Robust hematopoietic specification requires the ubiquitous Sp1 and Sp3 transcription factors [ATAC-Seq]. Robust hematopoietic specification requires the ubiquitous Sp1 and Sp3 transcription factors [ATAC-Seq]

Development requires the cooperation of tissue-specific and ubiquitously expressed transcription factors, such as Sp-family members. However, the molecular details of how ubiquitous factors participate in developmental processes are still unclear. We previously showed that during the differentiation of embryonic stem cells lacking Sp1 DNA binding activity (Sp1deltaDBD/deltaDBD cells), early blood progenitors are formed. However, gene expression during differentiation becomes progressively deregulated and terminal differentiation is severely compromised. Here we studied the cooperation of Sp1 and its closest paralogue Sp3 in hematopoietic development and demonstrate that Sp1 and Sp3 binding sites largely overlap. Sp3 cooperates with Sp1deltaDBD/deltaDBD but is unable to support hematopoiesis in the complete absence of Sp1. Using single cell gene expression analysis, we show that the lack of Sp1 DNA binding leads to a distortion of cell fate decision timing, indicating that stable chromatin bi nding of Sp1 is required to maintain robust differentiation trajectories. Overall design: ATAC-Seq in A17 or E14 cells in ESC, Flk+Cells, with either WT, Sp1deltaDBD, Sp1hetrozygous, Sp1KO

生物体发育依赖于组织特异性转录因子与遍在表达转录因子的协同作用，例如Sp家族（Sp-family）成员。然而，遍在表达转录因子参与发育进程的具体分子机制仍未阐明。此前我们的研究发现，在缺失Sp1 DNA结合活性的胚胎干细胞（Sp1deltaDBD/deltaDBD细胞）的分化过程中，可生成早期血液祖细胞。但在分化过程中，基因表达会逐渐出现调控紊乱，终末分化过程也会受到严重阻碍。本研究聚焦Sp1及其最近缘旁系同源基因Sp3在造血发育中的协同作用，证实Sp1与Sp3的结合位点存在广泛重叠。Sp3可与Sp1deltaDBD/deltaDBD发挥协同作用，但在Sp1完全缺失的情况下无法支持造血过程。通过单细胞基因表达分析，我们发现Sp1 DNA结合功能的缺失会扰乱细胞命运决定的时序，这表明Sp1的稳定染色质结合对于维持稳健的分化轨迹至关重要。实验整体设计：对A17或E14细胞在胚胎干细胞（ESC）、Flk+细胞两种状态下进行转座酶可及性测序（ATAC-Seq），细胞基因型分别为野生型（WT）、Sp1deltaDBD、Sp1杂合子（Sp1hetrozygous）以及Sp1敲除型（Sp1KO）。