Role of ABCC1 in export of sphingosine-1-phosphate from mast cells

Abstract

Mast cells play a pivotal role in inflammatory and immediate-type allergic reactions by secreting a variety of potent inflammatory mediators, including sphingosine-1-phosphate (S1P). However, it is not known how S1P is released from cells. Here, we report that S1P is exported from mast cells independently of their degranulation and demonstrate that it is mediated by ATP binding cassette (ABC) transporters. Constitutive and antigen-stimulated S1P release was inhibited by MK571, an inhibitor of ABCC1 (MRP1), but not by inhibitors of ABCB1 (MDR-1, P-glycoprotein). Moreover, downregulation of ABCC1 with small interfering RNA, which decreased its cell surface expression, markedly reduced S1P export from both rat RBL-2H3 and human LAD2 mast cells. Transport of S1P by ABCC1 influenced migration of mast cells toward antigen but not degranulation. These findings have important implications for S1P functions in mast cell-mediated immune responses.secretion ͉ sphingolipids ͉ multidrug resistance ͉ allergic responses ͉ sphingosine kinase S phingosine-1-phosphate (S1P) is a pleiotropic lipid mediator that has been implicated in cancer, immunity, and allergy (1-4). S1P is a ligand for a family of five specific G protein-coupled receptors, designated S1P 1-5 (collectively referred to as S1PRs) (5), through which it regulates many different biological responses, including growth, survival, differentiation, cytoskeleton rearrangements, motility, angiogenesis, vascular maturation, lymphocyte trafficking, and mast cell functions (reviewed in refs. 1-3, 5, and 6). Moreover, like its precursors, sphingosine (Sph) and ceramide, S1P may also have intracellular functions (1, 2, 7). S1P is produced in cells by phosphorylation of Sph, the backbone of all sphingolipids, catalyzed by two closely related Sph kinase (SphK) isoenzymes. SphK1 activity is enhanced by numerous external stimuli, including growth factors, ligands for G protein-coupled receptors, and proinflammatory cytokines, and by cross-linking of Ig receptors (reviewed in refs. 1 and 6). To date, only EGF (8) and cross-linking of Ig receptors (4) have been shown to stimulate SphK2.Although S1P secretion has been demonstrated in only a few types of cells (platelets, astroglial cells, and mast cells) activation of SphK1 involves its translocation to the plasma membrane where its substrate Sph resides (9 -11). Increased S1P formation, in turn, activates S1P receptors on the same and͞or neighboring cells in an autocrine or paracrine manner. It has been demonstrated that this type of ''inside-out'' signaling by S1P is critical for migratory responses of fibroblasts and smooth muscle cells toward PDGF (9, 12) and human breast cancer cells toward EGF (8). Similarly, S1P secreted by mast cells is important for migration of mast cells toward antigen (Ag) and might be involved in the movement of mast cells to sites of inf lammation (13). In addition, S1P provokes human airway smooth muscle contraction and may promote inf lammation and airway remodeling in asthma (14). Because S1P's level...

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][4][5] receptors (data not shown). S1P at 1 M was nearly as effective as Ag or ionomycin at stimulating secretion of TNF-␣ and IL-6 ( Figure 1B,C), 2 very important pleiotropic cytokines that elicit a general inflammatory response and have been implicated in IgE-associated chronic allergic diseases.…”

“…Having established that S1P can induce degranulation and cytokine release from LAD2 cells (Figure 1), and is secreted by them upon antigenic stimulation, 5 we next examined the involvement of the SphKs that produce S1P in Ag-induced activation of human MCs by down-regulation of their expression. LAD2 cells express similar levels of SphK1 and SphK2 mRNA and siRNA targeted to SphK1 reduced expression of SphK1 but not SphK2 as determined by QPCR and Western blotting ( Figure 2A, and data not shown).…”

“…4,5 Because there are numerous important functional differences between human and murine MCs (reviewed in Bischoff 12 ), it was important to examine the functions of S1P and the kinases that produce it in human MCs. To this end, we used the LAD2 human mast cell line that is closely related to CD34 ϩ -derived hMCs and expresses functional Fc⑀RI receptors.…”

“…S1P levels are elevated in the bronchoalveolar lavage (BAL) fluid of asthmatics after Ag challenge, suggesting that secretion of S1P by MCs is of relevance in inflammatory responses and asthma. 1 S1P is a ligand for 5 G protein-coupled receptors (GPCRs), designated S1P [1][2][3][4][5] , through which it exerts many of its actions. 2,3 Indeed, S1P is secreted by activated MCs, 4,5 and studies with rodent MCs have shown that this S1P is able to rapidly bind and activate its receptors, S1P 1 and S1P 2 , in an autocrine manner.…”

The role of sphingosine-1-phosphate in the tumor microenvironment and its clinical implications

“…S1P export requires a carrier or transporter, and studies have suggested the involvement of ATP-binding cassette (ABC) transporters in this process from various types of cultured cells. 22 S1P is exported from mast cells via ABCC1 (also known as multidrug-resistant protein 1; MRP1), 32 from astrocytes via ABCA1, 33 from endothelial cells via ABCA1 and ABCC1, 34 and from thyroid carcinoma cells via ABCC1. 35 Furthermore, we demonstrated that ABCC1 and ABCG2 (also known as breast cancer resistance protein; BCRP) are involved in estradiol-mediated transport of S1P and dihydro-S1P out of MCF-7 human breast cancer cells.…”