β-arrestin-1 and β-arrestin-2 Restrain MRGPRX2-Triggered Degranulation and ERK1/2 Activation in Human Skin Mast Cells

Abstract

As a novel receptor that efficiently elicits degranulation upon binding to one of its numerous ligands, MRGPRX2 has moved to the center of attention in mast cell (MC) research. Indeed, MRGPRX2 is believed to be a major component of pseudo-allergic reactions to drugs and of neuropeptide-elicited MC activation in skin diseases alike. MRGPRX2 signals via G proteins which organize downstream events ultimately leading to granule discharge. Skin MCs require both PI3K and ERK1/2 cascades for efficient exocytosis. β-arrestins act as opponents of G proteins and lead to signal termination with or without subsequent internalization. We recently demonstrated that ligand-induced internalization of MRGPRX2 requires the action of β-arrestin-1, but not of β-arrestin-2. Here, by using RNA interference, we find that both isoforms counter skin MC degranulation elicited by three MRGPRX2 agonists but not by FcεRI-aggregation. Analyzing whether this occurs through MRGPRX2 stabilization under β-arrestin attenuation, we find that reduction of β-arrestin-1 indeed leads to increased MRGPRX2 abundance, while this is not observed for β-arrestin-2. This led us speculate that β-arrestin-2 is involved in signal termination without cellular uptake of MRGPRX2. This was indeed found to be the case, whereby interference with β-arrestin-2 has an even stronger positive effect on ERK1/2 phosphorylation compared to β-arrestin-1 perturbation. Neither β-arrestin-1 nor β-arrestin-2 had an impact on AKT phosphorylation nor affected signaling via the canonical FcεRI-dependent route. We conclude that in skin MCs, β-arrestin-2 is chiefly involved in signal termination, whereas β-arrestin-1 exerts its effects by controlling MRGPRX2 abundance.

CREB Is Activated by the SCF/KIT Axis in a Partially ERK-Dependent Manner and Orchestrates Survival and the Induction of Immediate Early Genes in Human Skin Mast Cells

“…A well-established and efficient siRNA method for skMCs was utilized [ 29 , 53 , 57 , 58 , 99 , 100 , 101 , 102 ]. In brief, skMCs were transfected twice (on day 0 and day 1) by CREB-targeting siRNA or control siRNA (each at 1 µM) for a total of 2 or 3 d in Accell ® medium (Dharmacon, Lafayette, CO, USA) (supplemented with Non-Essential Amino Acids and L-Glutamine (both from Carl Roth)).…”

MRGPRX2 signaling involves the Lysyl-tRNA synthetase and MITF pathway

“…MRGPRX2-triggered degranulation depends on Gαi, Gαq, Ca ++ channels, ERK, and PI3K ( 14 , 15 ). Cytoskeleton dynamics and granule exocytosis need Cdc42 GTPase activation and the participation of unconventional class I myosins, MYO1F ( 16 ), and β1 and β2 arrestins regulate the kinetics and the extent of MRGPRX2 cellular activity ( 17 ). The present study adds more insights into MRGPRX2 signals and demonstrates the involvement of the LysRS-MITF pathway.…”

“…Furthermore, GTPase activation (Cdc42) upstream of the unconventional class I myosin 1f (MYO1F) regulates actin cytoskeleton dynamics and granule release ( 16 ). Lately, β arrestins 1 and 2 (involved in GPCR desensitization) have been proposed to interfere with MRGPRX2-dependent degranulation ( 17 ).…”

The MRGPR family of receptors in immunity

“…Notably, unlike AG-30/5C and LL-37, codeine and C48/80 induce MRGPRX2 internalization through β-arrestin 20 , 26 , 104 , which may function to restrain hyperactivation of MRGPRX2. 105 Downstream of these pathways, MRGPRX2-mediated degranulation in response to C48/80 and substance P requires ERK and PI3K in primary human mast cells. 106 , 107 Similarly, C48/80- and LL-37-induced chemokines also appear to rely on ERK activation in the LAD2 human mast cell line.…”