Listeria monocytogenes GshF contributes to oxidative stress tolerance via regulation of the phosphoenolpyruvate-carbohydrate phosphotransferase system

Glutathione (GSH) is an essential component of the glutaredoxin (Grx) system, and it is synthesized by the enzyme glutathione synthase GshF in Listeria monocytogenes. GSH plays a crucial role in regulating Listeria virulence by modifying the virulence factors LLO and PrfA. In this study, we investigated the involvement of L. monocytogenes GshF in oxidative tolerance and intracellular infection. Our findings revealed that the deletion of gshF resulted in a significant reduction in bacterial growth in vitro when exposed to diamide and copper ions stress. More importantly, this deletion also impaired the efficiency of invasion and proliferation in macrophages and mice organs. Furthermore, GshF influenced global transcriptional profiles, including carbohydrate and amino acid metabolism, particularly those related to the phosphoenolpyruvate-carbohydrate phosphotransferase system (PTS) genes lmo1997-lmo2004, under oxidative stress conditions. In the wild-type strain, the transcription of lmo1997-lmo2004 was notably downregulated in response to copper ions and diamide stress compared to normal conditions. However, in the absence of gshF, the transcripts of lmo1997-lmo2004 were upregulated in response to these stress conditions. Notably, the deletion of iiB(man) (lmo2002) enhanced oxidative stress tolerance to copper ions, whereas overexpression of iiB(man) reduced this resistance. In conclusion, our study provides the first evidence that L. monocytogenes GshF plays a crucial role in bacterial antioxidation through the regulation of iiB(man). IMPORTANCE Listeria monocytogenes has developed various mechanisms to withstand oxidative stress, including the thioredoxin and glutaredoxin systems. However, the specific role of the glutathione synthase GshF, responsible for synthesizing GSH in L. monocytogenes, in oxidative tolerance remains unclear. This study aimed to elucidate the relationship between GshF and oxidative tolerance in L. monocytogenes by examining the efficiency of invasion and proliferation in macrophages and mice organs, as well as analyzing global transcriptional profiles under oxidative stress conditions. The results revealed that GshF plays a significant role in L. monocytogenes' response to oxidative stress. Notably, GshF acts to suppress the transcription of phosphoenolpyruvate-carbohydrate phosphotransferase system genes lmo1997-lmo2004, among which iiB(man) (lmo2002) was identified as the most critical gene for resisting oxidative stress. These findings enhance our understanding of how L. monocytogenes adapts to its environment and provide valuable insights for investigating the environmental adaptation mechanisms of other pathogenic bacteria.