Dataset Information

Intestinal Salmonella typhimurium infection leads to miR-29a induced caveolin 2 regulation.

ABSTRACT:

Background

Salmonella are able to modulate host cell functions facilitating both uptake and resistance to cellular host defence mechanisms. While interactions between bacterial modulators and cellular proteins have been the main focus of Salmonella research, relatively little is known about mammalian gene regulation in response to Salmonella infection. A major class of mammalian gene modulators consists of microRNAs. For our study we examined interactions of microRNAs and regulated mRNAs in mammalian intestinal Salmonella infections using a piglet model.

Methodology/principal findings

After performing microRNA as well as mRNA specific microarray analysis of ileal samples from Salmonella infected as well as control piglets, we integrated expression analysis with target prediction identifying microRNAs that mainly regulate focal adhesion as well as actin cytoskeleton pathways. Particular attention was given to miR-29a, which was involved in most interactions including Caveolin 2. RT-qPCR experiments verified up-regulation of miR-29a after infection while its predicted target Caveolin 2 was significantly down-regulated as examined by transcript and protein detection. Reporter gene assays as well as RNAi experiments confirmed Caveolin 2 to be a miR-29a target. Knock-down of Caveolin 2 in intestinal epithelial cells resulted in retarded proliferation as well as increased bacterial uptake. In addition, our experiments showed that Caveolin 2 regulates the activation of the small Rho GTPase CDC42 but apparently not RAC1 in human intestinal cells.

Conclusions/significance

Our study outlines for the first time important regulation pathways in intestinal Salmonella infection pointing out that focal adhesion and organisation of actin cytoskeleton are regulated by microRNAs. Functional relevance is shown by miR-29a mediated Caveolin 2 regulation, modulating the activation state of CDC42. Further analysis of examined interactions may support the discovery of novel strategies impairing the uptake of intracellular pathogens.

SUBMITTER: Hoeke L

PROVIDER: S-EPMC3691122 | biostudies-literature | 2013

REPOSITORIES: biostudies-literature

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Publications

Intestinal Salmonella typhimurium infection leads to miR-29a induced caveolin 2 regulation.

Hoeke Lena L Sharbati Jutta J Pawar Kamlesh K Keller Andreas A Einspanier Ralf R Sharbati Soroush S

PloS one 20130624 6

<h4>Background</h4>Salmonella are able to modulate host cell functions facilitating both uptake and resistance to cellular host defence mechanisms. While interactions between bacterial modulators and cellular proteins have been the main focus of Salmonella research, relatively little is known about mammalian gene regulation in response to Salmonella infection. A major class of mammalian gene modulators consists of microRNAs. For our study we examined interactions of microRNAs and regulated mRNAs ...[more]

PMID: 23826261

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Project description:BackgroundIntestinal fibrosis is a common and serious complication of Crohn's disease characterized by the accumulation of fibroblasts, deposition of extracellular matrix, and formation of scar tissue. Although many factors including cytokines and proteases contribute to the development of intestinal fibrosis, the initiating mechanisms and the complex interplay between these factors remain unclear.MethodsChronic infection of mice with Salmonella enterica serovar Typhimurium was used to induce intestinal fibrosis. A murine protease-specific CLIP-CHIP microarray analysis was employed to assess regulation of proteases and protease inhibitors. To confirm up- or downregulation during fibrosis, we performed quantitative real-time polymerase chain reaction (PCR) and immunohistochemical stainings in mouse tissue and tissue from patients with inflammatory bowel disease. In vitro infections were used to demonstrate a direct effect of bacterial infection in the regulation of proteases.ResultsMice develop severe and persistent intestinal fibrosis upon chronic infection with Salmonella enterica serovar Typhimurium, mimicking the pathology of human disease. Microarray analyses revealed 56 up- and 40 downregulated proteases and protease inhibitors in fibrotic cecal tissue. Various matrix metalloproteases, serine proteases, cysteine proteases, and protease inhibitors were regulated in the fibrotic tissue, 22 of which were confirmed by quantitative real-time PCR. Proteases demonstrated site-specific staining patterns in intestinal fibrotic tissue from mice and in tissue from human inflammatory bowel disease patients. Finally, we show in vitro that Salmonella infection directly induces protease expression in macrophages and epithelial cells but not in fibroblasts.ConclusionsIn summary, we show that chronic Salmonella infection regulates proteases and protease inhibitors during tissue fibrosis in vivo and in vitro, and therefore this model is well suited to investigating the role of proteases in intestinal fibrosis.

Project description:To cause a systemic infection, Salmonella must respond to many environmental cues during mouse infection and express specific subsets of genes in a temporal and spatial manner, but the regulatory pathways are poorly established. To unravel how micro-environmental signals are processed and integrated into coordinated action, we constructed in-frame non-polar deletions of 83 regulators inferred to play a role in Salmonella enteriditis Typhimurium (STM) virulence and tested them in three virulence assays (intraperitoneal [i.p.], and intragastric [i.g.] infection in BALB/c mice, and persistence in 129X1/SvJ mice). Overall, 35 regulators were identified whose absence attenuated virulence in at least one assay, and of those, 14 regulators were required for systemic mouse infection, the most stringent virulence assay. As a first step towards understanding the interplay between a pathogen and its host from a systems biology standpoint, we focused on these 14 genes. Transcriptional profiles were obtained for deletions of each of these 14 regulators grown under four different environmental conditions. These results, as well as publicly available transcriptional profiles, were analyzed using both network inference and cluster analysis algorithms. The analysis predicts a regulatory network in which all 14 regulators control the same set of genes necessary for Salmonella to cause systemic infection. We tested the regulatory model by expressing a subset of the regulators in trans and monitoring transcription of 7 known virulence factors located within Salmonella pathogenicity island 2 (SPI-2). These experiments validated the regulatory model and showed that the response regulator SsrB and the MarR type regulator, SlyA, are the terminal regulators in a cascade that integrates multiple signals. Furthermore, experiments to demonstrate epistatic relationships showed that SsrB can replace SlyA and, in some cases, SlyA can replace SsrB for expression of SPI-2 encoded virulence factors.

Dataset Information

Intestinal Salmonella typhimurium infection leads to miR-29a induced caveolin 2 regulation.

Background

Methodology/principal findings

Conclusions/significance

Publications

Intestinal Salmonella typhimurium infection leads to miR-29a induced caveolin 2 regulation.

Similar Datasets

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