Rictor is pivotal for the emergence and function of embryonic hematopoietic stem cells

Abstract

Engineering hematopoietic stem cells (HSC) from pluripotent stem cells has tremendous clinical significance. Despite efficient production of hematopoietic progenitor cells (HPC), the generation of authentic HSC remains unsuccessful, reflecting poor understanding of regulatory mechanisms specifically regarding HSC ontogeny. Here, we disrupted Rictor, a core mTORC2 component, at distinct developmental stages with Cre/LoxP strategy. Rictor deficiency from embryonic endothelial stage (using Tie2-Cre) had minimal effect on the generation of definitive HPC in vivo and in vitro. In striking contrast, the endothelial emergence of first cohort of HSC was severely blocked in the AGM region. only 1 out of 15 recipients of Tie2-Cre;Rictorf/f AGM cells was successfully repopulated with markedly lower chimerism (15.3%), whereas 12 out of 19 recipients of the control AGM cells were repopulated with a much higher long-term chimerism (54.5 ± 25.1%). Interestingly, deleting Rictor from developing hematopoietic stage (using Vav-Cre), other than within adult bone marrow (using Mx1-Cre), significantly impaired the HSC function in vivo. When E11.5 control AGM and E12.5 FL cells were transplanted, all recipients were repopulated with an average long-term chimerism of 58.2±2.1% and 77.2±11.1% respectively as compared with the markedly reduced chimerism of Vav-Cre;Rictorf/f AGM group (29.9±14.6%) and FL group (37.0±16.3%) respectively. We thus demonstrate that Rictor is vital for endothelial transition to HSC (other than HPC) and for HSC function at embryonic stage (other than in adult), suggesting its hierarchy-and stage-restricted regulatory trait.