Epigenetic and transcriptomic alterations in key inflammatory pathways are established in RUNX1 deficient hematopoietic progenitors and are propagated to neutrophils [HI-C]. Epigenetic and transcriptomic alterations in key inflammatory pathways are established in RUNX1 deficient hematopoietic progenitors and are propagated to neutrophils [HI-C]

Epigenome and transcriptome characterization of RUNX1 deficient hematopoietic cells. We hypothesized that epigenetic alterations in key inflammatory pathway genes are acquired in RUNX1 deficient granulocyte-monocyte progenitors (GMPs) and are propagated to neutrophils. We deleted RUNX1 using Cebpa-Cre, which deletes primarily in GMPs. We analyzed the basal transcriptional changes caused by the loss of RUNX1 in purified GMPs and neutrophils by bulk RNA-seq. We evaluated if the loss of RUNX1 leads to changes in chromatin accessibility in GMPs and neutrophils by ATAC-seq. We determined that there was increased chromatin accessibility of transposable elements (TEs) in RUNX1 deficient cells. To determine if we could detect dsRNA from TEs, we pulled down dsRNA using the 9D5 antibody and performed RNA-seq. To gain insight into how RUNX1 loss affects active enhancers, we performed H3K27ac ChIP-seq on control and Runx1 deficient neutrophils. We also evaluated if RUNX1 mutations in patient neutrophils lead to changes in chromatin accessibility by ATAC-seq. We then performed Hi-C to evaluate the global high-order chromatin organization in GMPs and neutrophils. Finally, we performed RUNX1 CUT&RUN to determine RUNX1 occupancy sites in GMPs. Overall design: Examination of transcriptome, chromatin accessibility, and histone modifications in RUNX1 mouse deficient GMPs and neutrophils, and patient neutrophils. Please note that each processed data file was generated from both A and B replicates, and is linked to the corresponding *_A sample records.

RUNX1缺陷型造血细胞的表观基因组与转录组特征分析。本研究提出假说：关键炎症通路基因的表观遗传改变，可在RUNX1缺陷型粒单核细胞祖细胞（granulocyte-monocyte progenitors, GMPs）中产生，并传递至中性粒细胞。我们利用Cebpa-Cre系统敲除RUNX1，该系统主要在粒单核细胞祖细胞中实现基因敲除。我们通过批量RNA测序（bulk RNA-seq），分析了纯化的粒单核细胞祖细胞与中性粒细胞中，RUNX1缺失所导致的基础转录组变化。我们通过转座酶可及性测序（ATAC-seq），评估了RUNX1缺失是否会改变粒单核细胞祖细胞与中性粒细胞的染色质可及性。研究发现，RUNX1缺陷细胞中转座元件（transposable elements, TEs）的染色质可及性显著提升。为检测转座元件产生的双链RNA（double-stranded RNA, dsRNA），我们使用9D5抗体富集双链RNA并开展了RNA测序。为探究RUNX1缺失如何调控活性增强子，我们对对照组与Runx1缺陷型中性粒细胞开展了H3K27ac染色质免疫沉淀测序（ChIP-seq）。我们还通过转座酶可及性测序（ATAC-seq），评估了患者中性粒细胞中RUNX1突变是否会改变染色质可及性。随后我们通过Hi-C技术，分析了粒单核细胞祖细胞与中性粒细胞的全局高阶染色质结构。最后，我们通过RUNX1 CUT&RUN技术，确定了粒单核细胞祖细胞中RUNX1的结合位点。整体实验设计：分析RUNX1缺陷型小鼠粒单核细胞祖细胞、中性粒细胞，以及患者中性粒细胞中的转录组、染色质可及性与组蛋白修饰。请注意，所有处理后的数据文件均由A、B两个生物学重复样本生成，并与对应的*_A样本记录相关联。