Tracing haematopoietic stem cell formation at single-cell resolution

Abstract

Haematopoietic stem cells (HSCs) are derived early from embryonic precursors, such as haemogenic endothelial cells and pre-haematopoietic stem cells (pre-HSCs), the molecular identity of which still remains elusive. Here we use potent surface markers to capture the nascent pre-HSCs at high purity, as rigorously validated by single-cell-initiated serial transplantation. Then we apply single-cell RNA sequencing to analyse endothelial cells, CD45(-) and CD45(+) pre-HSCs in the aorta-gonad-mesonephros region, and HSCs in fetal liver. Pre-HSCs show unique features in transcriptional machinery, arterial signature, metabolism state, signalling pathway, and transcription factor network. Functionally, activation of mechanistic targets of rapamycin (mTOR) is shown to be indispensable for the emergence of HSCs but not haematopoietic progenitors. Transcriptome data-based functional analysis reveals remarkable heterogeneity in cell-cycle status of pre-HSCs. Finally, the core molecular signature of pre-HSCs is identified. Collectively, our work paves the way for dissection of complex molecular mechanisms regulating stepwise generation of HSCs in vivo, informing future efforts to engineer HSCs for clinical applications.

Human haematopoietic stem cell development: from the embryo to the dish

“…However, although IAHCs disappear by CS 17 (35-38 dpc), HSCs are still detectable at this stage indicating that functional HSCs in the human AGM region do not entirely correlate with the presence of IAHCs (Tavian et al, , 1999. As in the mouse, the number of IAHCs significantly exceeds the low numbers of HSCs in the AGM , suggesting that many cells in human IAHCs could be nascent HSCs, that cannot yet be detected by direct transplantation, as inferred from in vitro modelling of mouse HSC development (Taoudi et al, 2008;Rybtsov et al, 2011;Zhou et al, 2016). The development of a similar in vitro co-culture approach for recapitulating human HSC development has been unexpectedly challenging.…”

“…These bud predominantly from the endothelial floor of the dorsal aorta, which accordingly coexpress endothelial and haematopoietic markers. The search for markers that would accurately identify developing HSCs and separate them from non-self-renewing progenitors is an important goal towards understanding the mechanisms that underlie HSC development Zhou et al, 2016). Important events that occur downstream of HSC emergence include colonisation and expansion of HSCs in the foetal liver and subsequent lodging in the adult bone marrow.…”

Will targeting PI3K/Akt/mTOR signaling work in hematopoietic malignancies?

“…Raptor and PRAS40 belong specifically to mTORC1, while Rictor, mSin1 and Protor are specific components of mTORC2 (56,57). mTOR complexes play critical roles in both normal hematopoiesis and leukemogenesis (58,59). mTOR hyperactivation was observed in various cancers of 123 individual studies in the cBioportal (http://www.cbioportal.org/, data not shown).…”

Understanding development and stem cells using single cell-based analyses of gene expression

“…These investigations range from defining the molecular profiles of hematopoietic stem cells (HSCs) from mouse embryos (Zhou et al, 2016) and the transcriptional landscape of heart development (DeLaughter et al, 2016;Li et al, 2016a), to comparing cell type diversity in the embryonic midbrain between human and mouse (La Manno et al, 2016) and obtaining initial cellular censuses of the murine spleen (Jaitin et al, 2014), cortex and hippocampus (Zeisel et al, 2015). We list many of these studies in Table 2 but note that more studies are being published every month.…”