Regulation of mutY and Nature of Mutator Mutations in Escherichia coli Populations under Nutrient Limitation

Abstract

Previous analysis of aerobic, glucose-limited continuous cultures of Escherichia coli revealed that G:C-to-T:A (G:C3T:A) transversions were the most commonly occurring type of spontaneous mutation. One possible explanation for the preponderance of these mutations was that nutrient limitation repressed MutY-dependent DNA repair, resulting in increased proportions of G:C3T:A transversions. The regulation of the mutYdependent DNA repair system was therefore studied with a transcriptional mutY-lacZ fusion recombined into the chromosome. Expression from the mutY promoter was fourfold higher under aerobic conditions than under anaerobic conditions. But mutY expression was higher in glucose-or ammonia-limited chemostats than in nutrient-excess batch culture, so mutY was not downregulated by nutrient limitation. An alternative explanation for the frequency of G:C3T:A transversions was the common appearance of mutY mutator mutations in the chemostat populations. Of 11 chemostat populations screened in detail, six contained mutators, and the mutator mutation in four cultures was located in the region of mutY at 66 min on the chromosome. The spectrum of mutations and rate of mutation in these isolates were fully consistent with a mutY-deficiency in each strain. Based on PCR analysis of the region within and around mutY, isolates from three individual populations contained deletions extending at least 2 kb upstream of mutY and more than 5 kb downstream. In the fourth population, the deletion was even longer, extending at least 5 kb upstream and 5 kb downstream of mutY. The isolation of mutY mutator strains from four independent populations with extensive chromosomal rearrangements suggests that mutY inactivation by deletion is a means of increasing mutation rates under nutrient limitation and explains the observed frequency of G:C3T:A mutations in glucose-limited chemostats.There is an increasing awareness that mutational processes in bacteria are under environmental control or at least susceptible to stress responses (3,9,12,13,35,37). Most of the studies on these influences used batch cultured bacteria or colonies on plates with their constantly changing or spatially nonuniform environments. The complexity of the stress, be it starvation, slow growth or external challenge, can be simplified by using a steady-state application of particular stresses as can be achieved in chemostat culture (14). As an initial study in this direction, the mutational changes in three genes were analyzed in glucose-limited chemostats of Escherichia coli (23,24). Mutations in malT, mlc, and mgl genes were identified and consisted of mainly base changes under steady-state glucose limitation. The majority of nucleotide substitutions were G:Cto-T:A (G:C3T:A) transversions in all three genes. This communication addresses possible causes of the mutational spectrum observed under glucose limitation, since it differs markedly from the generally observed spectrum of spontaneous mutations in E. coli (29,34).A clue as to the origin of malT, mlc, and mgl ...

Ex Uno Plures: Clonal Reinforcement Drives Evolution of a Simple Microbial Community

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