Transcriptomic analysis of naïve B cells and antigen-specific activated B cells and plasma cells from 4dpi NP-LPS immunized mice [NPLPS_PC]. Transcriptomic analysis of naïve B cells and antigen-specific activated B cells and plasma cells from 4dpi NP-LPS immunized mice [NPLPS_PC]

Little is known about the earliest transcriptomic events in plasma cell differentiation. Indeed, activation of UPR-affiliated gene expression is seen in in-vitro plasma cell differentiation prior to Blimp1 upregulation and in the absence of Xbp1 as we show in linked companion series. In this study, Blimp1 reporter mice were immunized with the T-independent antigen NP-LPS four days prior to spleen harvest and naïve B cells, NP-binding activated B cells and NP-binding plasma cells were analyzed by bulk RNA-seq. Here we report the activation of ER remodeling gene expression in antigen-specific activated B cells in-vivo prior to upregulation of Blimp1 transcription. Overall design: Samples are three celltypes from three biological replicates (mice) 4 days post NP-LPS immunization. cDNA was prepared from RNA with 3' polyA capture method. Transcript abundance was measured by Kallisto psuedo-alignment and normalized (voom) and differential expression was assessed using limma.

目前学界对浆细胞分化过程中最早出现的转录组学事件尚所知甚少。正如我们在相关配套研究系列中所展示的，在体外浆细胞分化模型中，于B淋巴细胞诱导成熟蛋白1（B-lymphocyte-induced maturation protein 1, Blimp1）上调之前且缺乏X盒结合蛋白1（X-box binding protein 1, Xbp1）的情况下，即可观察到未折叠蛋白应答（Unfolded Protein Response, UPR）相关基因表达的激活。本研究中，我们将Blimp1报告小鼠（Blimp1 reporter mice）经T非依赖性抗原（T-independent antigen）NP-LPS免疫，于免疫后4天收取脾脏，并通过批量转录组测序（bulk RNA-seq）分析初始B细胞（naïve B cells）、结合NP的活化B细胞以及结合NP的浆细胞。本研究报道了在Blimp1转录上调之前，体内抗原特异性活化B细胞中内质网（Endoplasmic Reticulum, ER）重塑基因表达的激活现象。实验整体设计：样本为经NP-LPS免疫后4天的3只生物学重复小鼠的3种细胞类型。总RNA采用3'端polyA捕获法制备cDNA；转录本丰度通过Kallisto伪比对进行定量，并使用voom方法完成归一化处理；随后借助limma工具开展差异表达分析。