Gene expression profiles of LPS-stimulated RAW264.7 macrophages overexpressing Tbc1d23 novel innate immune gene
Ontology highlight
ABSTRACT:
ORGANISM(S): Mus musculus
SUBMITTER: Scott Alper
PROVIDER: E-GEOD-30840 | biostudies-arrayexpress |
REPOSITORIES: biostudies-arrayexpress
Publications
Spatiotemporal inhibition of innate immunity signaling by the Tbc1d23 RAB-GAP.
De Arras Lesly L Yang Ivana V IV Lackford Brad B Riches David W H DW Prekeris Rytis R Freedman Jonathan H JH Schwartz David A DA Alper Scott S
Journal of immunology (Baltimore, Md. : 1950) 20120206 6
We previously identified Tbc1d23 as a candidate novel regulator of innate immunity using comparative genomics RNA interference screens in Caenorhabditis elegans and mouse macrophages. Using Tbc1d23 knockout mice and macrophages engineered to overexpress Tbc1d23, we now show that Tbc1d23 is a general inhibitor of innate immunity signaling, strongly inhibiting multiple TLR and dectin-signaling pathways. Tbc1d23 likely acts downstream of the TLR-signaling adaptors MyD88 and Trif and upstream of the ...[more]
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Project description:We previously identified Tbc1d23 as a candidate novel regulator of innate immunity using comparative genomics RNAi screens in C. elegans and mouse macrophages. Using mice with an engineered knockout mutation in Tbc1d23 and macrophages engineered to overexpress Tbc1d23, we now show that Tbc1d23 is a general but not universal inhibitor of the innate immune response, inhibiting multiple Toll-like receptor (TLR) and Dectin signaling pathways but not NOD signaling pathways. Tbc1d23 likely acts downstream of the TLR signaling adaptors MyD88 and Trif and upstream of the transcription factor XBP1. Importantly, like XBP1, Tbc1d23 affects the maintenance but not initiation of inflammatory cytokine production induced by lipopolysaccharide (LPS). The identification of a novel temporal regulator of innate immunity signaling validates the comparative genomics approach for innate immunity gene discovery.
2012-04-04 | GSE30840 | GEO
Project description:Background: Toll-like family of receptors recognizes pathogen-associated molecular patterns (PAMPs) from different organisms. TLR4 is the receptor for bacterial lipopolysaccharide (LPS), dsRNA viral mimic poly(I:C) binds to TLR3, and bacterial CpG DNA signals through TLR9. TLR4 signaling is mediated by adaptor molecules Myd88 and TRIF while TLR3 pathway involves only the TRIF adaptor and TLR9 signals solely through Myd88. Methods: To identify genes other than those in TLR pathways that mediate macrophage response to different PAMPs, RAW264.7 cells were stimulated with LPS, poly(I:C), or CpG DNA, and RNA was profiled on microarrays 6 hrs and 24 hrs post-treatment. Gene expression data were analyzed to determine genes, pathways and transcriptional networks that are in common and unique to each of the three TLR stimuli. Potentially novel candidates revealed by this analysis were tested for their role in innate immunity using RNA interference. Results: Many genes are differentially regulated by LPS and poly(I:C) at both 6 hrs and 24 hrs following treatment, while CpG DNA elicits a much less pronounced transcriptional response. By analyzing gene expression data for networks and pathways, we prioritized differentially expressed genes that are in common to all three PAMPs as well as those shared by LPS and poly(I:C). Knockdown by RNA interference of two genes, Plec1 and TPST1, inhibited production of IL-6 in response to LPS in cultured macrophages. Conclusions: We have identified novel innate immunity genes that may be important in macrophage response to LPS, poly(I:C), and CpG DNA stimuli. Our results provide potential biomarkers and therapeutic targets that should be further investigated in mice and human populations. Keywords: time course For each treatment (Sigma LPS, LIST LPS, and media only), three biological replicates (separate macrophage cultures and RNA isolations) were profiled. Each sample was labeled with Cy3 and Cy5 and co-hybridized with Stratagene Universal Mouse Reference (dye flip techical replicates). Expression at 2 timepoints (6 and 24 hours post-treatment) was assessed.
