IL-4-BATF signaling directly modulates IL-9 producing mucosal mast cell (MMC9) function in experimental food allergy

IL-4-BATF signaling directly modulates IL-9 producing mucosal mast cell (MMC9) function in experimental food allergy In this study we identified the IL-4 signaling-mediated regulation of small intestinal (SI) MMC9s using bulk RNAseq. We also identified the MMC9 transcriptome at the single cell level using single cell RNAseq. For the first goal, we employed an adoptive transfer model of food allergy, where ovalbumin (OVA)-sensitized wild type (WT) mice are reconstituted with bone marrow progenitors of either WT, IL-4RaY709F (IL-4RA gain of function mutant) or IL-4Ra-/- (loss of function mutant) mice, followed by 7 repeated oral Ova challenges (3 times per week). This approach generated SI-MMC9s of WT, IL-4RaY709F and IL-4Ra-/- origins. SI-MMC9s were sort purified as described in the study, and their gene expressions identified by bulk RNAseq. Each RNA sample was obtained by pooling from 12-20 biological replicates. For the second goal, we performed single cell RNAseq on 81 sort purified SI-WT MMC9s from mice that had undergone Ova sensitization, followed repeated oral Ova challenges. We obtained scRNA-seq reads from 81 SI Lin- IL-4GFPhigh IL-17RB2- c-Kit+ ST2+ β7-integrinlow cells that were aligned to the reference mouse genome (GRCm38) to obtain relative gene expression as RPKM as described in the bulk sequencing analyses.