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Updated: May 8, 2026
Applying an Inducible Expression System to Study Interference of Bacterial Virulence Factors with Intracellular Signaling
Published on: June 25, 2015
Development of a tunable wide-range gene induction system useful for the study of streptococcal toxin-antitoxin
Zhoujie Xie1, Fengxia Qi, Justin Merritt
1Department of Microbiology and Immunology, University of Oklahoma Health Sciences Center, Oklahoma City, Oklahoma, USA.
Abstract:
Despite the plethora of genetic tools that have been developed for use in Streptococcus mutans, the S. mutans genetic system still lacks an effective gene induction system exhibiting low basal expression and strong inducibility. Consequently, we created two hybrid gene induction cassettes referred to as Xyl-S1 and Xyl-S2. Both Xyl-S cassettes are xylose inducible and controlled by the Bacillus megaterium xylose repressor. The Xyl-S cassettes each demonstrated >600-fold-increased reporter activity in the presence of 1.2% (wt/vol) xylose. However, the Xyl-S1 cassette yielded a much higher maximum level of gene expression, whereas the Xyl-S2 cassette exhibited much lower uninduced basal expression. The cassettes also performed similarly in Streptococcus sanguinis and Streptococcus gordonii, which suggests that they are likely to be useful in a variety of streptococci. We demonstrate how both Xyl-S cassettes are particularly useful for the study of toxin-antitoxin (TA) modules using both the previously characterized S. mutans mazEF TA module and a previously uncharacterized HicAB TA module in S. mutans. HicAB TA modules are widely distributed among bacteria and archaea, but little is known about their function. We show that HicA serves as the toxin component of the module, while HicB serves as the antitoxin. Our results suggest that, in contrast to that of typical TA modules, HicA toxicity in S. mutans is modest at best. The implications of these results for HicAB function are discussed.
Insights
Researchers developed novel xylose-inducible gene expression systems (Xyl-S1 and Xyl-S2) for Streptococcus mutans, offering improved control for studying bacterial genetic systems and toxin-antitoxin modules.
Area of Science:
- Microbiology
- Molecular Biology
- Bacterial Genetics
Background:
- Streptococcus mutans lacks effective gene induction systems with low basal expression and high inducibility.
- Existing genetic tools in S. mutans are insufficient for precise gene expression control.
Purpose of the Study:
- To engineer novel, highly inducible gene expression cassettes for S. mutans.
- To characterize the performance of these cassettes in controlling gene expression.
- To apply these systems for the study of toxin-antitoxin modules in streptococci.
Main Methods:
- Construction of two hybrid gene induction cassettes (Xyl-S1 and Xyl-S2) using the Bacillus megaterium xylose repressor.
- Evaluation of reporter gene activity under varying xylose concentrations.
- Application of the cassettes to study the mazEF and HicAB toxin-antitoxin modules in S. mutans.
Main Results:
- Both Xyl-S cassettes demonstrated over 600-fold induction with 1.2% xylose.
- Xyl-S1 achieved higher maximum gene expression, while Xyl-S2 showed lower basal expression.
- The cassettes were effective in Streptococcus sanguinis and Streptococcus gordonii.
- Characterization of the HicAB toxin-antitoxin module, identifying HicA as the toxin and HicB as the antitoxin.
Conclusions:
- The Xyl-S cassettes provide a robust and versatile tool for inducible gene expression in S. mutans and related streptococci.
- These systems facilitate the study of essential genetic elements like toxin-antitoxin modules.
- The HicAB module in S. mutans exhibits modest toxin activity, suggesting unique regulatory functions.
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