Transcriptomic data on the role of PEST-domain-enriched tyrosine phosphatase in the regulation of antigen-mediated activation and antiallergic action of glucocorticoids in mast cells

Protein tyrosine phosphatases and glucocorticoids are known to regulate allergic and antiallergic action in activated mast cells. Here we provide RNA sequencing and quantitative real-time PCR data from bone marrow derived mast cells, for wild-type and PEST-domain-enriched tyrosine phosphatase (PEP) null mice, activated by immunoglobulin E sensitization and dinitrophenol treatment, and additionally treated with the glucocorticoid dexamethasone. The transcriptomics experiment was performed in duplicate with a total of 16 samples (GSE108972).


Value of the data
This dataset is a transcriptomic analysis of bone marrow derived mast cells (BMMCs) from PEP þ/ þ and PEP À / À mice activated by antigen and additionally treated with glucocorticoids.
These data provide RNA-seq and qRT-PCR gene expression results to show the role of PEP in antigen-mediated mast cell activation and the inhibition of mast cell activation by glucocorticoids.
These data will serve as a reference point for antigen-mediated activation of mast cells and the antiallergic action of glucocorticoids.

Preparation of the biological material
BMMCs from PEP þ/þ and PEP À / À mice were isolated by culturing bone marrow cells from the femur and tibia of male C57BL/6 mice aged 8-10 weeks in mast cell medium (IMDM supplemented with 10% foetal calf serum, 2 mM L-glutamine, 1 mM pyruvate, 100 ng stem cell factor/kit ligand, 5 ng/ml IL3, 50 mM β-mercaptoethanol, 1% penicillin/streptomycin 10,000 U/ml) in T-75 flask at 37°C, Table 1 Outline of the different treatments administered prior to the transcriptomics study.

Sample name
Genotype Treatment 5% CO 2 and 95% humidity in an incubator. The culture medium was changed every three days by centrifuging the suspended cells at 1300 rpm for 7 min and re-suspending the resulting pellet in fresh medium as previously described [1]. The BMMCs were used after 4 weeks in culture as described in Table 1.

Preparation of libraries, sequencing and data analysis
For the RNAseq experiment, total RNA was extracted from 3 Â 10 6 PEP þ/ þ and PEP À / À BMMCs using innuPREP RNA Mini Kit (Analytik Jena AG, Jena, Germany). A total of 1 mg RNA was used to prepare mRNA sequencing libraries for each sample using the TruSeq stranded mRNA kit v2 (Illumina), according to the vendor´s protocol. Next, 10 pM of multiplexed libraries were used to generate clusters in 2 lanes of a high-throughput flowcell. A HiSeq. 1500 was used to obtain pairedend reads of 50 bases using the sequencing by synthesis (SBS v3 kit Illumina). The cluster detection and the base calling were done with the software RTA (v1.13 Illumina) and demultiplexing with the software CASAVA (v1.8.1 Illumina). The sequencing resulted in 576 million reads with a mean quality Phred score of 35.2. The quality of the sequencing data was first assessed with FASTX toolkit (v0.0.13 [http://hannonlab.cshl.edu/fastx_toolkit/]) and no pre-processing was needed. The alignment of the sequencing reads was done with Tophat2 (v2.0.11) [2] against the mouse reference genome (GRCm38 v75). The raw count per gene was calculated using HTSeq (v0.5.3) [3]. The normalization of the counts, the differential expression analysis were done using the R software packages including DESeq. 2 [4]. The reproducibility of the biological replicates for all the conditions examined was assessed by Principal Component Analysis (Fig. 1). The significantly deregulated genes for each paired comparison were subjected to hierarchical clustering using hclust and gplot R packages. These were done for the activation of the mast cells by antigen (DNP) alone (Fig. 2) or together with DEX treatment (Fig. 3).