Colon tissues were collected from UC patients for bulk RNAseq and lamina propria cells were isolated for MRGPRX2 activation studies and single-cell RNA sequencing (scRNAseq). Genetic association of all protein altering GPCR SNPs was performed in an Ashkenazi Jewish UC case-control cohort. Variants of MRGPRX2 were transfected into CHO and HMC-1.1 cells to detect genotype-dependent effects on β-arrestin recruitment, IP-1 accumulation, and phosphoERK.
Mast cell-specific mediators and ADM (adrenomedullin, proteolytic precursor of PAMP-12, an MRGPRX2 agonist) are upregulated in inflamed compared to uninflamed UC. MRGPRX2 stimulation induces carboxypeptidase secretion from inflamed UC. Of all protein-altering GPCR alleles, a unique variant of MRGPRX2, Asn62Ser, was most associated, bioinformatically predicted to alter arrestin recruitment. We validated that the UC protective serine allele enhances beta-arrestin recruitment, decreases IP-1, and increases phosphoERK with MRGPRX2 agonists. scRNASeq defines that ADM is expressed by activated fibroblasts and epithelial cells, and that IFNG is a key, upstream regulator of mast cell gene expression.
Inflamed UC regions are distinguished by MRGPRX2-mediated activation of mast cells, with decreased activation observed with a UC-protective genetic variant. These results define cell modules of UC activation and a new therapeutic target.
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