An atlas of gene regulatory networks for T memory cells in youth and old age [RNA-seq]

We previously identified a population of IL-10-producing, T follicular helper-like cells ("Tfh10"), linked to suppressed vaccine responses in aged mice. Here, we applied scRNA-seq and scATAC-seq to characterize Tfh10 - and the full CD4+ memory T cell (CD4+TM) compartment - in young and old mice. Unprecedented scRNA-seq coverage of the CD4+TM compartment and parallel chromatin accessibility measurements (scATAC-seq) enabled identification of 13 CD4+TM populations, which we validated as a reference through comprehensive cross-comparison to aging cell atlases and scRNA-seq studies reporting Tfh10 in other contexts. In addition, we compared sc-resolved populations to flow-sorted CD4+TM populations (Treg, Tfh10 and "nonTfh, non-Treg IL-10-") subjected to bulk RNA-seq and bulk ATAC-seq. Beyond robust characterization of age- and cell-type-dependent transcriptional landscapes, we used integrative computational modeling to predict the underlying regulatory mechanisms: We inferred gene regulatory networks (GRNs) that describe transcription-factor control of gene expression in each T-cell population and how these circuits change with age. Furthermore, we integrated our data with prior, pan-cell scRNA-seq studies to identify intercellular-signaling networks driving age-dependent changes in CD4+TM. Our atlas of finely resolved CD4+TM subsets, GRNs and cell-cell communication networks is a critical resource for analysis of biologic processes operative in memory T cells in youth and old age. The resource presents new opportunities to manipulate regulatory circuits in CD4+TM, which, long-term, could improve immune responses in the elderly. CD4+ memory T cells were harvested from spleens of young (<=4 months) and old (>=18 months) mice and subject to scRNA-seq, scATAC-seq, bulk RNA-seq, or bulk ATAC-seq.我们此前鉴定出一群产生白细胞介素10（Interleukin 10, IL-10）的滤泡辅助样T细胞（T follicular helper-like cells, "Tfh10"），该细胞群与老年小鼠的疫苗应答受抑制密切相关。本研究借助单细胞RNA测序（single-cell RNA sequencing, scRNA-seq）与单细胞转座酶可及性测序（single-cell assay for transposase-accessible chromatin using sequencing, scATAC-seq），对年轻与老年小鼠体内的Tfh10及完整CD4+记忆T细胞（CD4+ memory T cell, CD4+TM）区室开展了系统性表征。本次研究对CD4+TM区室实现了前所未有的scRNA-seq覆盖度，并结合平行染色质可及性检测，成功鉴定出13个CD4+TM细胞亚群；我们通过与衰老细胞图谱、以及其他研究场景中报道Tfh10的scRNA-seq工作进行全面交叉比对，验证了该细胞分类参考标准的可靠性。此外，我们将单细胞分辨率解析得到的细胞群，与经流式分选的CD4+TM细胞群（调节性T细胞（regulatory T cell, Treg）、Tfh10以及"非Tfh、非Treg的IL-10阴性细胞"）进行了比对，上述分选得到的细胞群均接受了批量RNA测序与批量ATAC测序。除了精准刻画年龄与细胞类型依赖的转录组特征外，我们还通过整合计算建模预测了潜在的调控机制：我们推断出基因调控网络（gene regulatory networks, GRNs），该网络描述了各T细胞群体中转录因子对基因表达的调控作用，以及这些调控环路如何随年龄发生动态改变。进一步地，我们将本研究数据与已发表的全细胞scRNA-seq研究进行整合，鉴定出驱动CD4+TM年龄依赖性变化的细胞间信号传导网络。本研究构建的高分辨率CD4+TM细胞亚群图谱、基因调控网络及细胞间通信网络，是研究青年与老年个体记忆T细胞生物学过程的关键资源。该资源为靶向调控CD4+TM的调控环路提供了全新途径，从长期来看，有望改善老年群体的免疫应答能力。本研究从小鼠脾脏中分离获取了年轻（≤4月龄）与老年（≥18月龄）小鼠的CD4+记忆T细胞，并分别开展了scRNA-seq、scATAC-seq、批量RNA测序或批量ATAC测序。