| SH2-containing inositol 5′-phosphatase (SHIP) is a 145kDa protein that becomes both tyrosine phosphorylated and associated with the adapter protein Shc following the stimulation of hemopoietic cells with a variety of extracellular stimuli. SHIP typically acts as a negative regulator of hemopoietic cell activation, at least in part, by hydrolyzing the phosphatidylinositol 3′-kinase (PI3K) generated second messenger phosphatidylinositol 3,4,5-thsphosphate (PIP3). To gain further insight into SHIP's role in mast cell development and activation, we generated bone marrow-derived mast cells (BMMCs) from SHIP+/+ and −/− mice. We found that mature BMMCs from these mice exhibit comparable receptor expression profiles, total granularity and granular content, despite the faster rate of development observed in the absence of SHIP. Following stimulation of these cells, we found that IgE (Immunglobulin E) + antigen (Ag)-induced degranulation, arachidonic acid (AA) metabolism, and proinflammatory cytokine production are substantially higher in SHIP−/− than +/+ BMMCs. Focusing on cytokine production, we demonstrate herein that SHIP negatively regulates interleukin (IL)-6 production by inhibiting nuclear factor κB (NFκB) activity. Using various pathway inhibitors we determined that the PI-3K/Protein Kinase (PK)B and PKC pathways, which are elevated in IgE+Ag-induced SHIP−/− cells, elevate IL-6 mRNA synthesis, at least in part, by enhancing the phosphorylation of IκB and NFκB DNA binding. Conversely, the Erk and p38 pathways, which are also elevated in SHIP−/− cells, enhance IL-6 mRNA synthesis by increasing the transactivation potential of NFκB. Taken together, our results are consistent with a model in which SHIP negatively regulates NFκB activity and IL-6 synthesis by reducing IgE+Ag-induced PIPS levels and thus PKB, PKC, Erk and p38 activation.; Although IgE binding to mast cells is thought to be a passive pre-sensitization step in the current mast cell paradigm, we observed that SHIP−/− BMMCs degranulate in response to IgE alone, unlike their wildtype counterparts. We explored this phenomenon further and found that monomeric IgE (mIgE), in the absence of Ag, stimulates multiple phosphorylation events in normal BMMCs. While mIgE does not induce degranulation or leukotriene synthesis, it leads to a more potent production of cytokines than IgE+Ag. (Abstract shortened by UMI.)... |
