Activation of Adhesion GPCR EMR2/ADGRE2 Induces Macrophage Differentiation and Inflammatory Responses via Gα16/Akt/MAPK/NF-κB Signaling Pathways

Abstract

EMR2/ADGRE2 is a human myeloid-restricted adhesion G protein-coupled receptor critically implicated in vibratory urticaria, a rare type of allergy caused by vibration-induced mast cell activation. In addition, EMR2 is also highly expressed by monocyte/macrophages and has been linked to neutrophil migration and activation. Despite these findings, little is known of EMR2-mediated signaling and its role in myeloid biology. In this report, we show that activation of EMR2 via a receptor-specific monoclonal antibody promotes the differentiation of human THP-1 monocytic cell line and induces the expression of pro-inflammatory mediators, including IL-8, TNF-α, and MMP-9. Using specific signaling inhibitors and siRNA knockdowns, biochemical and functional analyses reveal that the EMR2-mediated signaling is initiated by Gα16, followed by the subsequent activation of Akt, extracellular signal-regulated kinase, c-Jun N-terminal kinase, and nuclear factor kappa-light-chain-enhancer of activated B cells. Our results demonstrate a functional role for EMR2 in the differentiation and inflammatory activation of human monocytic cells and provide potential targets for myeloid cell-mediated inflammatory disorders.

G Protein-Coupling of Adhesion GPCRs ADGRE2/EMR2 and ADGRE5/CD97, and Activation of G Protein Signalling by an Anti-EMR2 Antibody

“…Further confirmation of EMR2 G protein coupling in mammalian cells was provided by IP 1 assays, in which EMR2-CTF mediated IP 1 accumulation was dependent the presence of G α16 , in keeping with the biological pathway data of I. et al 36 . In our experiment there was no coupling to G α15 , as seen by Gupte et al 34 , but we saw coupling through a G α16z49 chimera, supporting the G αz coupling hypothesis suggested from the cAMP assay data.…”

“…The G protein-couplings of EMR2 and CD97 described here agree with data from more biologically relevant systems. For EMR2, G α16 coupling is consistent with the expression of both receptor and G protein in immune cells, where EMR2 activation of G α16 is linked to macrophage-like differentiation of THP-1 cells to mediate inflammatory responses 36 . For CD97, transfection into COS-7 cells induced a SRE-Luciferase reporter through G α12/13 -mediated Rho activation, and this has been linked to CD97's role in cancer invasion and metastasis.…”

“…Despite the relatively early discovery of EMR2 and CD97 within the aGPCR family, evidence of their G protein-coupling has been limited. EMR2 was reported to couple to rodent G α15 in a recombinant assay 34 , and to mediate inflammatory responses through G α16 activation 36 . CD97 coupling via G α12/13 has been indicated, involving heterodimerization with the lysophosphatidic acid receptor LPAR1 31 ; and the PTX-sensitive lysophosphatidylethanolamine (LPE) response of LPAR1 in MDA-MB-231 cells requires CD97 42 .…”

“…Published reports indicate that 2A1 needs to be immobilised to effect cytokine release. In our experiments, 2A1 was immobilised as described 35,36 , but experiments performed under various conditions failed to elicit any specific G protein signalling. We did also observe inhibitory effects mediated by 2A1 on control assays; therefore, for technical reasons we cannot draw conclusions on 2A1's ability to activate EMR2 G protein signalling from these systems.…”

Adhesion GPCRs in Kidney Development and Disease

“…According to the here analyzed databases aGPCRs which are related to vascularization ( Adgre1, Adgre5, Adgra2 ) (Wang et al, 2005 ; Cullen et al, 2011 ; Wandel et al, 2012 ) or inflammatory processes ( Adgre1, Adgre2, Adgre5 ) (Gray et al, 1996 ; Taylor et al, 2005 ; Hamann et al, 2007 ; Kuan-Yu et al, 2017 ) are >2.5-fold upregulated in ccRCC (Table 2 ). This reflects the high degree of vascularization of ccRCCs (Prasad et al, 2006 ) and the role of inflammatory related gene expression in its progression (Tan et al, 2011 ).…”