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RGS13 Controls G Protein-Coupled Receptor-Evoked Responses of Human Mast Cells¹

Geetanjali Bansal^2,*, Jeffrey A. DiVietro^*, Hye Sun Kuehn, Sudhir Rao^3,, Karl H. Nocka^4,, Alasdair M. Gilfillan^4, and Kirk M. Druey^5,*

^* Molecular Signal Transduction Section and Mast Cell Biology Section, Laboratory of Allergic Diseases, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Bethesda, MD 20892; and UCB Pharma, Inc., Cambridge, MA 02139

IgE-mediated mast cell degranulation and release of vasoactive mediators induced by allergens elicits allergic responses. Although G protein-coupled receptor (GPCR)-induced signals may amplify IgE-dependent degranulation, how GPCR signaling in mast cells is regulated remains incompletely defined. We investigated the role of regulator of G protein signaling (RGS) proteins in the modulation of these pathways in human mast cells. Several RGS proteins were expressed in mast cells including RGS13, which we previously showed inhibited IgE-mediated mast cell degranulation and anaphylaxis in mice. To characterize how RGS13 affects GPCR-mediated functions of human mast cells, we analyzed human mast cell lines (HMC-1 and LAD2) depleted of RGS13 by specific small interfering RNA or short hairpin RNA and HMC-1 cells overexpressing RGS13. Transient RGS13 knockdown in LAD2 cells lead to increased degranulation to sphingosine-1-phosphate but not to IgE-Ag or C3a. Relative to control cells, HMC-1 cells stably expressing RGS13-targeted short hairpin RNA had greater Ca²⁺ mobilization in response to several natural GPCR ligands such as adenosine, C5a, sphingosine-1-phosphate, and CXCL12 than wild-type cells. Akt phosphorylation, chemotaxis, and cytokine (IL-8) secretion induced by CXCL12 were also greater in short hairpin RGS13-HMC-1 cells compared with control. RGS13 overexpression inhibited CXCL12-evoked Ca²⁺ mobilization, Akt phosphorylation and chemotaxis. These results suggest that RGS13 restricts certain GPCR-mediated biological responses of human mast cells.

The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore be hereby marked advertisement in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.

¹ This research was supported by the Intramural Research Program, National Institute of Allergy and Infectious Diseases, National Institutes of Health.

² Current address: Diabetes Branch, NIDDK, National Institutes of Health, Bethesda, MD 20892.

³ Current address: Merck Research Laboratories, Boston, MA 02115.

⁴ Current address: Wyeth Research, Cambridge, MA 02140.

⁵ Address correspondence and reprint requests to Dr. Kirk M. Druey, Laboratory of Allergic Diseases, National Institute of Allergy and Infectious Diseases, National Institutes of Health, 10 Center Drive, Bethesda, MD 20892. E-mail address: kdruey{at}niaid.nih.gov

⁶ Abbreviations used in this paper: PLC, phospholipase C; S1P, sphingosine-1-phosphate; GPCR, G protein-coupled receptor; RGS, regulator of G protein signaling; shRNA, short hairpin RNA; BMMC, bone marrow-derived mast cell; siRNA, small interfering RNA.