The induction of several SOS genes of Escherichia coli by fluoroquinolones has been studied. Three different SOS gene fusions (recA::lacZ, umuC::lacZ and sulA::lacZ) have been introduced into the E.coli MC1061 strain to study the induction of these SOS genes in the same genetic background. Data on the basal level of expression of these fusions, as well as their induction by mitomycin C and N-methyl-N'-nitro-N-nitrosoguanidine are presented. Using these strains, we have found that, like nalidixic acid, ofloxacin, enoxacin and ciprofloxacin are strong inducers of the SOS genes tested, umuC gene expression being the highest. Furthermore, fluoroquinolones produced a significant increase in the reversion of the base substitution hisG428 mutation in the TA102 Salmonella tester strain, while no effect was found in strains TA98, TA100, TA1537 and TA1535. These data indicate that the error-prone repair pathway can participate in mutagenesis induced by fluoroquinolones and also that the damage produced by these chemicals may be similar to that produced by nalidixic acid.

The effect of plasmid pKM101 on the survival of Escherichia coli AB1157, growing in minimal medium, in the presence of a 4-quinolone DNA gyrase inhibitor was investigated. The presence of this plasmid decreased susceptibility to the quinolone ciprofloxacin, whereas mucAB genes present in a multicopy plasmid did not. The same effect of pKM101 was detected in a recA430 mutant, confirming that it was not really related to the SOS response. In contrast, when survival assays were performed under amino acid starvation conditions, pKM101 did not confer protection against ciprofloxacin. All of these results indicated that the synthesis of a product(s), different from MucAB, which was encoded by the plasmid pKM101 increased the rate of survival of the AB1157 strain in the presence of quinolone. To identify the gene(s) responsible for this phenotype, several plasmid derivatives carrying different portions of pKM101 were constructed. The 2.2-kb region containing korB, traL, korA, and traM genes was sufficient to decrease susceptibility to quinolone. This plasmidic fragment also made the AB1157 host strain grow more slowly (the Slo phenotype). Moreover, the suppression of the Slo phenotype by addition of adenine to the cultures abolished the decreased susceptibility to quinolone. These results are evidence that the protection against quinolone conferred by this region of pKM101 in strain AB1157 is a direct consequence of the slow growth rate.pKM101 is a 35.4-kb self-transmissible broad-host-range IncN plasmid, which was derived from plasmid R46 (17) by a spontaneous deletion of 14 kb (2, 12). This plasmid has been extensively studied because it contains the mucAB genes (18), which are analogous to the umuDC genes whose products are involved in error-prone repair of DNA damage that can occur as a consequence of the SOS response. The ability of pKM101 to increase bacterial mutability has resulted in its introduction into Salmonella typhimurium strains used in the Ames test, enhancing the sensitivity of this system to detect chemicals as mutagens (16).Besides mucAB, other genes of pKM101 that participate in plasmid replication, stable maintenance, and host range have been described. The conjugal transfer system of pKM101 consists of a mobilization gene cluster, providing functions required for DNA processing and mobilization, and a cluster of tra genes involved in the synthesis of the pilus and putative mating pore (23). Near this second cluster, the kilA and kilB genes were described to encode potentially lethal products whose lethality was prevented by the products of two other genes, korA and korB (24). In addition, the kilA gene, which is defined to a region between two Tn5 insertions, was also thought to retard cell growth of certain Escherichia coli strains on defined minimal medium (24), and this phenomenon had previously been called the Slo phenotype (11,22). More recently, this cluster of tra genes has been sequenced, and, on the basis of DNA sequence information, the kilB (renamed traE), korA, and korB genes have bee...

