Single cells transitioning to hematopoietic fate in vivo show pulsatile Gata2 expression

Cell fate is established through coordinated gene expression programs in individual cells. Regulatory networks that include the Gata2 transcription factor play central roles in hematopoietic fate establishment. Whereas Gata2 is essential to the embryonic development and function of hematopoietic stem cells that form the adult hierarchy, little is known of the in vivo expression dynamics of Gata2 in single cells. Here we examine Gata2 expression in single aortic cells as they establish hematopoietic fate in Gata2Venus mouse embryos. Time-lapse imaging reveals rapid pulsatile level changes in Gata2 reporter expression in cells undergoing endothelial-to-hematopoietic-transition. Moreover, Gata2 reporter pulsatile expression is dramatically altered in Gata2+/- aortic cells, which undergo fewer transitions and are reduced in hematopoietic potential. Our novel finding of dynamic pulsatile expression of Gata2 suggests a highly unstable genetic state in single cells concomitant with their transition to hematopoietic fate. This reinforces the notion that threshold levels of Gata2 influence fate establishment and has implications for transcription factor-related hematologic dysfunctions. At embryonic day 10.5, HP/SC populations (CD31+ckit+) were sorted into Gata2Venus high and intermediate populations.细胞命运通过单个细胞内协同调控的基因表达程序得以确立。包含Gata2转录因子（Gata2 transcription factor）的调控网络在造血命运确立过程中发挥核心作用。尽管Gata2对于构成成体造血层级的造血干细胞（hematopoietic stem cells）的胚胎发育与功能至关重要，但目前对于Gata2在单细胞水平的体内表达动态仍知之甚少。本研究针对Gata2Venus小鼠胚胎中，主动脉单个细胞确立造血命运过程的Gata2表达展开分析。延时成像（time-lapse imaging）结果显示，在内皮-造血转化（endothelial-to-hematopoietic-transition）过程中，细胞的Gata2报告基因表达呈现快速的脉动式水平变化。此外，Gata2杂合子（Gata2+/-）主动脉细胞的Gata2报告基因脉动表达发生显著改变，这类细胞的转化事件更少且造血潜能（hematopoietic potential）降低。本研究发现Gata2存在动态脉动表达这一全新现象，提示单个细胞向造血命运转化时伴随高度不稳定的遗传状态。这一结果进一步支持了Gata2阈值水平影响命运确立的观点，并为转录因子相关血液系统功能异常（hematologic dysfunctions）的研究提供了新的启示。在胚胎第10.5天，我们将造血祖细胞/干细胞（HP/SC）群（CD31+ckit+）分选得到Gata2Venus高表达及中等表达两个亚群。