Abstract
BackgroundThe S. mutans LrgA/B holin-like proteins have been shown to affect biofilm formation and oxidative stress tolerance, and are regulated by oxygenation, glucose levels, and by the LytST two-component system. In this study, we sought to determine if LytST was involved in regulating lrgAB expression in response to glucose and oxygenation in S. mutans.ResultsReal-time PCR revealed that growth phase-dependent regulation of lrgAB expression in response to glucose metabolism is mediated by LytST under low-oxygen conditions. However, the effect of LytST on lrgAB expression was less pronounced when cells were grown with aeration. RNA expression profiles in the wild-type and lytS mutant strains were compared using microarrays in early exponential and late exponential phase cells. The expression of 40 and 136 genes in early-exponential and late exponential phase, respectively, was altered in the lytS mutant. Although expression of comYB, encoding a DNA binding-uptake protein, was substantially increased in the lytS mutant, this did not translate to an effect on competence. However, a lrgA mutant displayed a substantial decrease in transformation efficiency, suggestive of a previously-unknown link between LrgA and S. mutans competence development. Finally, increased expression of genes encoding antioxidant and DNA recombination/repair enzymes was observed in the lytS mutant, suggesting that the mutant may be subjected to increased oxidative stress during normal growth. Although the intracellular levels of reaction oxygen species (ROS) appeared similar between wild-type and lytS mutant strains after overnight growth, challenge of these strains with hydrogen peroxide (H2O2) resulted in increased intracellular ROS in the lytS mutant.ConclusionsOverall, these results: (1) Reinforce the importance of LytST in governing lrgAB expression in response to glucose and oxygen, (2) Define a new role for LytST in global gene regulation and resistance to H2O2, and (3) Uncover a potential link between LrgAB and competence development in S. mutans.
Highlights
The S. mutans LrgA/B holin-like proteins have been shown to affect biofilm formation and oxidative stress tolerance, and are regulated by oxygenation, glucose levels, and by the LytST two-component system
It was shown that oxygen and glucose metabolism have a pronounced effect on lrg and cid expression in S. mutans, but the specific role of LytS, if any, in this regulation was not addressed [11,37]
Mutation of lytS resulted in a severe loss of stationary phase lrgAB induction in cells grown in 11 mM glucose, lrgAB expression was not completely abolished
Summary
The S. mutans LrgA/B holin-like proteins have been shown to affect biofilm formation and oxidative stress tolerance, and are regulated by oxygenation, glucose levels, and by the LytST two-component system. An emerging trend in the study of bacterial biofilms has been a focus on the contribution of bacterial cell death and autolysis to biofilm adherence, maturation, and dispersal It has been demonstrated in a wide variety of bacteria that death and lysis of a subpopulation of cells can facilitate biofilm formation due to the release of DNA into the extracellular environment (eDNA) [17,18,19,20,21,22]. Characterization of S. aureus cid and lrg mutants has revealed that these operons have opposing effects on cell death and murein hydrolase activity [27,29] These observations, combined with the fact that LrgA and CidA share structural features with the bacteriophage lambda family of holin proteins [29], have led to the hypothesis that CidA and LrgA control cell death and lysis in a manner analogous to effector and inhibitor holins, respectively [26,30]. The molecular details of how Cid and Lrg function to control cell death and lysis have not yet been completely elucidated, the fact that cid and lrg homologues have been identified in a wide variety of bacterial and archeal genomes supports a fundamental and conserved role for this system in cell physiology [30,36]
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