2010-10-01 | E-GEOD-21548 | biostudies-arrayexpress
Project description:The innate immune system is vital to rapidly responding to pathogens and Toll-like receptors (TLRs) are a critical component of this response. Nanovesicular exosomes play a role in immunity, but to date their exact contribution to the dissemination of the TLR response is unknown. To understand the effect of exosomal cargo released from locally stimulated cells on distal cell expression, we collected exosomes from local ovarian adenocarcinoma (HEY) cells that were either unstimulated (control-exosomes), stimulated with pIC (pIC-exosomes), or lipopolysaccharide (LPS-exosomes) for 48 hours. The three groups of exosomes were added to naïve (distal) cells and the gene expression profiles were compared between local TLR stimulation (for 6 hours) and distal stimulation mediated by exosomes at the 48-hour time point The goal of the study was to delineate the differential effector function of TLR-exosomes based on the innate immune activation state (control-, LPS-, pIC-stimulation) of the cell of origin in vitro. We used microarrays to understand the changes in gene expression in 2 samples of local hey cells either unstimulated(control) or stimulated with poly(I:C) or LPS each. We also profiled 2 samples of distal naïve cells that were exposed to either control, LPS or poly(I:C) exosomes from local cells. Here we show that exosomes from TLR stimulated cells (TLR-exosomes) can largely recapitulate TLR activation in distal cells in vitro. We can abrogate the action-at-a-distance signaling of exosomes by UV irradiation, demonstrating that RNA is crucial for their effector function. This work definitively establishes the differential effector function for TLR-exosomes in communicating the activation state of the cell of origin. HEY cells were either unstimulated (Local control) or stimulated with LPS or poly (I:C) for 6 hours and the RNA from these cells was profiled using a microarray. Exosomes were collected from local cells that were unstimulated or stimulated for 48 hours and added to distal (naive) HEY cells that were exposed to control exosomes, LPS exosomes or pIC exosomes and then the RNA was collected for analysis by microarray. mRNA expression was captured on Affymetrix U133 Plus 2 chips. mRNA microarray data were analyzed using the Expression Console software (Affymetrix) and Bioconductor tools written in the R statistical programming language. Pre-processing of raw signal intensities and normalization was performed using GCRMA (R). Linear modelling of the transformed data was determined by using Limma in R with the Benjamini and Hochberg correction. Differentially expressed probesets were identified using a threshold 5% FDR correction and a fold change ⥠1.4 was applied
2016-05-17 | E-GEOD-81248 | biostudies-arrayexpress
Project description:Molecular Pathways and Transcriptional Networks Involved in the Macrophage Response to LPS, poly(I:C) and CpG DNA Stimulation Background: Toll-like family of receptors recognizes pathogen-associated molecular patterns (PAMPs) from different organisms. TLR4 is the receptor for bacterial lipopolysaccharide (LPS), dsRNA viral mimic poly(I:C) binds to TLR3, and bacterial CpG DNA signals through TLR9. TLR4 signaling is mediated by adaptor molecules Myd88 and TRIF while TLR3 pathway involves only the TRIF adaptor and TLR9 signals solely through Myd88. Methods: To identify genes other than those in TLR pathways that mediate macrophage response to different PAMPs, RAW264.7 cells were stimulated with LPS, poly(I:C), or CpG DNA, and RNA was profiled on microarrays 6 hrs and 24 hrs post-treatment. Gene expression data were analyzed to determine genes, pathways and transcriptional networks that are in common and unique to each of the three TLR stimuli. Potentially novel candidates revealed by this analysis were tested for their role in innate immunity using RNA interference. Results: Many genes are differentially regulated by LPS and poly(I:C) at both 6 hrs and 24 hrs following treatment, while CpG DNA elicits a much less pronounced transcriptional response. By analyzing gene expression data for networks and pathways, we prioritized differentially expressed genes that are in common to all three PAMPs as well as those shared by LPS and poly(I:C). Knockdown by RNA interference of two genes, Plec1 and TPST1, inhibited production of IL-6 in response to LPS in cultured macrophages. Conclusions: We have identified novel innate immunity genes that may be important in macrophage response to LPS, poly(I:C), and CpG DNA stimuli. Our results provide potential biomarkers and therapeutic targets that should be further investigated in mice and human populations. For each treatment (LPS, polyIC, CpG DNA, media only), three biological replicates (separate macrophage cultures and RNA isolations) were profiled. Each sample was labeled with Cy3 and Cy5 and co-hybridized with Stratagene Universal Mouse Reference (dye flip techical replicates). Expression at 2 timepoints (6 and 24 hours post-treatment) was assessed.
