P17 induces chemotaxis and differentiation of monocytes via MRGPRX2-mediated mast cell-line activation.
J Allergy Clin Immunol
; 149(1): 275-291, 2022 01.
Article
em En
| MEDLINE
| ID: mdl-34111449
ABSTRACT
BACKGROUND:
P17, a peptide isolated from Tetramorium bicarinatum ant venom, is known to induce an alternative phenotype of human monocyte-derived macrophages via activation of an unknown G protein-coupled receptor (GPCR).OBJECTIVE:
We sought to investigate the mechanism of action and the immunomodulatory effects of P17 mediated through MRGPRX2 (Mas-related G protein-coupled receptor X2).METHODS:
To identify the GPCR for P17, we screened 314 GPCRs. Upon identification of MRGPRX2, a battery of in silico, in vitro, ex vivo, and in vivo assays along with the receptor mutation studies were performed. In particular, to investigate the immunomodulatory actions, we used ß-hexosaminidase release assay, cytokine releases, quantification of mRNA expression, cell migration and differentiation assays, immunohistochemical labeling, hematoxylin and eosin, and immunofluorescence staining.RESULTS:
P17 activated MRGPRX2 in a dose-dependent manner in ß-arrestin recruitment assay. In LAD2 cells, P17 induced calcium and ß-hexosaminidase release. Quercetin- and short hairpin RNA-mediated knockdown of MRGPRX2 reduced P17-evoked ß-hexosaminidase release. In silico and in vitro mutagenesis studies showed that residue Lys8 of P17 formed a cation-π interaction with the Phe172 of MRGPRX2 and [Ala8]P17 lost its activity partially. P17 activated LAD2 cells to recruit THP-1 and human monocytes in Transwell migration assay, whereas MRGPRX2-impaired LAD2 cells cannot. In addition, P17-treated LAD2 cells stimulated differentiation of THP-1 and human monocytes, as indicated by the enhanced expression of macrophage markers cluster of differentiation 11b and TNF-α by quantitative RT-PCR. Immunohistochemical and immunofluorescent staining suggested monocyte recruitment in mice ears injected with P17.CONCLUSIONS:
Our data provide novel structural information regarding the interaction of P17 with MRGPRX2 and intracellular pathways for its immunomodulatory action.Palavras-chave
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Base de dados:
MEDLINE
Assunto principal:
Peptídeos
/
Receptores Acoplados a Proteínas G
Idioma:
En
Ano de publicação:
2022
Tipo de documento:
Article