Abstract
It is still an open question whether mutation rate can vary across the bacterial chromosome. In this study, the occurrence of mutations within the same mutational target sequences at different chromosomal locations of Pseudomonas putida was monitored. For that purpose we constructed two mutation detection systems, one for monitoring the occurrence of a broad spectrum of mutations and transposition of IS element IS1411 inactivating LacI repressor, and another for detecting 1-bp deletions. Our results revealed that both the mutation frequency and the spectrum of mutations vary at different chromosomal positions. We observed higher mutation frequencies when the direction of transcription of the mutational target gene was opposite to the direction of replisome movement in the chromosome and vice versa, lower mutation frequency was accompanied with co-directional transcription and replication. Additionally, asymmetry of frameshift mutagenesis at homopolymeric and repetitive sequences during the leading and lagging-strand replication was found. The transposition frequency of IS1411 was also affected by the chromosomal location of the target site, which implies that regional differences in chromosomal topology may influence transposition of this mobile element. The occurrence of mutations in the P. putida chromosome was investigated both in growing and in stationary-phase bacteria. We found that the appearance of certain mutational hot spots is strongly affected by the chromosomal location of the mutational target sequence especially in growing bacteria. Also, artificial increasing transcription of the mutational target gene elevated the frequency of mutations in growing bacteria.
Highlights
Mutations provide a raw material for population diversity and adaptation; the mutation rate has profound effects on population diversity and evolution dynamics
These test systems have not been applicable for the detection of mutations in the bacterial chromosome because of too low mutation frequency
To elucidate mechanisms which could affect the frequency of mutations at different chromosomal positions, we examined the effect of orientation of the target gene on mutagenic processes in the bacterial chromosome
Summary
Mutations provide a raw material for population diversity and adaptation; the mutation rate has profound effects on population diversity and evolution dynamics. Over the years, a number of detection systems have been developed to study mutagenesis in bacteria [2]. Some of these mutation detection systems allow to monitor mutations in plasmids whereas others in bacterial chromosome. Measurement of lac reversion frequencies for the two orientations in the chromosome of E. coli in the absence of DNA mismatch repair indicated that the lagging strand replication may be more accurate than leading strand replication [9,10]. Under the conditions of constitutive expression of the SOS system the replication of the lagging strand appeared to be the major source of the mutations [11], whereas the following studies demonstrated lack of strand bias in UV-induced mutagenesis in E. coli [12]
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