GSE160131
Single cell hibernation cultures identify molecules dispensable for haematopoietic stem cell self-renewal
Advances in the isolation and gene expression profiling of single haematopoietic stem cells (HSCs) have permitted in-depth resolution of their molecular program.  However, long-term HSCs (LT-HSCs) can only be isolated to near purity from adult mouse bone marrow, thereby precluding studies of their molecular program in different physiological states. Here, we describe a powerful 7-day culture system that removes single LT-HSCs from their native bone marrow microenvironment without compromising their functional potential. LT-HSCs are maintained as single cells and retain full HSC function compared to freshly isolated LT-HSCs with respect to divisional kinetics, colony forming cell potential (73.9%), and transplantation into primary (53%) and secondary (44%) recipients. Comparison of molecular profiles of single hibernating LT-HSCs to freshly isolated counterparts showed a high degree of similarity, but also identified key factors dispensable for LT-HSC function in transplantation assays including members of the AP1 complex (Jun, Fos, and Ncor2), Sult1a1 and Cish. Overall design: We performed single-cell RNA-sequencing on LT-HSCs cultured in hibernating conditions for 7 days (n=106) and compared them to freshly isolated single LT-HSCs (n=165) and also to LT-HSCs stimulated with SCF for 16 hours (from both HSC+SCF (n=63) and hibHSC+SCF(n=127).