5):507-14. doi: 10.1093/mutage/13.5.507.
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-->A:T transitions, consistent with the major role of the O6-alkylguanine miscoding lesion in mutagenesis by alkylating agents. Specific sites for G:C-->A:T transitions were recovered more or less frequently in one genetic background versus the others, giving statistically significant differences among the spectra (P < 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-->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.
在缺乏核苷酸切除修复的情况下，大肠杆菌组成型ogt基因编码的DNA烷基转移酶（ATase）的额外缺陷导致N-丁基-N-亚硝基脲（BNU）诱导的突变显著增加。无论是否存在Ogt ATase，在0-8 mM BNU浓度范围内，Uvr+细菌中几乎未检测到诱变反应，这表明在这些浓度下诱导的大多数前诱变DNA损伤能被核苷酸切除修复系统有效识别和修复。在Uvr- Ogt+细菌中检测到对BNU诱变的易感性增加，但Uvr- Ogt-双突变体表现出更高的敏感性。这些数据表明，Ogt ATase在很大程度上可以替代（A）BC核酸外切酶的修复能力。从Uvr+ Ogt-、Uvr- Ogt+和Uvr- Ogt-细菌中回收了由6 mM BNU在大肠杆菌lacI基因初始部分诱导的正向突变。通过DNA序列分析对总共454个独立突变进行了表征。BNU诱导的谱以G:C→A:T转换为主，这与O6-烷基鸟嘌呤错配损伤在烷基化剂诱变中的主要作用一致。在一种遗传背景与其他遗传背景相比，G:C→A:T转换的特定位点或多或少频繁出现，导致谱之间存在统计学显著差异（P < 10(-6)）。我们研究了（A）BC核酸外切酶和Ogt ATase的DNA修复对与BNU诱导的G:C→A:T转换相关的5'-侧翼碱基的影响；检测到与先前报道的其他烷基亚硝基脲不同的偏好。我们讨论了这些差异可能是由BNU除了产生预期的线性链加合物外还产生支链衍生物，以及关于O6-丁基鸟嘌呤损伤的初始分布及其可修复性的可能偏好所引起的。
