Transactivation of Sphingosine-1–Phosphate Receptors by FcεRI Triggering Is Required for Normal Mast Cell Degranulation and Chemotaxis

Abstract

Mast cells secrete various substances that initiate and perpetuate allergic responses. Cross-linking of the high-affinity receptor for IgE (FcɛRI) in RBL-2H3 and bone marrow–derived mast cells activates sphingosine kinase (SphK), which leads to generation and secretion of the potent sphingolipid mediator, sphingosine-1–phosphate (S1P). In turn, S1P activates its receptors S1P1 and S1P2 that are present in mast cells. Moreover, inhibition of SphK blocks FcɛRI-mediated internalization of these receptors and markedly reduces degranulation and chemotaxis. Although transactivation of S1P1 and Gi signaling are important for cytoskeletal rearrangements and migration of mast cells toward antigen, they are dispensable for FcɛRI-triggered degranulation. However, S1P2, whose expression is up-regulated by FcɛRI cross-linking, was required for degranulation and inhibited migration toward antigen. Together, our results suggest that activation of SphKs and consequently S1PRs by FcɛRI triggering plays a crucial role in mast cell functions and might be involved in the movement of mast cells to sites of inflammation.

Distinct roles of sphingosine kinases 1 and 2 in human mast-cell functions

“…4 In agreement, FTY720-phosphate, an agonist of all S1P receptors except S1P 2 , did not induce degranulation. 19 Moreover, quantitative real-time PCR (QPCR) revealed that like rodent MCs, LAD2 cells as well as cord blood-derived human MCs and human skin MCs express similar levels of S1P 1 and S1P 2 , but do not express S1P 3-5 receptors (data not shown).…”

“…Secretion of granules was determined by measuring the release of the granule marker ␤-hexosaminidase with a colorimetric assay in which the production of p-nitrophenol from p-nitrophenyl-N-acetyl-␤-D-glucosaminide is measured. 4 Values are expressed as the percentage of total cellular ␤-hexosaminidase released into the medium. Chemotaxis was measured in transwells (Costar, Cambridge, MA) with 8-m pore size.…”

“…4,6,7 In human BMMCs and murine MCs, SphK1 translocates to the plasma membrane within minutes of Fc⑀RI clustering. 4,6 SphK1 interacts with the protein tyrosine kinases, Src kinase members Lyn and Fyn (Fc⑀RI proximal kinases that initiate the signaling events following cross-linking of this receptor), but not Src or other tyrosine kinases, such as Syk. Following interaction with Lyn, SphK1 is recruited to membrane rafts where conformational changes may occur in both kinases that favor their respective activities.…”

“…Moreover, secreted S1P regulates mast-cell degranulation and cytokine and chemokine production in an autocrine and/or paracrine manner through activation of their S1P receptors. 4 In this study, we examined the roles of S1P formed by SphK1 and SphK2 in regulation of the functions of hMCs. We discovered that S1P is a much more potent regulator of human mast cells than of rodent mast cells, and is capable of inducing robust degranulation and cytokine and chemokine production at physiological concentrations.…”

“…4,6-8 Down-regulation of SphK1 but not SphK2 in these MCs inhibits calcium mobilization, degranulation, and migration induced by Ag. 4,6,7 In human BMMCs and murine MCs, SphK1 translocates to the plasma membrane within minutes of Fc⑀RI clustering. 4,6 SphK1 interacts with the protein tyrosine kinases, Src kinase members Lyn and Fyn (Fc⑀RI proximal kinases that initiate the signaling events following cross-linking of this receptor), but not Src or other tyrosine kinases, such as Syk.…”