Search

Introduction: Recent studies have revealed that microRNAs (miRNAs, miRs) are important for self-renewal, differentiation, and cellular reprogramming of somatic cells into induced pluripotent stem cells (iPSC); however, their functional roles and target genes that are regulated by human PSC–specific miRs including hsa-miR-302 clusters remain largely unknown. Analysis of their target gene will give us the opportunity to understand the functional roles of such miRs. Methods: We analyzed the expression profiles of miRs in 4 somatic cell lines, 8 human iPSC lines derived from 4 different cell types, 3 human ESC lines, and embryoid bodies differentiated from the human ESCs to identify human PSC–specific miRs. We also analyzed the simultaneous expression profiles of miRs and mRNAs to identify candidate targets of human PSC–specific miRs. Then, we constructed a vector for overexpressing one of the target gene to dissect the functions of human PSC–specific miR in maintenance of self-renew and differentiation. Results: We focused on hsa-miR-302 cluster as a human PSC–specific miR and identified 22 candidate targets of hsa-miR-302 cluster that were moderately expressed in undifferentiated human PSCs and up-regulated in differentiated cells. Deleted in azoospermia-associated protein 2 (DAZAP2), one such target, was directly repressed by hsa-miR-302a, -302b, -302c and -302d, but not by hsa-miR-367. Overexpression of DAZAP2 caused a decrease in cell proliferation of undifferentiated human iPSCs, although morphology and undifferentiated marker gene expression was not affected. In addition, neural differentiation was suppressed in DAZAP2-overexpressing human iPSCs. Conclusion: Our study revealed that hsa-miR-302 cluster controls the cell proliferation of human PSCs and the neural differentiation of human PSCs by repression of DAZAP2, thereby highlighting an additional function of human PSC–specific miRs in maintaining pluripotency.

A bstract Runt-related transcription factor 1 (Runx1) is a master hematopoietic transcription factor essential for hematopoietic stem cell (HSC) emergence. Runx1-deficient mice die during early embryogenesis due to the inability to establish definitive hematopoiesis. Here, we have used human pluripotent stem cells (hPSCs) as model to study the role of RUNX1 in human embryonic hematopoiesis. Although the three RUNX1 isoforms a, b, and c were induced in CD45+ hematopoietic cells, RUNX1c was the only isoform induced in hematoendothelial progenitors (HEPs)/hemogenic endothelium. Constitutive expression of RUNX1c in human embryonic stem cells enhanced the appearance of HEPs, including hemogenic (CD43+) HEPs and promoted subsequent differentiation into blood cells. Conversely, specific deletion of RUNX1c dramatically reduced the generation of hematopoietic cells from HEPs, indicating that RUNX1c is a master regulator of human hematopoietic development. Gene expression profiling of HEPs revealed a RUNX1c-induced proinflammatory molecular signature, supporting previous studies demonstrating proinflammatory signaling as a regulator of HSC emergence. Collectively, RUNX1c orchestrates hematopoietic specification of hPSCs, possibly in cooperation with proinflammatory signaling. S tem C ells 2017;35:2253-2266 [ABSTRACT FROM AUTHOR]