The evolution of Great Apes has shaped the functional enhancers' landscape in human embryonic stem cells.

High-throughput functional assays of enhancer activity have recently enabled the genome-scale definition of molecular, structural, and biochemical features of these genomic regulatory regions. To infer the evolutionary origin of DNA sequences operating as functional enhancers in human embryonic stem cells (hESC), we examined the patterns of evolutionary conservation and divergence in the genome-wide functional enhancers' landscape of hESC. We show that a prominent majority (up to 94%) of DNA sequences identified in hESC as functional enhancers are conserved in humans and our closest evolutionary relatives, Chimpanzee and Bonobo. More than 91% of functional enhancers that are highly conserved in both Chimpanzee and Bonobo, are conserved among other Great Apes and >75% are conserved in the Rhesus genome. In striking contrast, <5% of DNA sequences operating in hESC as functional enhancers are conserved in rodents. Conserved in primates enhancers' sequences are complemented by 1619 sequences of enhancers that are specific to humans. Enhancers that harbor human-specific sequences appear enriched among the invariant enhancer module maintaining activity in different pluripotent states and these regions are associated with pluripotency- and embryonic-lineage-related genes. However, functional enhancers make up only a minority of all conserved in primates or human-specific transcription factor binding sites. Our analyses revealed that sequences that are conserved during ~8 million years of primate evolution dominate the genomic landscape of functional enhancers in both primed and naïve hESC. Collectively, these observations revealed thousands of evolutionarily conserved sequences that function as a core regulatory network in human embryonic stem cells which has recently undergone further extension after divergence of modern humans from our closest relatives, Chimpanzee and Bonobo.