NHGRI Hematopoiesis Section, GMBB, Bethesda, MD, USA.
Pennsylvania State University, University Park, PA, USA.
Epigenetics Chromatin. 2018 May 28;11(1):22. doi: 10.1186/s13072-018-0195-z.
Enhancers and promoters are cis-acting regulatory elements associated with lineage-specific gene expression. Previous studies showed that different categories of active regulatory elements are in regions of open chromatin, and each category is associated with a specific subset of post-translationally marked histones. These regulatory elements are systematically activated and repressed to promote commitment of hematopoietic stem cells along separate differentiation paths, including the closely related erythrocyte (ERY) and megakaryocyte (MK) lineages. However, the order in which these decisions are made remains unclear.
To characterize the order of cell fate decisions during hematopoiesis, we collected primary cells from mouse bone marrow and isolated 10 hematopoietic populations to generate transcriptomes and genome-wide maps of chromatin accessibility and histone H3 acetylated at lysine 27 binding (H3K27ac). Principle component analysis of transcriptional and open chromatin profiles demonstrated that cells of the megakaryocyte lineage group closely with multipotent progenitor populations, whereas erythroid cells form a separate group distinct from other populations. Using H3K27ac and open chromatin profiles, we showed that 89% of immature MK (iMK)-specific active regulatory regions are present in the most primitive hematopoietic cells, 46% of which contain active enhancer marks. These candidate active enhancers are enriched for transcription factor binding site motifs for megakaryopoiesis-essential proteins, including ERG and ETS1. In comparison, only 64% of ERY-specific active regulatory regions are present in the most primitive hematopoietic cells, 20% of which containing active enhancer marks. These regions were not enriched for any transcription factor consensus sequences. Incorporation of genome-wide DNA methylation identified significant levels of de novo methylation in iMK, but not ERY.
Our results demonstrate that megakaryopoietic profiles are established early in hematopoiesis and are present in the majority of the hematopoietic progenitor population. However, megakaryopoiesis does not constitute a "default" differentiation pathway, as extensive de novo DNA methylation accompanies megakaryopoietic commitment. In contrast, erythropoietic profiles are not established until a later stage of hematopoiesis, and require more dramatic changes to the transcriptional and epigenetic programs. These data provide important insights into lineage commitment and can contribute to ongoing studies related to diseases associated with differentiation defects.
增强子和启动子是与谱系特异性基因表达相关的顺式作用调节元件。先前的研究表明，不同类别的活性调节元件存在于开放染色质区域，并且每种类别都与特定的组蛋白翻译后标记物相关联。这些调节元件被系统地激活和抑制，以促进造血干细胞沿着不同的分化途径（包括密切相关的红细胞（ERY）和巨核细胞（MK）谱系）作出特定的命运决定。然而，这些决定的顺序尚不清楚。
为了描述造血过程中细胞命运决定的顺序，我们从鼠骨髓中采集原代细胞，并分离出 10 种造血细胞群，以生成转录组和染色质可及性及组蛋白 H3 赖氨酸 27 乙酰化（H3K27ac）的全基因组图谱。转录组和开放染色质图谱的主成分分析表明，巨核细胞谱系的细胞与多能祖细胞群密切相关，而红细胞则形成一个与其他群体不同的独立群体。使用 H3K27ac 和开放染色质图谱，我们发现 89%的未成熟巨核细胞（iMK）特异性活性调节区域存在于最原始的造血细胞中，其中 46%含有活性增强子标记物。这些候选活性增强子富含与巨核细胞生成必需蛋白（包括 ERG 和 ETS1）结合的转录因子结合位点基序。相比之下，只有 64%的 ERY 特异性活性调节区域存在于最原始的造血细胞中，其中 20%含有活性增强子标记物。这些区域没有富集任何转录因子的保守序列。全基因组 DNA 甲基化的纳入确定了 iMK 中显著水平的新生甲基化，但 ERY 中没有。
我们的结果表明，巨核细胞生成谱在造血早期就已建立，并存在于大多数造血祖细胞群体中。然而，巨核细胞生成并不构成“默认”的分化途径，因为巨核细胞生成伴随着大量的新生 DNA 甲基化。相比之下，红细胞生成谱直到造血的后期阶段才建立，并且需要对转录和表观遗传程序进行更剧烈的改变。这些数据为谱系决定提供了重要的见解，并为与分化缺陷相关的疾病的相关研究提供了帮助。
