The hsa-miR-302 cluster controls ectodermal differentiation of human pluripotent stem cell via repression of DAZAP2

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.

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.

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.

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.