Unknown,Transcriptomics,Genomics,Proteomics

Dataset Information

Transcription profiling of mouse ANA-1 macrophages infected with Chlamydia pneumoniae

ABSTRACT: This experiment is an additional experiment to GSE6688. Mouse macrophages (ANA-1 cells) were infected in vitro with C. pneumoniae with a M.O.I. of 10. Twenty two genes were significantly upregulated. Examples of the most upregulated genes in mouse macrophages after C. pneumoniae infection are serum amyloid A3 (saa3), a protein that is mainly produced by activated macrophages during tissue injury or inflammation, MIP-2 (cxcl2) and irg1. Expression levels of all genes induced by C. pneumoniae in macrophages in vitro correlated with the results obtained from infected lungs from wild type mice (GSE6688), suggesting that this cell type participates in host defense in vivo against C. pneumoniae. Experiment Overall Design: ANA-1 macrophages were infected with Chlamydia pneumoniae or left untreated. After 8h total RNA was extracted. Two biological replicates were performed resulting in 4 arrays in total

ORGANISM(S): Mus musculus

SUBMITTER: Joerg Mages

PROVIDER: E-GEOD-6690 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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Publications

MyD88-dependent changes in the pulmonary transcriptome after infection with Chlamydia pneumoniae.

Rodríguez Nuria N Mages Jörg J Dietrich Harald H Wantia Nina N Wagner Hermann H Lang Roland R Miethke Thomas T

Physiological genomics 20070320 2

Chlamydia pneumoniae, an intracellular bacterium, causes pneumonia in humans and mice. Toll-like receptors and the key adaptor molecule myeloid differentiation factor-88 (MyD88) play a critical role in inducing immunity against this microorganism and are crucial for survival. To explore the influence of MyD88 on induction of immune responses in vivo on a genome-wide level, wildtype (WT) or MyD88(-/-) mice were infected with C. pneumoniae on anesthesia, and the pulmonary transcriptome was analyze ...[more]

PMID: 17374847

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Project description:Purpose: To further define the enhanced metabolic activity of Chlamydia pneumoniae under hypoxia, a transcriptome screen was performed. Next-genration sequencing (NGS) has revolutioned systems-based analysis of transcriptomic pathways. The goals of this study are to compare the transcriptomic profile of C. pneumoniae, grown within HEp-2 cells under normoxic and hypoxic conditions. Methods: Total RNA of C. pneumoniae infected HEp-2 cells cultured under normoxia (20% O2) or hypoxia (2% O2) was isolated by NucleoSpin RNA kit (Macherey Nagel). Human rRNA was depleted by RiboZero rRNA removal kit (Epicentre) in order to enrich bacterial RNA. The tagged cDNA libraries, in the size range of 250 –400 bp, were pooled and single-read sequencing (read length 50 bp) was performed on Illumina HiSeq 2000 by BGI-Hong Kong. Illumina reads were mapped to the Chlamydia genome (GeneBank, version AE001363.1) by TopHat (version TopHat v1.0.12. Gene expression was determined by the Htseq package and data was normalized using the RPKM conversion. Finally, the differential expression analysis was done using the Bioconductor package NOISeq version 2.6.0. The NOISeq-sim function included in the package allows for differential expression estimates in absence of replication by simulating replicates considering that reads counts follow a multinominal distribution. Results: Ranking of differentially expressed chlamydial genes by NOISeq-sim revealed 153 upregulated and 18 downregulated candidate genes. The expression profile of selected genes was validated using qRT-PCR. Most of the upregulated genes under hypoxia belong to the transcriptional and translational machinery or have unknown function. Moreover, transporters show increased expression under hypoxia such as the ATP/ADP translocase (Cpn0614) responsible for NAD uptake. Within the group of metabolic genes, numerous genes belonging to the energy metabolism and nucleotide metabolism were upregulated under hypoxia. Furthermore, thioredoxin reductase (trxB) belonging to the chlamydial redox system was upregulated. Conclusions: Our study represents the first detailed transcriptomic analysis of C. pneumoniae infected HEp-2 cells under hypoxic conditions, generated by RNA-seq technology. Our results show an increased expression of trxB, a part of the chlamydial redox system, which might reduce reactive oxygen species (ROS) under hypoxia. Interestingly, infection of hypoxic cells indeed resulted in a decreased ROS generation compared to hypoxic non-infected cells. We conclude that enhanced growth of C. pneumoniae is the result of hypoxia-induced mitochondrial dysfunction and the associated metabolic switch. Particularly, mitochondrial hyperpolarization is a central mechanism which promotes C. pneumoniae growth under hypoxic conditions. Further, our data imply that mitochondrial dysfunction could also play a major role for other intracellular pathogens, since mitochondrial dysfunction usually occurs under hypoxia and could thereby influence intracellular growth.

Dataset Information

Transcription profiling of mouse ANA-1 macrophages infected with Chlamydia pneumoniae

Publications

MyD88-dependent changes in the pulmonary transcriptome after infection with Chlamydia pneumoniae.

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