AbstractThe first cell fate commitment during mammalian development is the specification of the inner cell mass and trophectoderm. This irreversible cell fate commitment should be epigenetically regulated, but the precise mechanism is largely unknown in humans. Here, we show that naïve human embryonic stem (hES) cells can transdifferentiate into trophoblast stem (hTS) cells, but primed hES cells cannot. Our transcriptome and methylome analyses reveal that a primate-specific miRNA cluster on chromosome 19 (C19MC) is active in naïve hES cells but epigenetically silenced in primed ones. Moreover, genome and epigenome editing using CRISPR/Cas systems demonstrate that C19MC is essential for hTS cell maintenance and C19MC-reactivated primed hES cells can give rise to hTS cells. Thus, we reveal that C19MC activation confers differentiation potential into trophoblast lineages on hES cells. Our findings are fundamental to understanding the epigenetic regulation of human early development and pluripotency.