A purine metabolic checkpoint that prevents autoimmunity and autoinflammation (T cell RNA Seq dataset)

In this study, we investigate how FAMIN activity in dendritic cells determines the release of mediators that affect priming, activation and gene transcription of CD8+ T cells. Supernatant from Famin-/- and Famin+/+ splenic CD11c+ dendritic cells were harvested following overnight incubation. CD8+ T cells were isolated from spleens and lymph nodes of OT-I;Rag–/– mice, stimulated with anti-CD3 and CD28 in the presence of supernatant obtained from Famin-/- or Famin+/+ dendritic cells for 24 hours. Total RNA was extracted using the RNeasy Mini Kit (QIAGEN, 74104) in accordance with the manufacturer's instructions. Libraries were prepared using TruSeq stranded mRNA library prep kit (Illumina, 20020594). Sequencing of libraries was performed using an Illumina NextSeq 500 platform with NextSeq 500-Mid Output kit generating 2x150bp end reads. FastQ files were quality-checked and any residual adaptor sequences were removed using TrimGalore. Reads were then aligned to the reference genome Ensembl Mus_musculus.GRCm38 using STAR. Differential gene expression analysis was performed on read count files using edgeR.

本研究旨在探究树突状细胞（dendritic cells）中FAMIN的活性如何调控介导CD8+ T细胞（CD8+ T cells）致敏、活化及基因转录的介质释放。将Famin基因敲除纯合子（Famin-/-）与野生型（Famin+/+）脾脏CD11c+树突状细胞经过夜培养后收集其上清液。从OT-I;Rag–/–小鼠的脾脏及淋巴结中分离CD8+ T细胞，在Famin-/-或Famin+/+树突状细胞上清液存在的条件下，以抗CD3抗体与抗CD28抗体刺激培养24小时。严格按照试剂盒说明书，使用RNeasy Mini Kit（QIAGEN, 74104）提取总RNA。采用TruSeq链特异性mRNA文库制备试剂盒（Illumina, 20020594）构建测序文库。使用Illumina NextSeq 500测序平台搭配NextSeq 500中端输出试剂盒完成文库测序，生成2×150bp双端读段。使用TrimGalore对FastQ文件进行质量质控并去除残留接头序列。随后使用STAR软件将读段比对至参考基因组Ensembl Mus_musculus.GRCm38。最后使用edgeR软件对读段计数文件开展差异基因表达分析。