Differential homeostatic supercoil regulation in Escherichia coli and Salmonella enterica serovar Typhimurium.

摘要

Escherichia coli and Salmonella enterica serovar Typhimurium are closely related gram-negative bacterium. They share a high degree of homology among conserved genes and have very similar genetic maps. In addition, most defined processes are nearly indistinguishable between the two. However, several persisting and unexplained differences exist in the literature. Paramount among these is the observation that a deletion of the gene encoding topoisomerase I is lethal in E. coli without a compensatory mutation, but viable in Salmonella. To address this disparity, we examined the effect of the Salmonella gyrB652 allele in E. coli. The gyrB652 transversion has a complex phenotype in Salmonella as a result of an inefficient DNA gyrase enzyme. We found that this mutation, which is viable in Salmonella, is lethal in E. coli. We also found that E. coli, but not Salmonella, is very sensitive to altered DNA gyrase levels. This led us to compare the level of negative supercoiling in each species. Using two-dimensional chloroquine electrophoresis, we found that E. coli contains about 15% more unconstrained negative supercoiling than Salmonella. We confirmed these measurements using supercoil-sensitive chromosomal reporter cassettes. We further demonstrated this by showing phenotypic differences in physiological systems that have a supercoil component in their regulation. These include chromosome condensation via the SMC protein MukB, replication initiation via SegA, and the lysis/lysogeny pathway of phage Mu. Further analysis showed that both species have a similar chromosomal supercoil fluctuation pattern through the growth phase. However, the supercoil fluctuations of plasmid DNA in the two species exhibited a stark difference. This may indicate that, in addition to differences in global supercoiling levels, there may be differences in plasmid maintenance between the species.