A network including TGFβ/Smad4, Gata2 and p57 regulates proliferation of mouse hematopoietic stem/progenitor cells [ChIP-seq]

Transforming growth factor-β (TGFβ) is a potent inhibitor of hematopoietic stem cell (HSC) proliferation. However, the precise mechanism for this effect is unknown. Here, we have identified the transcription factor Gata2, previously described as an important regulator of HSC function, as an early and direct target gene for TGFβ-induced Smad signaling in hematopoietic stem and progenitor cells (HSPCs). Interestingly, TGFβ-induced Gata2 upregulation is critical for subsequent transcriptional activation of the TGFβ signaling effector molecule p57 and resulting growth arrest of HSPCs. Importantly, both Gata2 and p57 are abundantly expressed in freshly isolated highly purified HSCs, demonstrating the relevance of this circuit in HSC regulation within the HSC niche. Our results connect key molecules involved in HSC self-renewal and reveal a functionally relevant network regulating proliferation of primitive hematopoietic cells. To identify TGFβ targets downstream of Gata2, we carried out a ChIP-Seq experiment on TGFβ-induced Lhx2 cells. Interestingly, there was a large overlap between the GATA2-bound genes and genes differentially expressed after 2h TGFβ induction. One sample of 1x10^8 cells (treated with 10 ng/ml TGFβ for 2h) was sequenced.

转化生长因子-β（TGFβ）是造血干细胞（hematopoietic stem cell，HSC）增殖的强效抑制剂，但其介导该效应的确切分子机制尚未阐明。本研究鉴定发现，转录因子Gata2——此前被证实为HSC功能的重要调控因子——是造血干祖细胞（hematopoietic stem and progenitor cells，HSPCs）中TGFβ诱导的Smad信号通路的早期直接靶基因。值得注意的是，TGFβ诱导的Gata2上调，对于后续TGFβ信号通路效应分子p57的转录激活以及由此引发的HSPCs生长阻滞至关重要。尤为关键的是，Gata2与p57在新鲜分离的高纯度HSCs中均呈高表达状态，这表明该调控环路在造血干细胞龛内的HSC生理调控中具有实际相关性。本研究结果串联了参与HSC自我更新的核心分子，并揭示了调控原始造血细胞增殖的功能相关调控网络。为鉴定Gata2下游的TGFβ靶基因，我们对TGFβ诱导的Lhx2细胞开展了染色质免疫共沉淀测序（ChIP-Seq）实验。结果显示，GATA2结合靶基因与TGFβ诱导2小时后发生差异表达的基因存在大量重叠。本实验对1×10⁸个经10 ng/ml TGFβ处理2小时的细胞进行了单样本测序。