Impact of the overexpression of the tyrosine kinase receptor RET in the hematopoietic potential of induced pluripotent stem cells (iPSCs)

Abstract

Previous studies have suggested that the tyrosine kinase receptor RET plays a significant role in the hematopoietic potential in mice and could also be used to expand cord-blood derived hematopoietic stem cells (HSCs). The role of RET in human iPSC-derived hematopoiesis has not been tested so far. To test the implication of RET on the hematopoietic potential of iPSCs, we activated its pathway with the lentiviral overexpression of RETWT or RETC634Y mutation in normal iPSCs. An iPSC derived from a patient harboring the RETC634Y mutation (iRETC634Y) and its CRISPR-corrected isogenic control iPSC (iRETCTRL) were also used. The hematopoietic potential was tested using 2D cultures and evaluated regarding the phenotype and the clonogenic potential of generated cells. Hematopoietic differentiation from iPSCs with RET overexpression (WT or C634Y) led to a significant reduction in the number and in the clonogenic potential of primitive hematopoietic cells (CD34+/CD38-/CD49f+) as compared to control iPSCs. Similarly, the hematopoietic potential of iRETC634Y was reduced as compared to iRETCTRL. Transcriptomic analyses revealed a specific activated expression profile for iRETC634Y compared to its control with evidence of overexpression of genes which are part of the MAPK network with negative hematopoietic regulator activities. RET activation in iPSCs is associated with an inhibitory activity in iPSC-derived hematopoiesis, potentially related to MAPK activation.