Dataset Information

Analysis of A to I editing of miRNA in macrophages exposed to Salmonella.

ABSTRACT: The main mediator of the lipopolysaccharide (LPS) response in macrophages is activation of Toll-like receptor 4 (TLR4). This generates interferon-beta (INF-beta) production that stimulates increased expression of the RNA editing enzyme ADAR1. To determine if there is an increase in RNA editing in mature miRNA in response to TLR4 activation upon Salmonella infection of macrophages we analyzed small RNA deep sequencing data. Interestingly, we found that direct infection of macrophage cell lines with Salmonella does not result in an increase of edited mature miRNA. Thus, despite elevated levels of ADAR1 during TLR4 activation of macrophages mediated by Salmonella infection, ADAR1 does not result in redirection of miRNA. The most common consequence of ADAR activity on miRNA is a reduction in the mature miRNA level due to interference with miRNA processing of pri-miRNA. However, we found very few miRNAs with reductions in level, and no significant difference between miRNAs previously reported to be edited and those reported to be not edited. In particular, we did not see significant decrease in mir-22 and mir-142, nor editing of pri-mir-22 or pri-mir-142 in infected RAW macrophages. Thus, ADAR1 has very little, if any, effect on the miRNA machinery following TL4 activation by Salmonella infection.

SUBMITTER: Heale BS

PROVIDER: S-EPMC3073258 | biostudies-literature | 2010 Sep-Oct

REPOSITORIES: biostudies-literature

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Analysis of A to I editing of miRNA in macrophages exposed to Salmonella.

Heale Bret S E BS Eulalio Ana A Schulte Leon L Vogel Jörg J O'Connell Mary A MA

RNA biology 20100901 5

The main mediator of the lipopolysaccharide (LPS) response in macrophages is activation of Toll-like receptor 4 (TLR4). This generates interferon-beta (INF-beta) production that stimulates increased expression of the RNA editing enzyme ADAR1. To determine if there is an increase in RNA editing in mature miRNA in response to TLR4 activation upon Salmonella infection of macrophages we analyzed small RNA deep sequencing data. Interestingly, we found that direct infection of macrophage cell lines wi ...[more]

PMID: 21037424

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Project description:BackgroundMacrophages play essential roles in both innate and adaptive immune responses. Bacteria require endotoxin, a complex lipopolysaccharide, for outer membrane permeability and the host interprets endotoxin as a signal to initiate an innate immune response. The focus of this study is kinetic and global transcriptional analysis of the chicken macrophage response to in vitro stimulation with endotoxin from Salmonella typhimurium-798.ResultsThe 38535-probeset Affymetrix GeneChip Chicken Genome array was used to profile transcriptional response to endotoxin 1, 2, 4, and 8 hours post stimulation (hps). Using a maximum FDR (False Discovery Rate) of 0.05 to declare genes as differentially expressed (DE), we found 13, 33, 1761 and 61 DE genes between endotoxin-stimulated versus non-stimulated cells at 1, 2, 4 and 8 hps, respectively. QPCR demonstrated that endotoxin exposure significantly affected the mRNA expression of IL1B, IL6, IL8, and TLR15, but not IL10 and IFNG in HD 11 cells. Ingenuity Pathway Analysis showed that 10% of the total DE genes were involved in inflammatory response. Three, 9.7, 96.8, and 11.8% of the total DE inflammatory response genes were significantly differentially expressed with endotoxin stimulation at 1, 2, 4 and 8 hps, respectively. The NFKBIA, IL1B, IL8 and CCL4 genes were consistently induced at all times after endotoxin treatment. NLRC5 (CARD domain containing, NOD-like receptor family, RCJMB04_18i2), an intracellular receptor, was induced in HD11 cells treated with endotoxin.ConclusionsAs above using an in vitro model of chicken response to endotoxin, our data revealed the kinetics of gene networks involved in host response to endotoxin and extend the known complexity of networks in chicken immune response to Gram-negative bacteria such as Salmonella. The induction of NFKBIA, IL1B, IL8, CCL4 genes is a consistent signature of host response to endotoxin over time. We make the first report of induction of a NOD-like receptor family member in response to Salmonella endotoxin in chicken macrophages.

Project description:UnlabelledHemophagocytes are cells of the monocyte lineage that have engulfed erythrocytes and leukocytes. Hemophagocytes frequently accumulate in patients with severe acute bacterial infections, such as those caused by Salmonella enterica, Brucella abortus, and Mycobacterium tuberculosis. The relationship between hemophagocytosis and infection is not well understood. In the murine liver, S. enterica serovar Typhimurium resides within hemophagocytic macrophages containing leukocytes. Here we show that S. Typhimurium also resides within hemophagocytes containing erythrocytes. In cell culture, S. Typhimurium benefits from residence within hemophagocytes by accessing iron, but why macrophages hemophagocytose is unknown. We show that treatment of macrophages with a cocktail of the proinflammatory cytokine interferon gamma (IFN-γ) and lipopolysaccharide (LPS) stimulates engulfment of nonsenescent erythrocytes. Exposure of resting or IFN-γ-treated macrophages to live, but not to heat-killed, S. Typhimurium cells also stimulates erythrocyte engulfment. Single-cell analyses show that S. Typhimurium-infected macrophages are more likely to erythrophagocytose and that infected macrophages engulf more erythrocytes than uninfected macrophages within the same culture well. In addition, macrophages containing erythrocytes harbor more bacteria. However, S. Typhimurium does not promote macrophage engulfment of polystyrene beads, suggesting a role for a ligand on the target cell. Finally, neither of the two S. Typhimurium type 3 secretion systems, T3SS1 or T3SS2, is fully required for hemophagocytosis. These results indicate that infection of macrophages with live S. Typhimurium cells stimulates hemophagocytosis.ImportanceMacrophages are white blood cells (leukocytes) that engulf and destroy pathogens. Hemophagocytes, a subset of macrophages, are characteristic of severe acute infection in patients with, for instance, typhoid fever, brucellosis, tuberculosis, and leishmaniasis. Each of these diseases has the potential to become chronic. Hemophagocytes (blood-eating cells) engulf and degrade red and white blood cells for unknown reasons. The bacterial pathogen Salmonella acquires the essential nutrient iron from murine hemophagocytes. We report that Salmonella stimulates macrophages to engulf blood cells, indicating that cells of this bacterium actively promote the formation of a specialized cellular niche in which they can acquire nutrients, evade killing by the host immune system, and potentially transition to chronic infection.

Dataset Information

Analysis of A to I editing of miRNA in macrophages exposed to Salmonella.

Publications

Analysis of A to I editing of miRNA in macrophages exposed to Salmonella.

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