The mutagenic events induced by ciprofloxacin, a potent antimicrobial agent, have been characterized. For this, a battery of His− mutants of Salmonella typhimurium (hisG428, hisG46, hisC9070, and hisG1775 targets) that detects the six possible transitions and transversions [Levin and Ames (1986): Environ Mutagen 8:9–28] and two additional His− strains (hisC3076 and hisD3052 targets) carrying frameshift mutations have been used. Our results indicate that GC→TA transversions are the major base‐pair substitution induced by ciprofloxacin and that GC→AT transitions are also produced, but to a lesser degree. However, we cannot discard the fact that AT→TA transversions are also induced. In addition, the data indicate that the mutational specificity of ciprofloxacin depends on the location of the target. Intragenic base‐pair substitutions are the most frequent mutations at the hisG428 target when it is on the chromosome, whereas 3 or 6 base‐pair deletions are the major mutagenic events when this target is on the plasmid pAQ1. We have shown that ciprofloxacin also induces deletions/insertions at the hisC3076 and hisD3052 frameshift targets. Therefore, this inhibitor of DNA gyrase promotes a wide pattern of mutations including different kinds of base‐pair substitutions, 3 or 6 base‐pair deletions, and insertions/deletions resulting in frameshifts. All of these mutagenic events require the MucAB proteins involved in the error‐prone repair, with the exception of base‐pair insertions/deletions at the hisD3052 target, which are independent of the presence of plasmid pKM101. © 1996 Wiley‐Liss, Inc.

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Salmonella typhimurium has a SOS regulon which resembles that of Escherichia coli. recA mutants of S. typhimurium have already been isolated, but no mutations in lexA have been described yet. In this work, two different lexA mutants of S. typhimurium LT2 have been constructed on a sulA background to prevent cell death and further characterized. The lexA552 and lexA11 alleles contain an insertion of the kanamycin resistance fragment into the carboxy-and amino-terminal regions of the lexA gene, respectively. SOS induction assays indicated that both lexA mutants exhibited a LexA(Def) phenotype, although SOS genes were apparently more derepressed in the lexA11 mutant than in the lexA552 mutant. Like lexA(Def) of E. coli, both lexA mutations only moderately increased the UV survival of S. typhimurium, and the lexA552 strain was as mutable as the lexA ؉ strain by UV in the presence of plasmids encoding MucAB or E. coli UmuDC (UmuDC Ec ). In contrast, a lexA11 strain carrying any of these plasmids was nonmutable by UV. This unexpected behavior was abolished when the lexA11 mutation was complemented in trans by the lexA gene of S. typhimurium. The results of UV mutagenesis correlated well with those of survival to UV irradiation, indicating that MucAB and UmuDC Ec proteins participate in the error-prone repair of UV damage in lexA552 but not in lexA11. These intriguing differences between the mutagenic responses of lexA552 and lexA11 mutants to UV irradiation are discussed, taking into account the different degrees to which the SOS response is derepressed in these mutants.The SOS regulon, which plays an important role in bacterial responses to DNA damage, has been most extensively studied in Escherichia coli (41). This regulon is composed of more than 20 genes (see reference 12) and is involved in several physiological responses such as DNA repair and mutagenesis, whose expression is transcriptionally regulated by LexA and RecA proteins. LexA repressor binds to similar operators of most SOS genes, including recA and lexA. Treatments that cause DNA damage or stalled DNA replication lead to the activation of the RecA coprotease which facilitates the autocatalytic cleavage of LexA and the subsequent derepression of SOScontrolled genes (see reference 13). The umuDC operon, as well as the analogous mucAB operon, is induced as part of the SOS response, and its products are required for most UV and chemical mutagenesis in E. coli. The UmuD protein is processed to the mutagenically active form UmuDЈ by activated RecA through a mechanism similar to that of the cleavage of LexA (2, 22, 34). The mechanism of SOS mutagenesis is not yet clarified, but biochemical evidence suggests that the UmuCUmuDЈ complex and RecA help DNA polymerase III holoenzyme to facilitate error-prone translesion DNA synthesis (29).The closely related species Salmonella typhimurium has a SOS regulatory system which resembles that of E. coli (37). Several SOS loci such as recA (28), uvrB (31), uvrD (25), sulA (4), and several other din (damage-inducible) loci...