MRGPRX2 Signaling in Mast Cells and Mastocytosis (Urticaria Pigmentosa)
Saudi Digital Library
 The neuropeptide substance P (SP) contributes to neurogenic inflammation, pain and atopic dermatitis in mice through the activation of mast cells via MAS-related G protein-coupled receptor-B2 (MrgprB2). SP also induces degranulation in human MCs through MRGPRX2, which is the human counterpart of MrgprB2. MRGPRX2 contains six tyrosine residues, five located in transmembrane domains and one in the receptor’s intracellular loop 2 (ICL2). Tyr279 in MRGPRX2’s seventh transmembrane domain contained within the motif NPxxY is important for coupling to G proteins. In addition to G protein, selected MRGPRX2 ligands also couple to β-arrestin-mediated signaling. However, the roles of MRGPRX2’s tyrosine residues on G protein and β-arrestin-mediated signaling is unknown. To address this question, we prepared cDNA encoding MRGPRX2 mutants Y67A, Y89A, Y113A, Y137A, Y222A or Y279A and generated transient transfectants in RBL-2H3 cells. We found that while cells expressing wild-type (WT), Y89A and Y222A responded normally to SP for Ca2+ mobilization and degranulation, mutants Y67A, Y113A, Y137A and Y279A were unresponsive. To determine if G protein independent signaling mediates β-arrestin recruitment. We cloned Y279A mutant into Tango vector and determined the ability to activate β-arrestin by using SP. We found mutant Y279A failed to recruit β-arrestin. These findings suggest that four tyrosine residues at MRGPRX2 show loss of function phenotype. Moreover, this is the first demonstration that a single amino acid Tyr279 residue is involved in coupling to both G protein and β- arrestin.  Mastocytosis is a condition that occurs when MCs accumulate in the skin and/or internal organs. It is divided into two main categories systemic mastocytosis (SM) and cutaneous mastocytosis (CM). Urticaria pigmentosa is the most common type of CM that is mainly presented as generalized small macules and papules of a deep-brown color that affects the entire body. The etiology of the disease is mainly due to a mutation of a KIT gene. However, the MCs’ activation mechanism is still unknown. To address this question, we used immunohistochemistry (IHC) staining to confirm the presence of MCs. After that, we utilized double immunofluorescence (IF) staining to detect MRGPRX2 signals. We found that the number of MCs is higher in the diseased samples compared to the control. Moreover, we detected MRGPRX2 signals in all MCs of patients with urticaria pigmentosa. Although the mechanism of mastocytosis is still unknown, this is the first demonstration that MRGPRX2 is expressed at high level in mast cells of patients of urticaria pigmentosa. These findings may help for further investigation about the role of MRGPRX2 in this heterogeneous disease.