engleski

The rus-1 mutation suppresses cytological defects in recBC sbcBC ruv mutants of Escherichia coli

Background and purpose: The RuvABC proteins of Escherichia coli play an important role in the processing of Holliday junctions during homologous genetic recombination. We have previously found that a ruvC mutation drastically reduces cell viability and causes severe defects in chromosome segregation and cell division when introduced into the recBC sbcBC strain of E. coli. The aim of this work was to further study the mechanisms leading to morphological defects and reduced viability in recBC sbcBC ruv mutants. For this purpose, we have examined the effects of elevated rusA gene expression on cytological patterns of these mutants (the rusA gene codes for a resolvase that can substitute for RuvABC complex in the process of Holliday junction resolution). Material and methods: Different E. coli recBC sbcBC derivatives were constructed by P1 transduction. Cell growth was monitored by measuring the optical density at 600 nm. Cells were fixed with OsO4 and their chromosomes were visualized by staining with DAPI. Cell morphology was examined by combined phase-contrast and fluorescence microscopy. Results: The ruvA, ruvB and ruvC mutations cause chromosome segregation and cell division defects, and reduce cell viability when introduced into a recBC sbcBC background. All ruv-associated cytological defects could be suppressed by the rus-1 mutation which is known to induce synthesis of an alternative Holliday junction resolvase, the RusA protein. The suppression by rus-1 is the most efficient in the recBC sbcBC strain carrying a ruvA mutation. The suppressive effect of rus-1 depends critically on a functional recG gene. Conclusion: Our results suggest that in recBC sbcBC ruv mutants abortive recombination takes place during exponential growth. The unresolved Holliday junctions accumulated during this process cause cytological defects and affect cell viability.

Escherichia coli; rus-1; chromosome segregation; cell division; Holliday junction

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