IADR Abstract Archives
Characterization of a Putative Oxidative Stress Gene in Streptococcus Mutans
Objectives: The dental pathogen Streptococcus mutans faces multiple environmental stresses in oral biofilms, which includes peroxide (H2O2) stress generated by the metabolism of oral commensal bacteria. PerR is a conserved DNA-binding transcriptional repressor that becomes inactive during peroxide stress due to a protein conformational change. Among the few S. mutans genes containing highly conserved PerR-binding motifs, all have been characterized or are annotated to have roles in oxidative stress or metal homeostasis, with the exception of smu635 that encodes for an open reading frame with unknown function.
Methods: A smu635 deletion (△smu635) strain was created by replacement of the coding region with an antibiotic resistance cassette in both the UA159 wild-type and △perR background strains. Growth of the strains under acid or oxidative stress conditions was monitored using an automated growth reader. The association of Smu635 with metal homeostasis was explored by growing cells in the presence of the iron-dependent antibiotic streptonigrin or in sub-inhibitory concentrations of trace metals.
Results: When compared to UA159, the △smu635 strain showed a severe growth defect, even in the absence of stress. This growth defect was considerably less severe if the perR gene was also inactivated such that further characterizations of smu635 were conducted in the △perR background. When compared to △perR, the △perR△smu635 double mutant displayed heightened sensitivity to H2O2 and diamide stresses but not to low pH or metal intoxication. However, the △perR△smu635 strain was slightly more sensitive to streptonigrin than the UA159 and △perR strains suggesting that Smu635 may play a role in iron homeostasis.
Conclusions: The PerR-regulated gene smu635 is a previously uncharacterized oxidative stress gene. Studies to uncover the mechanism by which Smu635 mediates oxidative stress tolerance and, possibly, metal homeostasis are currently underway.
Methods: A smu635 deletion (△smu635) strain was created by replacement of the coding region with an antibiotic resistance cassette in both the UA159 wild-type and △perR background strains. Growth of the strains under acid or oxidative stress conditions was monitored using an automated growth reader. The association of Smu635 with metal homeostasis was explored by growing cells in the presence of the iron-dependent antibiotic streptonigrin or in sub-inhibitory concentrations of trace metals.
Results: When compared to UA159, the △smu635 strain showed a severe growth defect, even in the absence of stress. This growth defect was considerably less severe if the perR gene was also inactivated such that further characterizations of smu635 were conducted in the △perR background. When compared to △perR, the △perR△smu635 double mutant displayed heightened sensitivity to H2O2 and diamide stresses but not to low pH or metal intoxication. However, the △perR△smu635 strain was slightly more sensitive to streptonigrin than the UA159 and △perR strains suggesting that Smu635 may play a role in iron homeostasis.
Conclusions: The PerR-regulated gene smu635 is a previously uncharacterized oxidative stress gene. Studies to uncover the mechanism by which Smu635 mediates oxidative stress tolerance and, possibly, metal homeostasis are currently underway.