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Tetragenococcus halophilus is a moderately halophilic lactic acid bacterium widely used in high-salt food fermentation because of its coping ability under various stress conditions. Bacterial toxin-antitoxin (TA) modules are widely distributed and play important roles in stress response, but those specific for genus Tetragenococcus have never been explored. Here, a bona fide TA module named DinJ1-YafQ1tha was characterized in T. halophilus. The toxin protein YafQ1tha acts as a ribonuclease, and its overexpression severely inhibits Escherichia coli growth. These toxic effects can be eliminated by introducing DinJ1tha, indicating that YafQ1tha activity is blocked by the formed DinJ1-YafQ1tha complex. In vivo and in vitro assays showed that DinJ1tha alone or DinJ1-YafQ1tha complex can repress the transcription of dinJ1-yafQ1tha operon by binding directly to the promoter sequence. In addition, dinJ1-yafQ1tha is involved in plasmid maintenance and stress response, and its transcriptional level is regulated by various stresses. These findings reveal the possible roles of DinJ1-YafQ1tha system in the stress adaptation processes of T. halophilus during fermentation. A single antitoxin DinJ2tha without a cognate toxin protein was also found. Its sequence shows low similarity to that of DinJ1tha, indicating that this antitoxin may have evolved from a different ancestor. Moreover, DinJ2tha can cross-interact with noncognate toxin YafQ1tha and cross-regulate with dinJ1-yafQ1tha operon. In summary, DinJ-YafQtha characterization may be helpful in investigating the key roles of TA systems in T. halophilus and serves as a foundation for further research. KEY POINTS: • dinJ1-yafQ1tha is the first functional TA module characterized in T. halophilus and upregulated significantly upon osmotic and acidic stress. • DinJ2tha can exhibit physical and transcriptional interplay with DinJ1-YafQ1tha. • dinJ2tha may be acquired from bacteria in distant affiliation and inserted into the T. halophilus genome through horizontal gene transfer.


Xiaotong Luo, Jieting Lin, Junwei Yan, Xiaoxian Kuang, Hantao Su, Weifeng Lin, Lixin Luo. Characterization of DinJ-YafQ toxin-antitoxin module in Tetragenococcus halophilus: activity, interplay, and evolution. Applied microbiology and biotechnology. 2021 May;105(9):3659-3672

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PMID: 33877415

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