Hierarchic regulation of a metabolic pathway: H-NS, CRP, and SsrB control myo -inositol utilization by Salmonella enterica

Abstract

The metabolic island GEI4417/4436 of Salmonella enterica serovar Typhimurium ( S . Typhimurium) enables the degradation of myo -inositol (MI) as the sole carbon and energy source. The island encodes two regulatory factors, the autoregulated repressor IolR that inhibits the transcription of most iol genes in the absence of MI, and the activator ReiD that induces the expression of an iol gene operon that is not controlled by IolR. In this study, we investigated the putative role of global regulators in the control of MI utilization by S . Typhimurium. The histone-like nucleoid structuring protein H-NS is demonstrated here to interact with 16 regions of GEI4417/4436 and to silence the transcriptional activity of the promoters P reiD and P iolE , thus controlling the expression of genes initial for MI degradation. The cAMP-binding regulatory protein (CRP) is shown to bind numerous promoters in GEI4417/4436 and is required for the growth of S . Typhimurium strain 14028 in a minimal medium with MI. The binding kinetics of H-NS and CRP toward promoters of GEI4417/4436 were quantified by surface resonance spectroscopy, showing that H-NS weakly binds to promoters of initial genes and that CRP has a high affinity to promoters of all genes essential for MI utilization. Furthermore, three promoters of the metabolic island were identified here to belong to the virulon of the two-component system SsrA/SsrB of salmonellae as demonstrated by electrophoretic mobility shift assays and the use of chromosomal luciferase reporter fusions. IMPORTANCE The capacity to utilize myo-inositol (MI) as sole carbon and energy source is widespread among bacteria, among them the intestinal pathogen S . Typhimurium. This study elucidates the complex and hierarchical regulation that underlies the utilization of MI by S . Typhimurium under substrate limitation. A total of seven regulatory factors have been identified so far, allowing the pathogen an environment-dependent, efficient, and fine-tuned regulation of a metabolic property that provides growth advantages in different environments.