Synergy of NUP98-HOXA10 Fusion Gene and NrasG12D Mutation Preserves the Stemness of Hematopoietic Stem Cells on Culture Condition
Synergy of NUP98-HOXA10 Fusion Gene and NrasG12D Mutation Preserves the Stemness of Hematopoietic Stem Cells on Culture Condition
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NUP98-HOXA10融合基因和NrasG12D突变的协同作用在培养条件下保留造血干细胞的干性
DOI：
10.3390/cells8090951
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发表时间：
2019-09-01
期刊：
影响因子：
6
通讯作者：
Du, Juan
中科院分区：
文献类型：
--
作者：
Dong, Yong;Xia, Chengxiang;Du, Juan
天然造血干细胞（HSC）较为敏感，在体外培养条件下往往会失去干性、分化或死亡，这对于在从多能干细胞进行方案筛选时维持真正的类HSC细胞（如果能产生的话）增加了技术挑战。内源性基因的基因编辑是否能从基因上使HSC能够承受培养压力并保持干性，在很大程度上仍然未知。在本研究中，我们发现NUP98 - HOXA10HD融合以及内源性Nras突变修饰（NrasG12D）都促进了HSC的植入竞争力。此外，这两种基因修饰的协同作用赋予了HSC在体内超强的竞争力。令人惊讶的是，单个NAV - HSC在经过体外培养后成功地保持了其干性，并显示出强大的多谱系植入能力。从机制上讲，NUP98 - HOXA10HD融合和NrasG12D突变显著改变了涉及细胞周期、细胞分裂和DNA复制的多个途径，并在HSC的背景下显著调节了包括Hoxa9、Prdm16、Hoxb4、Trim27和Smarcc1在内的干性相关基因。因此，我们开发了一种超灵敏的转基因模型，该模型可报告在培养条件下单细胞水平上HSC的存在，这可能有利于在体外从多能干细胞筛选真正的HSC再生方案。
Natural hematopoietic stem cells (HSC) are susceptible and tend to lose stemness, differentiate, or die on culture condition in vitro, which adds technical challenge for maintaining bona fide HSC-like cells, if ever generated, in protocol screening from pluripotent stem cells. It remains largely unknown whether gene-editing of endogenous genes can genetically empower HSC to endure the culture stress and preserve stemness. In this study, we revealed that both NUP98-HOXA10HD fusion and endogenous Nras mutation modifications (NrasG12D) promoted the engraftment competitiveness of HSC. Furthermore, the synergy of these two genetic modifications endowed HSC with super competitiveness in vivo. Strikingly, single NAV-HSC successfully maintained its stemness and showed robust multi-lineage engraftments after undergoing the in vitro culture. Mechanistically, NUP98-HOXA10HD fusion and NrasG12D mutation distinctly altered multiple pathways involving the cell cycle, cell division, and DNA replication, and distinctly regulated stemness-related genes including Hoxa9, Prdm16, Hoxb4, Trim27, and Smarcc1 in the context of HSC. Thus, we develop a super-sensitive transgenic model reporting the existence of HSC at the single cell level on culture condition, which could be beneficial for protocol screening of bona fide HSC regeneration from pluripotent stem cells in vitro.