Genetic Characterization of the Galactitol Utilization Pathway of Salmonella enterica Serovar Typhimurium

Abstract

Galactitol degradation by salmonellae remains underinvestigated, although this metabolic capability contributes to growth in animals (R. R. Chaudhuri et al., PLoS Genet 9:e1003456, 2013, https://doi.org/10.1371/journal.pgen.1003456). The genes responsible for this metabolic capability are part of a 9.6-kb gene cluster that spans from gatY to gatR (STM3253 to STM3262) and encodes a phosphotransferase system, four enzymes, and a transporter of the major facilitator superfamily. Genome comparison revealed the presence of this genetic determinant in nearly all Salmonella strains. The generation time of Salmonella enterica serovar Typhimurium strain ST4/74 was higher in minimal medium with galactitol than with glucose. Knockout of STM3254 and gatC resulted in a growth-deficient phenotype of S. Typhimurium, with galactitol as the sole carbon source. Partial deletion of gatR strongly reduced the lag phase of growth with galactitol, whereas strains overproducing GatR exhibited a near-zero growth phenotype. Luciferase reporter assays demonstrated strong induction of the gatY and gatZ promoters, which control all genes of this cluster except gatR, in the presence of galactitol but not glucose. Purified GatR bound to these two main gat gene cluster promoters as well as to its own promoter, demonstrating that this autoregulated repressor controls galactitol degradation. Surface plasmon resonance spectroscopy revealed distinct binding properties of GatR toward the three promoters, resulting in a model of differential gat gene expression. The cyclic AMP receptor protein (CRP) bound these promoters with similarly high affinities, and a mutant lacking crp showed severe growth attenuation, demonstrating that galactitol utilization is subject to catabolite repression. Here, we provide the first genetic characterization of galactitol degradation in Salmonella, revealing novel insights into the regulation of this dissimilatory pathway. IMPORTANCEThe knowledge of how pathogens adapt their metabolism to the compartments encountered in hosts is pivotal to our understanding of bacterial infections. Recent research revealed that enteropathogens have adapted specific metabolic pathways that contribute to their virulence properties, for example, by helping to overcome limitations in nutrient availability in the gut due to colonization resistance. The capability of Salmonella enterica serovar Typhimurium to degrade galactitol has already been demonstrated to play a role in vivo, but it has not been investigated so far on the genetic level. To our knowledge, this is the first molecular description of the galactitol degradation pathway of a pathogen.

High binding affinity of repressor IolR avoids costs of untimely induction of myo-inositol utilization by Salmonella Typhimurium

“…3). The molecular excess of protein over DNA for quantitative binding of most promoters tested was in a range between 5 to 21 and thus very low in comparison with other repressors tested under similar conditions1112, pointing to a strong affinity of IolR to its target sequences. The bandshift experiments with the promoters P iolT1 , P iolD1 , P iolR , P iolC1 and P reiD exhibited at least two retarded protein/DNA complexes, suggesting an oligomerization of IolR with increasing protein concentration.…”

Comparative genomics of Salmonella enterica serovar Montevideo reveals lineage-specific gene differences that may influence ecological niche association

“…Additional small horizontally acquired elements involving metabolic genes were identified, including an arsenic resistance island that was found present in all clades of Montevideo [ 46 ]. In addition, a tagatose utilization island tag , which spans STM3251 to STM3256 in Typhimurium [ 47 ], was found but restricted to members of clades I and II ( Fig. 2 ).…”

“…Given the apparent niche association of clade I isolates for cattle, and clade II isolates for poultry, it is possible that these metabolic islands contribute to bacterial colonization in livestock. In support of this suggestion, it has been shown that Typhimurium tag operon mutants were markedly attenuated in their ability to colonize the intestines of cattle, swine and poultry [ 47, 83 ], and in serovar Enteritidis, allantoin utilization genes are upregulated in chicken intestines [ 84, 85 ], while allB deletion mutants demonstrate reduced virulence in poultry [ 51 ]. However, further research is needed to understand how these metabolic capabilities actually impact fitness in colonizing livestock.…”

Salmonella enterica Serovar Typhimurium Strategies for Host Adaptation

“…S. Tm encodes approximately 300 unique sRNAs, and the expression of a subset of sRNAs is highly sensitive to signals encountered within the macrophage environment, such as nutrient starvation, and are controlled by key components of the SPI-2 regulatory system ( Kröger et al, 2012 , 2013 ; Amin et al, 2016 ; Colgan et al, 2016 ). Specifically, the sRNA PinT regulates SPI-2 expression and is critical for host adaptation in vivo ( Chaudhuri et al, 2013 ; Westermann et al, 2016 ). It is possible that post-transcriptional regulation helps S. Tm incorporate environmental signals sensed within the SCV to modulate survival.…”