myo-Inositol transport by Salmonella enterica serovar Typhimurium

Abstract

In Salmonella enterica serovar Typhimurium, the genomic island GEI4417/4436 has recently been identified to be responsible for myo-inositol (MI) utilization. Here, two of the four islandencoded permeases are identified as the MI transporters of this pathogen. In-frame deletion of iolT1 (STM4418) led to a severe growth defect, and deletion of iolT1 (STM4419) to a slight growth defect in the presence of MI. These phenotypes could be complemented by providing the putative transporter genes in trans. Bioluminescence-based reporter assays demonstrated a strong induction of their promoters P iolT1 and P iolT2 in the presence of MI but not of glucose. Deletion of iolR, which encodes the negative regulator of most genes involved in MI degradation, resulted in upregulation of P iolT1 and P iolT2 , indicating that the expression of IolT1 and IolT2 is repressed by IolR. This finding was supported by bandshift assays using purified IolR. Both transporters are located in the membrane when expressed in Escherichia coli. Heterologously expressed IolT1 had its optimal activity at pH 5.5. Together with the strongly reduced MI uptake in the presence of protonophores, this indicates that IolT1 operates as a proton symporter. Usinginositol, a saturable uptake activity of IolT1 with a K m value between 0.49 and 0.79 mM was determined in DH5a expressing IolT1, in S. enterica serovar Typhimurium strain 14028, and in mutant 14028 DiolT2. Phylogenetic analysis of IolT1 identified putative MI transporters in Gram-negative bacteria also able to utilize MI.

Metabolism of myo -Inositol by Legionella pneumophila Promotes Infection of Amoebae and Macrophages

“…Inositol metabolism in B. subtilis, S. enterica, or L. casei is regulated by IolR (37,38,54). In the absence of inositol, IolR binds as a repressor to the operator sequences of inositol transporter (iolT) and metabolism genes, and if inositol is available, a catabolic in- termediate of inositol metabolism acts as a derepressor of IolR (37,63).…”

“…This uptake defect was complemented by expressing lpg1653 from a plasmid. Thus, lpg1653 was identified as the inositol transporter gene iolT, and the corresponding protein was termed IolT, in reference to the described inositol transporters from B. subtilis and S. enterica (37,54). Analogous inositol uptake studies were performed with an L. pneumophila ⌬iolG mutant strain.…”

Bistability in myo -Inositol Utilization by Salmonella enterica Serovar Typhimurium

“…In environments lacking MI, most iol gene promoters are repressed by IolR (21). When MI is added, only a few molecules will enter the cell due to the repression of iolT1 encoding the major facilitator of MI (22). MI is degraded to intermediates such as DKP, which is now present at suboptimal levels but induces bistability by binding to IolR.…”

“…In rich medium, the negative regulator IolR represses all but one promoter of the iol divergon, including its own (21). A protein belonging to the major facilitator superfamily, IolT1, has recently been identified as the predominant MI transporter of S. Typhimurium 14028, and IolR was revealed to also inhibit the transcription of iolT1 (22).…”

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

“…A key player in MI utilization is the negative autoregulator IolR that represses in rich medium all but one promoter of the iol genes involved in MI degradation, including its own promoter2 and that of the iolT1 gene encoding the predominant MI transporter in S. Typhimurium 140286. An intermediate of MI degradation, 2-deoxy-5-keto-D-gluconic acid 6-phosphate (DKGP), has been shown to antagonize IolR binding, thus inducing the expression of iol genes78.…”