Data_Sheet_3_Epigenetic Landscapes of Single-Cell Chromatin Accessibility and Transcriptomic Immune Profiles of T Cells in COVID-19 Patients.PDF

T cells play a critical role in coronavirus diseases. How they do so in COVID-19 may be revealed by analyzing the epigenetic chromatin accessibility of cis- and trans-regulatory elements and creating transcriptomic immune profiles. We performed single-cell assay for transposase-accessible chromatin (scATAC) and single-cell RNA (scRNA) sequencing (seq) on the peripheral blood mononuclear cells (PBMCs) of severely ill/critical patients (SCPs) infected with COVID-19, moderate patients (MPs), and healthy volunteer controls (HCs). About 76,570 and 107,862 single cells were used, respectively, for analyzing the characteristics of chromatin accessibility and transcriptomic immune profiles by the application of scATAC-seq (nine cases) and scRNA-seq (15 cases). The scATAC-seq detected 28,535 different peaks in the three groups; among these peaks, 41.6 and 10.7% were located in the promoter and enhancer regions, respectively. Compared to HCs, among the peak-located genes in the total T cells and its subsets, CD4+ T and CD8+ T cells, from SCPs and MPs were enriched with inflammatory pathways, such as mitogen-activated protein kinase (MAPK) signaling pathway and tumor necrosis factor (TNF) signaling pathway. The motifs of TBX21 were less accessible in the CD4+ T cells of SCPs compared with those in MPs. Furthermore, the scRNA-seq showed that the proportion of T cells, especially the CD4+ T cells, was decreased in SCPs and MPs compared with those in HCs. Transcriptomic results revealed that histone-related genes, and inflammatory genes, such as NFKBIA, S100A9, and PIK3R1, were highly expressed in the total T cells, CD4+ T and CD8+ T cells, both in the cases of SCPs and MPs. In the CD4+ T cells, decreased T helper-1 (Th1) cells were observed in SCPs and MPs. In the CD8+T cells, activation markers, such as CD69 and HLA class II genes (HLA-DRA, HLA-DRB1, and HLA-DRB5), were significantly upregulated in SCPs. An integrated analysis of the data from scATAC-seq and scRNA-seq showed some consistency between the approaches. Cumulatively, we have generated a landscape of chromatin epigenetic status and transcriptomic immune profiles of T cells in patients with COVID-19. This has provided a deeper dissection of the characteristics of the T cells involved at a higher resolution than from previously obtained data merely by the scRNA-seq analysis. Our data led us to suggest that the T-cell inflammatory states accompanied with defective functions in the CD4+ T cells of SCPs may be the key factors for determining the pathogenesis of and recovery from COVID-19.

T细胞在冠状病毒疾病中发挥关键作用。解析其在新型冠状病毒肺炎（COVID-19）中的作用机制，可通过分析顺式和反式调控元件的表观染色质可及性，并构建转录组免疫谱来实现。本研究对感染COVID-19的重症/危重症患者（SCPs）、普通型患者（MPs）以及健康志愿者对照（HCs）的外周血单个核细胞（PBMCs）开展了转座酶可及性单细胞测序（scATAC-seq）与单细胞RNA（scRNA）测序（seq）。其中，分别利用scATAC-seq（9例样本）和scRNA-seq（15例样本）纳入76570和107862个单细胞，以分析染色质可及性与转录组免疫谱特征。scATAC-seq在三组样本中共检测到28535个差异峰，其中41.6%和10.7%的峰分别位于启动子区域与增强子区域。相较于健康志愿者对照，重症/危重症患者与普通型患者的总T细胞及其亚群（CD4+ T细胞、CD8+ T细胞）中，峰关联基因富集于丝裂原活化蛋白激酶（MAPK）信号通路、肿瘤坏死因子（TNF）信号通路等炎症通路。与普通型患者相比，重症/危重症患者CD4+ T细胞中的TBX21基序可及性更低。进一步的scRNA-seq结果显示，相较于健康志愿者对照，重症/危重症患者与普通型患者的T细胞比例（尤其是CD4+ T细胞）显著下降。转录组分析结果表明，在重症/危重症患者和普通型患者的总T细胞、CD4+ T细胞以及CD8+ T细胞中，组蛋白相关基因与NFKBIA、S100A9、PIK3R1等炎症基因均呈高表达。在CD4+ T细胞中，重症/危重症患者与普通型患者的辅助性T细胞1（Th1）细胞比例均有所降低。在CD8+ T细胞中，重症/危重症患者的CD69、人类白细胞抗原II类（HLA class II）基因（HLA-DRA、HLA-DRB1、HLA-DRB5）等活化标志物均显著上调。对scATAC-seq与scRNA-seq数据的整合分析显示，两种技术的结果存在一定一致性。综上，本研究构建了COVID-19患者T细胞的染色质表观遗传状态与转录组免疫谱全景图，相较于以往仅通过scRNA-seq分析得到的数据，本研究以更高分辨率深入解析了T细胞的特征。本研究结果提示，重症/危重症患者CD4+ T细胞的炎症状态伴随功能缺陷，可能是决定COVID-19发病机制与疾病转归的关键因素。