Dataset Information

OmpR and LeuO positively regulate the Salmonella enterica serovar Typhi ompS2 porin gene.

ABSTRACT: The Salmonella enterica serovar Typhi ompS2 gene codes for a 362-amino-acid outer membrane protein that contains motifs common to the porin superfamily. It is expressed at very low levels compared to the major OmpC and OmpF porins, as observed for S. enterica serovar Typhi OmpS1, Escherichia coli OmpN, and Klebsiella pneumoniae OmpK37 quiescent porins. A region of 316 bp, between nucleotides -413 and -97 upstream of the transcriptional start point, is involved in negative regulation, as its removal resulted in a 10-fold increase in ompS2 expression in an S. enterica serovar Typhi wild-type strain. This enhancement in expression was not observed in isogenic mutant strains, which had specific deletions of the regulatory ompB (ompR envZ) operon. Furthermore, ompS2 expression was substantially reduced in the presence of the OmpR D55A mutant, altered in the major phosphorylation site. Upon random mutagenesis, a mutant where the transposon had inserted into the upstream regulatory region of the gene coding for the LeuO regulator, showed an increased level of ompS2 expression. Augmented expression of ompS2 was also obtained upon addition of cloned leuO to the wild-type strain, but not in an ompR isogenic derivative, consistent with the notion that the transposon insertion had increased the cellular levels of LeuO and with the observed dependence on OmpR. Moreover, LeuO and OmpR bound in close proximity, but independently, to the 5' upstream regulatory region. Thus, the OmpR and LeuO regulators positively regulate ompS2.

SUBMITTER: Fernandez-Mora M

PROVIDER: S-EPMC400630 | biostudies-literature | 2004 May

REPOSITORIES: biostudies-literature

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OmpR and LeuO positively regulate the Salmonella enterica serovar Typhi ompS2 porin gene.

Fernández-Mora Marcos M Puente José Luis JL Calva Edmundo E

Journal of bacteriology 20040501 10

The Salmonella enterica serovar Typhi ompS2 gene codes for a 362-amino-acid outer membrane protein that contains motifs common to the porin superfamily. It is expressed at very low levels compared to the major OmpC and OmpF porins, as observed for S. enterica serovar Typhi OmpS1, Escherichia coli OmpN, and Klebsiella pneumoniae OmpK37 quiescent porins. A region of 316 bp, between nucleotides -413 and -97 upstream of the transcriptional start point, is involved in negative regulation, as its remo ...[more]

PMID: 15126450

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Project description:BackgroundTyphoid fever is an acute systemic infection of humans caused by Salmonella enterica subspecies enterica serovar Typhi (S. Typhi). In chronic carriers, the bacteria survive the harsh environment of the gallbladder by producing biofilm. The phenotype of S. Typhi biofilm cells is significantly different from the free-swimming planktonic cells, and studies have shown that they are associated with antibiotic resistance, immune system evasion, and bacterial persistence. However, the mechanism of this transition and the events leading to biofilm formation are unknown. High throughput sequencing was performed to identify the genes involved in biofilm formation and to postulate the mechanism of action.ResultsPlanktonic S. Typhi cells were cultured using standard nutrient broth whereas biofilm cells were cultured in a stressful environment using high shearing-force and bile to mimic the gallbladder. Sequencing libraries were prepared from S. Typhi planktonic cells and mature biofilm cells using the Illumina HiSeq 2500 platform, and the transcriptome data obtained were processed using Cufflinks bioinformatics suite of programs to investigate differential gene expression between the two phenotypes. A total of 35 up-regulated and 29 down-regulated genes were identified. The identities of the differentially expressed genes were confirmed using NCBI BLAST and their functions were analyzed. The results showed that the genes associated with metabolic processes and biofilm regulations were down-regulated while those associated with the membrane matrix and antibiotic resistance were highly up-regulated.ConclusionsIt is proposed that the biofilm phenotype of S. Typhi allows the bacteria to increase production of the membrane matrix in order to serve as a physical shield and to adhere to surfaces, and enter an energy conservation state in response to the stressful environment. Conversely, the planktonic phenotype allows the bacteria to produce flagella and increase metabolic activity to enable the bacteria to migrate and form new colonies of infection. This data provide a basis for further studies to uncover the mechanism of biofilm formation in S. Typhi and to discover novel genes or pathways associated with the development of the typhoid carrier state.

Project description:A Salmonella enterica serovar Typhi gene that is selectively up-regulated upon bacterial invasion of eukaryotic cells was characterized. The open reading frame encodes a 298-amino-acid hydrophobic polypeptide (30.8 kDa), which is predicted to be an integral membrane protein with nine membrane-spanning domains. The protein is closely related (87 to 94% reliability) to different transport and permease systems. Gene expression under laboratory conditions was relatively weak; however, sevenfold induction was observed in a high-osmolarity medium (300 mM NaCl). The growth pattern in a laboratory medium of a serovar Typhi strain Ty2 derivative containing a 735-bp in-frame deletion in this gene, named gaiA (for gene activated intracellularly), was not affected. In contrast, the mutant was partially impaired in intracellular survival in murine peritoneal macrophages, as well as in human monocyte-derived macrophages. However, in the case of human macrophages, this survival defect was modest and evident only at late infection times (24 h). Despite the distinct intracellular survival kinetics displayed in macrophages of different species, the gaiA null mutant was significantly affected in its potential to trigger apoptosis in both murine and human macrophages. Provision of the gaiA gene in trans resulted in complementation of these phenotypes. Interestingly, the absence of a functional gaiA gene caused a marked attenuation in the mouse mucin model, as shown by the increase (3 orders of magnitude) in the 50% lethal dose of the mutant strain over that of the parental strain Ty2 (P </= 0.05). Altogether, these data indicate that the product encoded by the gaiA gene is required for triggering apoptosis and bacterial survival within murine macrophages, which is consistent with the in vivo results obtained in the mouse mucin model. However, gaiA was not required for initial intracellular survival in human cells, indicating that its role in the natural host might be more complex than is suggested by the studies performed in the murine system.

Dataset Information

OmpR and LeuO positively regulate the Salmonella enterica serovar Typhi ompS2 porin gene.

Publications

OmpR and LeuO positively regulate the Salmonella enterica serovar Typhi ompS2 porin gene.

Similar Datasets

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