Transcriptomic and network analysis reveal methionine sulfoxide reductases dependent regulation of flagellar motility in Salmonella Typhimurium.

Salmonella enterica serovar Typhimurium (S. Typhimurium), a food-borne pathogen is the major cause of gastroenteritis and systemic infections in humans. Its survival and pathogenicity depend upon coordinated regulation of motility, oxidative stress response, and virulence-associated pathways. By repairing oxidized methionine (Met-SO) residues, the methionine sulfoxide reductases (Msrs) system protects bacterial proteins from oxidative damage and aid in bacterial survival in the host. Flagella enable S. Typhimurium to swim and swarm, facilitating movement through the intestinal mucus thus aid in effective colonization in the host. Mutations in key flagellar genes like fliC or flgE are known to severely impair motility and virulence of this bacterium. S. Typhimurium encodes five msrs. In this study, we examined the global transcriptional changes following deletion of all known msrs in S. Typhimurium (by constructing a Δ5 msr mutant strain). Gene Set Enrichment Analysis (GSEA) revealed significant downregulation of pathways involved in flagellar assembly, chemotaxis, and anaerobic respiration, indicating impaired motility in Δ5 msr mutant strain. Upregulated pathways are enriched in ribosome biogenesis, amino acid metabolism, and two-component regulatory systems. Co-expression network analysis identified key hub genes (msrA, fliC, spaO) forming tightly connected functional clusters. Protein-protein interaction (PPI) analysis of downregulated genes further demonstrated an integrated and highly connected network of flagellar and chemotaxis components. qRT-PCR validation confirmed significant downregulation of multiple Class II and Class III flagellar genes, supporting the transcriptional repression of genes involved in motility of the Δ5 msr mutant strain. Western blot analysis further revealed markedly reduced expression of FliC protein in the Δ5 msr mutant strain as compared to the wild-type strain of S. Typhimurium. Mannitol motility, swimming and swarming motility assays demonstrate defective motility of Δ5 msr mutant strain. Results of current study demonstrates that the deletion of msr genes results in altered transcriptional response in S. Typhimurium which culminates in reduction in transcription of class II and class III flagellar genes and compromised motility of this bacterium.