Date of Award

Spring 1987

Degree Type

Thesis - Restricted

Degree Name

Master of Science (MS)

Department

Biology

Abstract

The inhibition of bacterial DNA gyrase by quinolones such as enoxacin leads to a variety of cytopathic effects such as a decrease in DNA synthesis, chromosomal DNA fragmentation, elimination of some types of plasmids, filamentation, and ultimately cell death. This thesis investigates two of these cellular responses: a) the roles of altered chromosomal and plasmid gene expression in the loss of plasmids, and b) the roles of induced sos genes in the survivability of bacteria after gyrase A inhibition. Previous studies have clearly shown that some chrmomosomal genes as well as plasmid sequences are necessary for replication and segregation of plasmid molecules. Quinolone treatment does alter gene expression and therefore, the elimination of plasmids during quinolone treatment may be a result of altered expression of some genes involved in plasmid stability. To investigate the susceptibility of chromosomal and plasmid gene expression to gyrase A inhibition, several different bacterial strains were constructed in which the lac operon was present on different plasmids. The ability to induce B-galactosidase (lacz gene product) activity after enoxacin treatment by plasmid-located lac operons was compared to chromosomal lac operons. All of the bacterial strains showed a decrease in lacz expression, but at sub-lethal concentrations of enoxacin there was a difference in inducibility between plasmid and chromosomal lacz genes, suggesting that gyrase A inhibition exerts a differential effect on gene expression depending on the location of the gene. This drug effect was also shown to be gyrase-mediated since gene expression in a gyra (nalr) strain was not greatly affected. In addition to effects on chromosomal and plasmid topology, quinolones are known to induce the bacterial sos repair system. Since more than 20 different genes are involved in the elaboration of the damage-induced response, lt thus becomes relevant to determine the extent to which gene products of the sos system influences the bacteriocidal effects of quinolone treatment. To investigate the involvement of sos genes, isogenic strains with various mutations in sos genes were compared. My results showed that the sfia gene, which causes filamentation of bacteria after DNA damage, had no effect on bacterial survival after gyrase A inhibition by enoxacin. The sos genes in general did confer a greater cell survival of quinolone damage as seen by the lesser amount of cell death of lexa (Def) mutants, which have non-repressed expression of sos genes. Expression of reca was shown to be most essential to greater cell survival of enoxacin treatment since reca mutants were the most sensitive to enoxacin treatment.

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