ATAC-sequencing of Th1, Tfh, and memory-like splenic CD4+ T cells on day 14 post LCMV Cl13 infection. ATAC-sequencing of Th1, Tfh, and memory-like splenic CD4+ T cells on day 14 post LCMV Cl13 infection

CD4+ T cell-derived interleukin 21 (IL-21) sustains CD8+ T cell responses during chronic viral infection, but the helper subset that confers this protection remains unclear. Here, we applied scRNA and ATAC-seq approaches to determine the heterogeneity of CD4+ T cells during LCMV clone 13 infection. Overall design: ATAC-seq was performed according to a published protocol (Buenrostro, Giresi et al. 2013). 50,000 cells of CXCR6+, CXCR5+ and CXCR6-CXCR5- PD-1hi CD4 T cell subsets (per mouse, with n=3 separate mice) were used for library construction. 37 cycles paired-end sequencing was performed on a Nextseq 550 sequencer and about fifty million reads were generated for each sample.

CD4+ T细胞衍生的白细胞介素21（interleukin 21, IL-21）可在慢性病毒感染过程中维持CD8+ T细胞应答，但介导此类保护作用的辅助T细胞亚群仍未明确。本研究借助单细胞RNA测序（single-cell RNA sequencing, scRNA）与转座酶可及性染色质测序（Assay for Transposase-Accessible Chromatin using sequencing, ATAC-seq）技术，解析淋巴细胞性脉络丛脑膜炎病毒克隆13（Lymphocytic Choriomeningitis Virus Clone 13, LCMV Clone 13）感染期间CD4+ T细胞的异质性。
实验整体设计如下：ATAC-seq实验参照已发表的实验流程（Buenrostro、Giresi等，2013年）开展。每只小鼠取5×10^4个CXCR6+、CXCR5+及CXCR6-CXCR5- PD-1hi CD4+ T细胞亚群（共3只独立生物学重复小鼠）用于文库构建。随后在Nextseq 550测序仪上进行37轮双端测序，每个样本约生成5000万条测序读段（reads）。