Batf-mediated Epigenetic Control of Effector CD8+ T Cell Differentiation (HiChIP)

The response of naive CD8+ T cells to their cognate antigen involves rapid and broad changes in gene expression that are coupled with extensive chromatin remodeling, but the mechanisms governing these changes are not fully understood. In this study, we investigated how this process depends on the activity of the basic leucine zipper ATF-like transcription factor Batf, which is essential for the earliest phase of effector CD8+ T cell differentiation. Through genome scale profiling of multiple modalities, we characterized the role of Batf in chromatin organization at several levels, including the accessibility of key regulatory regions, the expression of nearby genes, and the interactions these regions make with each other and with key transcription factors. We quantified the dependencies between Batf and other transcription factors and identified a core transcription factor network that cooperated with Batf, including Irf4, and the transcription factors Runx3 and T-bet, which tended to co-localize with Batf and bind in regions whose accessibility and long-range interactions were mediated by Batf. We functionally demonstrated the synergistic activity of this network in initiating aspects of the effector T cells' transcriptional and chromatin accessibility profiles in an ectopically-induced fibroblast system. Using HiChIP, we further found that overexpressing all four factors in fibroblasts was required to recapitulate important aspects of the CD8+ T cell chromatin architecture. Our results provided a comprehensive resource for studying the epigenomic and transcriptomic landscape of effector differentiation of cytotoxic T cells and suggested various modes of dependencies between transcription factors in this process. Overall design: This submission contains the HiChIP portion of the data. P14 CD8+ T Cells and NIH/3T3 fibroblasts ectopically expressing Batf, Batf/Irf4, and Batf/Irf4/Runx3/T-bet were subjected to HiChIP for Batf. Additional samples were subjected to HiChIP for Ctcf.

初始CD8+ T细胞（naive CD8+ T cells）识别同源抗原后的应答反应，会发生快速且广泛的基因表达变化，并伴随大规模染色质重塑，但调控这些变化的分子机制尚未完全阐明。本研究旨在探究该过程如何依赖于碱性亮氨酸拉链ATF样转录因子（basic leucine zipper ATF-like transcription factor, Batf）的活性——该因子对于效应CD8+ T细胞分化的早期阶段至关重要。
通过全基因组多模态谱分析，我们从多个维度解析了Batf在染色质组织中的调控作用，涵盖关键调控区域的染色质可及性、邻近基因的表达情况，以及这些区域彼此间和与关键转录因子间的相互作用。我们定量分析了Batf与其他转录因子之间的调控依赖关系，并鉴定出一个与Batf协同发挥功能的核心转录因子网络：该网络包含干扰素调节因子4（Irf4）、Runt相关转录因子3（Runx3）以及T-bet，这些因子倾向于与Batf共定位，并结合在由Batf介导染色质可及性和远程相互作用的区域。
我们通过异位诱导的成纤维细胞体系，功能验证了该核心网络在启动效应T细胞转录组和染色质可及性特征方面的协同调控活性。进一步利用HiChIP（High-throughput Chromosome Conformation Capture followed by Immunoprecipitation）技术发现，在成纤维细胞中过表达全部这四个转录因子，才能重现CD8+ T细胞染色质结构的关键特征。
本研究结果为解析细胞毒性T细胞效应分化的表观基因组和转录组图谱提供了全面的数据资源，并揭示了该过程中不同转录因子间的多种调控依赖模式。
实验设计概况：本提交数据包含HiChIP实验部分。分别对P14 CD8+ T细胞，以及异位表达Batf、Batf/Irf4、Batf/Irf4/Runx3/T-bet的NIH/3T3成纤维细胞进行针对Batf的HiChIP实验；额外样本则针对Ctcf（CCCTC-binding factor）进行HiChIP实验。