Hematopoietic stem ell support through transcriptional coactivator TRAP220

TRAP/Mediator复合物是一种多蛋白共激活因子复合物，可介导多种激活因子的信号。已知TRAP220/MED1亚基作为配体依赖性核受体信号和一些其他激活因子（如GATA家族蛋白）的辅助激活因子。我们最近的研究表明，TRAP220不仅在脂肪细胞，而且在白细胞（粒细胞和单核细胞）的核受体依赖性分化的介导信号中起关键作用。骨髓造血是由造血细胞和生态位细胞实现的，后者由成骨细胞和其他类型的间充质细胞组成。在这项研究中，我们评估了TRAP220在生态位细胞中的作用。尽管TRAP220基因敲除小鼠在最终造血优势形成之前是致命的，但敲除小鼠胚胎成纤维细胞（MEF）是成功制备的。因此，已知在体外支持造血干细胞/祖细胞（HSPCs）的mef被用于检查其支持HSPCs的能力。事实上，与在对照mef上培养的骨髓细胞相比，在丝裂霉素c处理的敲除mef上培养6至8周的骨髓细胞在甲基纤维素培养基中的集落形成明显减少。这一结果明显证明了TRAP220在小生境细胞中的作用，它可以诱导介导HSPCs长期支持的信号。为了观察TRAP220在MEF中支持骨髓细胞的短期作用，我们随后评估了敲除MEF和对照MEF培养的骨髓细胞的生长和存活。培养2天后，敲除mef上的骨髓细胞计数明显低于对照组。为了了解细胞数量减少的机制，我们分析了骨髓细胞在mef上培养1周和2周后的DNA合成和凋亡情况。与对照组相比，敲除mef培养的骨髓细胞中BrdU的掺入明显减少。此外，敲除mef组的凋亡细胞少于对照组。这些结果表明，mef分泌信号分子，诱导骨髓细胞增殖应激，最终导致细胞凋亡。因此，小生境细胞中的TRAP220可能介导造血干细胞支持信号和造血祖细胞生长应激信号。这是首次发现基本转录辅激活因子复合物可能在造血生态位中发挥作用。少

TRAP/Mediator complex is a multiprotein coactivator complex that mediates the signals of a variety of activators. The TRAP220/MED1 subunit is known to act as a coactivator for ligand-dependent nuclear receptor signals and some other activators such as the GATA family proteins. We have recently shown that TRAP220 plays a key role in mediating signals of nuclear receptor-dependent differentiation of not only adipocytes but white blood cells (granulocytes and monocytes). Bone marrow hematopoiesis is achieved by both hematopoietic cells and niche cells, the latter of which are comprised of osteoblasts and other type(s) of mesenchymal cells. In this study we have evaluated the role of TRAP220 in niche cells.Although the TRAP220 knockout mice am lethal before definitive hematopoiesis is dominant, the knockout mouse embryonic fibroblasts (MEF) are successfully prepared. Therefore, the MEFs, known to support hematopoietic stem/progenitor cells (HSPCs)in vitro, are used to check the ability to … More support the HSPCs. Indeed, the colony formation within the methylcellulose media was prominently decreased for bone marrow cells cultured for 6 to 8 weeks on mitomycin C-treated knockout MEFs, compared to those cultured on control MEFs. This result apparently demonstrates the role of TRAP220 in niche cells, which induce signals that mediate long-term support of HSPCs.In order to see the short-term role of TRAP220 in MEF to support bone marrow cells, we then evaluated growth and survival of bone marrow cells cultured on knockout and control MEFs. The counts of bone marrow cells on knockout MEFs were significantly decreased compared to the control ever after the 2-days culture. To know the mechanism of decreased cell numbers, we analyzed DNA synthesis and apoptosis of bone marrow cells cultured on MEFs for one or two weeks. The incorporation of BrdU into bone marrow cells cultured on knockout MEFs was prominently decreased compared to the control. Furthermore, the apoptotic cells on knockout MEFs were less than those on the control. These results indicate that the MEFs secret the signal molecules that induce proliferation stress in bone marrow cells that eventually causes apoptosis simultaneously.Thus, the TRAP220 in niche cells might mediate the signals both of the HSPCs support and the growth stress in hematopoietic progenitors. This is the first finding that the basic transctiptional coactivator complex might play a role in the hematopoietic niche. Less