Toxin/antitoxin modules are genetic elements located on the plasmids or chromosomes of many bacteria and archaea. These modules are thought to play a role in a variety of cellular processes, including plasmid maintenance and organismal persistence under adverse conditions. Type II toxin/antitoxin modules contain a record for a protein that inhibits cell growth (toxin) and an antitoxin that prevents the action of the toxin. The toxin inhibits essential cellular processes such as DNA replication, translation or cell wall synthesis. Antitoxin opposes the action of the toxin by binding to it and neutralizing its effects.
The module we focused on is higBA2 from Vibrio cholerae. In this module, the antitoxin affects the regulation of its own operon by binding to DNA. The HigB2 toxin acts bacteriostatically by binding to the ribosome and cleaving mRNA. HigA2 antitoxin prevents its action through direct interaction with HigB2, and the binding and subsequent expression of the operon is conditioned by the toxin/antitoxin molar ratio. At high values of the ratio (toxin excess) the antitoxin does not bind to the DNA and the operon is expressed, but at low values (antitoxin excess) expression is disabled.
In this master's thesis, we investigated the influence of the higBA2 toxin/antitoxin system from the bacterium Vibrio cholerae on bacterial growth. We examined how the regulation of transcription can be studied in vivo, and how the presence of the higBA2 module affects bacterial growth in the presence of stress factors or a competitive bacterial strain. The results showed that the strain with higBA2 has a better ability to grow in competitive conditions, although we did not detect significant differences in the growth rate of individual strains. When mapping the transcriptional regulation of higBA2 via the reporter protein, we found that the presence of RBS in front of the reporter protein significantly improves the signal strength.
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