ABSTRACT
Chemokines mediate the signaling and migration of T cells, but little is known about the transcriptional events involved therein. Microarray analysis of CXC chemokine ligand (CXCL) 12-treated T cells revealed that Wnt ligands are significantly up-regulated during CXCL12 treatment. Real-time polymerase chain reaction and Western blot analysis confirmed that the expression of noncanonical Wnt pathway members (eg, Wnt5A) was specifically up-regulated during CXCL12 stimulation, whereas beta-catenin and canonical Wnt family members were selectively down-regulated. Wnt5A augmented signaling through the CXCL12-CXCR4 axis via the activation of protein kinase C. Moreover, Wnt5A expression was required for CXCL12-mediated T-cell migration, and rWnt5A sensitized human T cells to CXCL12-induced migration. Furthermore, Wnt5A expression was also required for the sustained expression of CXCR4. These results were further supported in vivo using EL4 thymoma metastasis as a model of T-cell migration. Together, these data demonstrate that Wnt5A is a critical mediator of CXCL12-CXCR4 signaling and migration in human and murine T cells.
Subject(s)
Cell Movement/immunology , Chemokine CXCL12/immunology , Proto-Oncogene Proteins/immunology , Signal Transduction/immunology , T-Lymphocytes/immunology , Wnt Proteins/immunology , Animals , Cell Line, Tumor , Cell Movement/genetics , Chemokine CXCL12/genetics , Chemokine CXCL12/metabolism , Humans , Mice , Mice, Mutant Strains , Proto-Oncogene Proteins/genetics , Proto-Oncogene Proteins/metabolism , Receptors, CXCR4/genetics , Receptors, CXCR4/immunology , Receptors, CXCR4/metabolism , Signal Transduction/genetics , T-Lymphocytes/metabolism , Up-Regulation/genetics , Up-Regulation/immunology , Wnt Proteins/genetics , Wnt Proteins/metabolism , Wnt-5a Protein , beta Catenin/genetics , beta Catenin/immunologyABSTRACT
Lipid rafts play an important role in signal integration and in the cellular activation of a number of cytokine and growth factor receptors. It has recently been demonstrated that flotillin proteins are recruited to lipid raft microdomains upon cellular activation and play a role in neural cell regeneration, receptor signaling and lymphocyte activation. However, little is known about the relevance of the flotillin proteins during T cell responses to chemoattractant stimulation. To this end, cytoplasmic and lipid raft fractions from human T cells were analyzed for flotillin protein redistribution prior to and after CXCL12 stimulation. Flotillin-1, but not flotillin-2, redistributes to lipid rafts upon CXCR4 ligation. Moreover, in CXCL12-treated T cells, flotillin-1 also associates with several raft proteins including LAT, CD48 and CD11a but not Lck. In addition, an increase in CXCR4 association with flotillin-1 in lipid rafts was observed after chemokine treatment. RNAi technology was also utilized to inhibit the expression of flotillin-1, resulting in an inhibition of CXCL12-mediated signaling, function and CXCR4 recruitment into lipid rafts. Together, these data suggest that the increased association of cellular flotillin-1 with lipid raft microdomains during chemokine exposure may play an important role in chemokine receptor signaling and receptor partitioning with lipid rafts.
Subject(s)
Chemokines, CXC/immunology , Membrane Microdomains/immunology , Membrane Proteins/immunology , Receptors, CXCR4/immunology , Signal Transduction/immunology , Blotting, Western , Cell Adhesion/immunology , Chemokine CXCL12 , Chemokines, CXC/metabolism , Chemotaxis, Leukocyte/immunology , Fluorescent Antibody Technique , Humans , Immunoprecipitation , Lymphocyte Activation/immunology , Membrane Microdomains/metabolism , Membrane Proteins/metabolism , Protein Transport/immunology , RNA, Small Interfering , Receptors, CXCR4/metabolism , TransfectionABSTRACT
Ghrelin, a recently described endogenous ligand for the growth hormone secretagogue receptor (GHS-R), is produced by stomach cells and is a potent circulating orexigen, controlling energy expenditure, adiposity, and growth hormone secretion. However, the functional role of ghrelin in regulation of immune responses remains undefined. Here we report that GHS-R and ghrelin are expressed in human T lymphocytes and monocytes, where ghrelin acts via GHS-R to specifically inhibit the expression of proinflammatory anorectic cytokines such as IL-1beta, IL-6, and TNF-alpha. Ghrelin led to a dose-dependent inhibition of leptin-induced cytokine expression, while leptin upregulated GHS-R expression on human T lymphocytes. These data suggest the existence of a reciprocal regulatory network by which ghrelin and leptin control immune cell activation and inflammation. Moreover, ghrelin also exerts potent anti-inflammatory effects and attenuates endotoxin-induced anorexia in a murine endotoxemia model. We believe this to be the first report demonstrating that ghrelin functions as a key signal, coupling the metabolic axis to the immune system, and supporting the potential use of ghrelin and GHS-R agonists in the management of disease-associated cachexia.