Unknown,Transcriptomics,Genomics,Proteomics

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Peritoneal and testicular macrophage mrna expression after exposure to UPEC CFT073

ABSTRACT: Ascending bacterial infections of the male genitourinary tract by Escherichia coli are an important cause of male infertility. Thus understanding mechanisms by which immunocompetent cells such as testicular macrophages (TM) respond to infection and how bacterial pathogens manipulate defense pathways is of great relevance. Following infection with uropathogenic E. coli (UPEC) whole genome expression profiling revealed induction of the calcium-dependent nuclear factor of activated T cells (NFAT) signaling pathway in TM as well as in peritoneal macrophages (PM) that were used in comparison. UPEC-dependent NFAT activation was confirmed in cultured TM and in TM using an in vivo infection model. Elevated expression of NFATC2-regulated anti-inflammatory cytokines was found in TM (IL-4, IL-13) and PM (IL-3, IL-4, IL-13). NFATC2 is activated by a rapid influx of calcium caused by the UPEC pore forming toxin alpha-hemolysin. Mutant analysis identified alpha-hemolysin as the main virulent factor responsible for UPEC-dependant suppression of IL-6 and TNF-a cytokine release from PM. Alpha-hemolysin caused differential activation of MAP kinase and AP-1 signaling pathways in TM and PM leading to differential expression profiles of key pro-inflammatory cytokines in PM (IL-1a, IL-1b and IL-6 downregulated) and TM (IL-1b and IL-6 upregulated). In contrast to PM, treatment with lipopolysaccharide did not result in activation of the NFkB pathway in TM as shown by lack of degradation of IkBa and lack of pro-inflammatory cytokine secretion (IL-6, TNF-a). In conclusion, these results propose a mechanism how TM can defend against microbes, while at the same time they are able to maintain the immune privileged status of the testis. 2 Macrophage populations ( peritoneal, testicular) infected with UPEC CFT073; timepoints of mrna harvest at 0 min (control), 30 min and 60 min; of each point 2 biological replicates; of each repliccate 2 technical replicates: summa summarum: 2*((1+1+1)*2*2)=24 samples

ORGANISM(S): Rattus norvegicus

SUBMITTER: Zbigniew Mikulski

PROVIDER: E-GEOD-24780 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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Project description:Cyanobacteria are complex prokaryotes, incorporating a Gram-negative cell wall and internal thylakoid membranes (TM). However, localisation of proteins within cyanobacterial cells is poorly understood. Using subcellular fractionation and quantitative proteomics we report the first subcellular map of the proteome of an entire cyanobacterial (or any other prokaryotic) cell, identifying ~67% of Synechocystis sp. PCC 6803 proteins, ~1000 more than previous studies. 1,712 proteins were assigned to six specific subcellular regions. Proteins involved in energy generation localised to TMs. The majority of transporters, with the exception of the TM localised copper importer CtaA, resided in the plasma membrane (PM). Most metabolic enzymes are soluble although numerous pathways terminated in the TM (notably those involved in peptidoglycan monomer, NADP+, heme, lipid and carotenoid biosynthesis), or the PM (specifically those catalysing lipopolysaccharide, molybdopterin, flavin adenine dinucleotide and menaquinone biosynthesis). Furthermore, we identified the proteins involved in the TM and PM electron transport chains. The majority of ribosomal proteins and enzymes synthesising the storage compound polyhydroxybuyrate formed distinct clusters within the data suggesting that they have similar subcellular distributions to one another as one would expect for proteins operating within multi-component protein structures. Moreover, heterogeneity within membrane and cytoplasmic regions is observed, indicating further cellular complexity. Cyanobacterial TM protein localisation is conserved in Arabidopsis thaliana chloroplasts, suggesting similar proteome organisation in higher photosynthetic organisms. The organisation of a cyanobacterial cell revealed here will significantly aid our understanding of these environmentally and biotechnological important organisms. The successful application of this technique in Synechocystis suggests it could be applied to mapping the proteomes of other cyanobacteria and complex single-celled organisms.

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Peritoneal and testicular macrophage mrna expression after exposure to UPEC CFT073

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