Identification of immune-activated hematopoietic stem cells

Hematopoietic stem cells (HSCs) are the source of both normal blood cells and leukemia [ 1 ]. The ability to identify and purify them using multiple surface markers, such as Lineage - cKit + Sca1 + CD34 − CD48 − CD150 + , facilitates the study of molecular mechanisms and dynamics of hematopoiesis [ 2 , 3 ]. However, the identification of HSCs under immune-stimulation conditions poses a major challenge due to considerable changes in their pivotal markers, including Sca1 and CD150, following stimulation [ 4 , 5 , 6 , 7 , 8 , 9 , 10 ]. Therefore, we employ the Fgd5 mCherry reporter mouse to improve identification of activated HSCs. Fgd5 is specifically expressed in HSCs of all hematopoietic cells, and the knocked-in Fgd5 mCherry reporter mouse was reported to specifically label HSCs under naive-state [ 11 ]. Here, we utilize Fgd5 mCherry to demonstrate the specific labeling of long-term multipotent cells under immune-stimulatory conditions, achieving better identification of activated-HSCs. RNA-Seq. data of purified Lineage - cKit + Sca1 + CD150 + Fgd5 mCherry+ (LSK150 + mC + ) reveal molecular-changes in HSCs upon stress. We further identify CD317 ( Bst2) and CD69 as new activation-markers of HSCs, demonstrating the response to stimulation at single-cell FACS resolution. This study thus presents the first mouse reporter for HSCs during immune stimulation, and highlights novel markers to monitor HSCs under stress conditions.