Next Generation Sequencing of LSK EpCAM+ and EpCAM- of Zeb1-/-

Zeb1, a zinc finger E-box binding homeobox epithelial-mesenchymal (EMT) transcription factor, confers properties of ‘stemness’, such as self-renewal, in cancer. Yet little is known about the function of Zeb1 in adult stem cells. Here, we used the hematopoietic system, as a well-established paradigm of stem cell biology, to evaluate Zeb1 mediated regulation of adult stem cells. We employed a conditional genetic approach using the Mx1-Cre system to specifically knockout (KO) Zeb1 in adult hematopoietic stem cells (HSCs) and their downstream progeny. Acute genetic deletion of Zeb1 led to rapid onset thymic atrophy and apoptosis driven loss of thymocytes and T cells. A profound cell-autonomous self-renewal defect and multi-lineage differentiation block was observed in Zeb1 KO HSCs. Loss of Zeb1 in HSCs activated transcriptional programs of deregulated HSC maintenance and multi-lineage differentiation genes, and of cell polarity, consisting of cytoskeleton, lipid metabolism/lipid membrane and cell adhesion related genes. Notably, Epithelial cell adhesion molecule (EpCAM) expression was prodigiously upregulated in Zeb1 KO HSCs, which correlated with their enhanced cell survival capacity and diminished differentiation in transplantation. Thus, Zeb1 acts as a crucial transcriptional regulator in hematopoiesis, co-ordinating HSC self-renewal and multi-lineage differentiation fates, in part, via EpCAM repression.

Zeb1作为一种锌指E盒结合同源盒上皮间质转化（Epithelial-Mesenchymal Transition, EMT）转录因子，可赋予癌细胞如自我更新一类的“干性（stemness）”特征。然而目前对Zeb1在成体干细胞中的功能却知之甚少。本研究以造血系统——干细胞生物学领域公认的经典研究范式——为模型，探究Zeb1对成体干细胞的调控作用。我们借助基于Mx1-Cre系统的条件性遗传学策略，特异性地在成体造血干细胞（hematopoietic stem cells, HSCs）及其下游子代细胞中敲除（knockout, KO）Zeb1。Zeb1的急性基因缺失会快速引发胸腺萎缩，并通过凋亡介导胸腺细胞与T细胞的丢失。在Zeb1基因敲除的造血干细胞中，可观察到严重的细胞自主性自我更新缺陷与多系分化阻滞。造血干细胞中Zeb1的缺失会激活多类转录程序，涵盖失调的造血干细胞维持与多系分化相关基因，以及涉及细胞骨架、脂质代谢/脂质膜和细胞黏附的细胞极性相关基因。值得注意的是，上皮细胞黏附分子（Epithelial cell adhesion molecule, EpCAM）的表达在Zeb1基因敲除的造血干细胞中被显著上调，这与其移植后增强的细胞存活能力以及减弱的分化潜能密切相关。综上，Zeb1是造血过程中的关键转录调控因子，可协调造血干细胞的自我更新与多系分化命运，其部分作用机制依赖于对EpCAM的转录抑制。