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Project description:The inflammatory response initiated by microbial products signaling through Toll-like receptors (TLRs) of the innate immune system is essential for host defense against infection. Because inflammation can be harmful to host tissues, the innate response is highly regulated. Negative regulation of TLR4, the receptor for bacterial lipopolysaccharide (LPS), results in LPS tolerance, defined as hyporesponsiveness to repeated stimulation with LPS. LPS tolerance is thought to protect the host from excessive inflammation by turning off TLR4 signal, which then shuts down TLR-induced genes. However, TLR signaling induces hundreds of genes with very different functions. We reasoned that genes with different functions should have different requirements for regulation. Specifically, genes encoding proinflammatory mediators should be transiently inactivated to limit tissue damage, while genes encoding antimicrobial effectors, which directly target pathogens, should remain inducible in tolerant cells to protect the host from infection. Using an in vitro system of LPS tolerance in macrophages, here we show that TLR-induced genes may indeed be divided into two distinct categories based on their functions and regulatory requirements. Further, we show these distinct groups are regulated by gene-specific, and not signal-specific mechanisms. Experiment Overall Design: We examined gene expression using affymetrix genechips in 3 groups of murine bone-marrow derived macrophages: Naive (untreated), Naive stimulated with LPS, and Tolerant stimulated with LPS. Two biological replicates were performed for each group.
2008-06-15 | E-GEOD-7348 | biostudies-arrayexpress
Project description:In animal production the use of probiotics supplements to promote animal health is increasing. The objective of this study was to assess the impact of probiotics administration on global gene expression in dairy cows. Lactating Holstein Friesian cows (n=10) from the North Carolina Agricultural and Technical State University dairy herd were used for the study. Treatment was a 10 ml oral drench of FASTtrak microbial pack (Probiotics) (Conklin Company, Kansas City, MO) at the recommended dose in water or water only (control). This treatment was carried out for 60 days. Whole blood was collected at the beginning (Day 0) and end of the study (Day 60) for microarray analysis. We employed microarray expression profiling as a discovery platform to identify genes with potential association with probiotics supplementation in cows. Gene expression analysis identified 10,859 differentially expressed genes- 1168 upregulated genes and 9691 downregulated gene. Results for pathway analysis showed significant pathways associated with innate immunity such as the Toll-like receptor (TLR) pathway, inflammation response and Wingless (Wnt) signaling pathway. Real-time PCR was used to validate gene expression of members of the TLR and Wnt signaling pathway. Treatment affected the expression of innate and adaptive immune response, cytokine and Wnt pathway genes. Daily administration of probiotics to dairy cows impacts global gene expression and particularly the expression of innate immune genes in dairy cows. Ten animals were enrolled in the study and an initial blood sample was collected (Day 0). Animals (n=5) received either daily supplementations with FASTtrak microbial pack (Probiotics) (Conklin Company, Kansas City, MO) or water daily (control animals) for 60 days. Blood samples were collected at the end of the study from probiotics-treated and control animals for RNA extraction and microarray analysis. In vitro effect of lipopolysaccharide (LPS) endotoxin treatment was evaluated using blood samples collected from probiotics-treated animals (Day 60 samples) to serve as positive control array. A pooled sample was generated by taking equal concentration of RNA from experimental animals in each group. Pooled samples from each group was hybridized on Agilent one color bovine v2 bovine (v2) 4x44KÂ array slides.
2015-11-21 | E-GEOD-75240 | biostudies-arrayexpress
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2013-03-22 | E-GEOD-45369 | biostudies-arrayexpress
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2016-05-17 | GSE81248 | GEO
Project description:Tight regulation of Toll-like receptor (TLR)-mediated inflammatory responses is important in innate immunity. Here, we show that T cell death-associated gene 51 (TDAG51/PHLDA1) is a novel coactivator of the transcription factor FoxO1, regulating inflammatory mediator production in the lipopolysaccharide (LPS)-induced inflammatory response. TDAG51 induction by LPS stimulation was mediated by the TLR2/4 signaling pathway in bone marrow-derived macrophages (BMMs). LPS-induced inflammatory mediator production was significantly decreased in TDAG51-deficient BMMs. In TDAG51-deficient mice, LPS- or pathogenic Escherichia coli infection-induced lethal shock was reduced by decreasing serum proinflammatory cytokine levels. The recruitment of 14-3-3 to FoxO1 was competitively inhibited by the TDAG51-FoxO1 interaction, leading to blockade of FoxO1 cytoplasmic translocation and thereby strengthening FoxO1 nuclear accumulation. TDAG51/FoxO1 double-deficient BMMs showed significantly reduced inflammatory mediator production compared with TDAG51- or FoxO1-deficient BMMs. TDAG51/FoxO1 double deficiency protected mice against LPS- or pathogenic E. coli infection-induced lethal shock by weakening the systemic inflammatory response. Thus, these results indicate that TDAG51 acts as a coactivator of the transcription factor FoxO1, leading to strengthened FoxO1 activity in the LPS-induced inflammatory response.
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