title: Selective regulation by delta-PKC and PI 3-kinase in the assembly of the antiapoptotic TNFR-1 signaling complex in neutrophils.
abstract: TNF is implicated in the attenuation of neutrophil constitutive apoptosis during sepsis. Antiapoptotic signaling is mediated principally through the TNF receptor-1 (TNFR-1). In adherent neutrophils, when beta-integrin signaling is activated, TNF phosphorylates TNFR-1 and activates prosurvival and antiapoptotic signaling. Previously, we identified the delta-PKC isotype and phosphatidylinositol (PI) 3-kinase as critical regulators of TNF signaling in adherent neutrophils. Both kinases associate with TNFR-1 in response to TNF and are required for TNFR-1 serine phosphorylation, NF-kappaB activation, and inhibition of apoptosis. The purpose of this study was to examine the role of delta-PKC and PI 3-kinase in the assembly of TNFR-1 signaling complex that regulates NF-kappaB activation and antiapoptotic signaling. Coimmunoprecipitation studies established that PI 3-kinase, delta-PKC, and TNFR-1 formed a signal complex in response to TNF. delta-PKC recruitment required both delta-PKC and PI 3-kinase activity, whereas PI 3-kinase recruitment was delta-PKC independent, suggesting that PI 3-kinase acts upstream of delta-PKC. An important regulatory step in control of antiapoptotic signaling is the assembly of the TNFR-1-TNFR-1-associated death domain protein (TRADD)-TNFR-associated factor 2 (TRAF2)-receptor interacting protein (RIP) complex that controls NF-kappaB activation. Inhibition of either delta-PKC or PI 3-kinase decreased TNF-mediated recruitment of RIP and TRAF2 to TNFR-1. In contrast, TRADD recruitment was enhanced. Thus delta-PKC and PI 3-kinase are positive regulators of TNF-mediated association of TRAF2 and RIP with TNFR-1. Conversely, these kinases are negative regulators of TRADD association. These results suggest that delta-PKC and PI 3-kinase regulate TNF antiapoptotic signaling at the level of the TNFR-1 through control of assembly of a TNFR-1-TRADD-RIP-TRAF2 complex.
authors.affiliation: Immunology Section, Rm. 1212H Abramson Bldg., Children's Hospital of Philadelphia, 3615 Civic Center Boulevard, Philadelphia, PA 19104, USA. kilpatrick@email.chop.edu
authors.forename: Laurie E
authors.initials: LE
authors.lastname: Kilpatrick
authors.forename: Shuang
authors.initials: S
authors.lastname: Sun
authors.forename: Helen M
authors.initials: HM
authors.lastname: Korchak
mesh_terms: D015703:Antigens, CD / Q000378:metabolism*; D017209:Apoptosis / Q000502:physiology*; D015153:Blotting, Western; D002448:Cell Adhesion / Q000502:physiology; D002478:Cells, Cultured; D004591:Electrophoresis, Polyacrylamide Gel; D006801:Humans; D016328:NF-kappa B / Q000378:metabolism; D009504:Neutrophils / Q000276:immunology / Q000378:metabolism*; D019869:Phosphatidylinositol 3-Kinases / Q000378:metabolism*; D011233:Precipitin Tests; D011493:Protein Kinase C / Q000378:metabolism*; D011506:Proteins / Q000378:metabolism; D053422:Receptor-Interacting Protein Serine-Threonine Kinases; D018124:Receptors, Tumor Necrosis Factor / Q000378:metabolism*; D047888:Receptors, Tumor Necrosis Factor, Type I; D015398:Signal Transduction / Q000502:physiology*; D047990:TNF Receptor-Associated Factor 1; D047992:TNF Receptor-Associated Factor 2
chemical_list: D015703:Antigens, CD; D016328:NF-kappa B; D011506:Proteins; D018124:Receptors, Tumor Necrosis Factor; D047888:Receptors, Tumor Necrosis Factor, Type I; D047990:TNF Receptor-Associated Factor 1; D047992:TNF Receptor-Associated Factor 2; D019869:Phosphatidylinositol 3-Kinases; C506249:RIPK1 protein, human; D053422:Receptor-Interacting Protein Serine-Threonine Kinases; D011493:Protein Kinase C