Metabolomics,Unknown,Transcriptomics,Genomics,Proteomics

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Protein phosphorylations, mediated by pro-inflammatory cytokines IL-32g and IL-17A/F

ABSTRACT: Global kinase activity induced by cytokines IL-32g and IL-17A/F were determined using peptide arrays representing phophorylation targets The objective of this study was to identify common and unique phosphorylation targets of pro-inflammatory cytokines IL-32 and IL-17. These cytokines are associated with the pathogenesis and severity of chronic inflammatory disorders, therefore signaling intermediates of these cytokines could be beneficial as alternate theraputic targets that may likely influence different inflammatory pathways. Phosphorylation of proteins is a critical mechanism in the regulation of cellular processes. This process is meticulously regulated by enzymes known as kinases, which are increasingly being identified as drug targets for a variety of diseases. In this study we interrogated kinase activities (kinome) induced in the presence of the human recombinant cytokines IL-32g and IL-17A/F employing peptide arrays representing 300 peptides, printed in triplicate, representing select phosphorylation events. Human macrophage-like THP-1 cells were used for this comparative kinome analysis. Macrophage-like THP-1 cells were stimulated with either IL-32g (20 ng/ml) or IL-17A/F (20 ng/ml) for 15 min, and the peptide arrays were used to comprehensively analyze protein phosphorylation profiles in the presence of these cytokines. Two independet biological experiments were performed and the kinoem analysis was done in triplicates for each. The phosphorylations of the peptides on the array were quantified in the cytokine-treated cells relative to the un-stimulated control cells. Differentially phosphorylated targets were defined as greater than 1.5 fold increase or decrease (p < 0.06) in phosphorylation compared to un-stimulated control cells.

ORGANISM(S): Homo sapiens

SUBMITTER: Neeloffer Mookherjee

PROVIDER: E-GEOD-28649 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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Project description:IL-17A is a pro-inflammatory cytokine that promotes host defense against infections and contributes to the pathogenesis of chronic inflammatory diseases. Dendritic cells (DC) are antigen-presenting cells responsible for adaptive immune responses. Here, we report that IL-17A induces intense remodeling of lipid metabolism in human monocyte-derived DC, as revealed by microarrays analysis. In particular NR1H3/LXR-a and its target genes were significantly upregulated in response to IL-17A. IL-17A induced accumulation of Oil Red O-positive lipid droplets in DC leading to the generation of lipid-laden DC. A lipidomic study established that all the analyzed lipid species, i.e phospholipids, cholesterol, triglycerides, cholesteryl esters were elevated in IL-17A-treated DC. The increased expression of membrane lipid transporters in IL-17A-treated DC as well as their enhanced ability to uptake the fatty acid Bodipy-FL-C16 suggested that lipid uptake was the main mechanism responsible for lipid accumulation in response to IL-17A. IL-17A-induced lipid laden DC were able to stimulate allogeneic T cell proliferation in vitro as efficiently as untreated DC, indicating that IL-17A-treated DC are potently immunogenic. This study, encompassed in the field of immunometabolism, points out for the first time IL-17A as a modulator of lipid metabolism in DC and provides a rationale to delineate the importance of lipid-laden DC in IL-17A-related inflammatory diseases. We used microarrays analysis to understand the impact of IL-17A on human monocyte-derived human dendritic cells. We found overexpression of many genes involved in lipid metabolism in IL-17A-treated dendritic cells compared to untreated dendritic cells. In particular NR1H3/LXR-a and its target genes were significantly upregulated in response to IL-17A. IL-17A induced accumulation of Oil Red O-positive lipid droplets in DC leading to the generation of lipid-laden DC. A lipidomic study established that all the analyzed lipid species, i.e phospholipids, cholesterol, triglycerides, cholesteryl esters were elevated in IL-17A-treated DC. The increased expression of membrane lipid transporters in IL-17A-treated DC as well as their enhanced ability to uptake the fatty acid Bodipy-FL-C16 suggested that lipid uptake was the main mechanism responsible for lipid accumulation in response to IL-17A. IL-17A-induced lipid laden DC were able to stimulate allogeneic T cell proliferation in vitro as efficiently as untreated DC, indicating that IL-17A-treated DC are potently immunogenic. This study, encompassed in the field of immunometabolism, points out for the first time IL-17A as a modulator of lipid metabolism in DC and provides a rationale to delineate the importance of lipid-laden DC in IL-17A-related inflammatory diseases. RNA was extracted from untreated in vitro-generated DC at day 0 (DC, 4 biological replicates ) or DC cultured for 12 days with IL-17A, in the absence or presence of IFN-g (DC-17 and DC-G17, 5 biological replicates)

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Protein phosphorylations, mediated by pro-inflammatory cytokines IL-32g and IL-17A/F

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