Enhancers are key regulatory elements that control the spatial and temporal gene expression, which is vital for cell differentiation and disease occurrence. It has been an important research focus to precisely identify DNA sequences that execute enhancer functions in the genome.

In the post-genomics era, with the assistance of next-generation sequencing, the researchers began to utilize the numerous signatures of chromatin to annotate important elements in the genome. Among these signatures, H3K4me1, H3K27ac, highly enriched histone variant H3.3, and a high histone turnover rate are remarkable markers for active enhancers. However, whether H3K27ac, the best indicator of active enhancers, could functionally determinate enhancer activity is less clear.

In a study published online in Genome Biology, the researchers from Institute of Biophysics of the Chinese Academy of Sciences reported that selective erasure of H3K27ac at enhancers do not impact active transcription, and enhancer activity does not solely depend on H3K27ac, instead it is a functional consequence of the synergistic action by multiple acetylation events at different histone sites.

To specifically target enhancer H3K27ac, the researchers mutated histone variant lysine 27 in H3.3 to arginine (H3.3K27R) and produced non-acetyl-modifiable H3.3 histones in mouse ESCs. H3K27ac is almost completely depleted at active enhancers in H3.3K27R mutant ESCs. Meanwhile, the transcriptome change is minimal.

They then applied the newly developed model, activity-by-contact, to define target genes of H3K27ac-marked active enhancers, and found that the transcription of these genes is not sensitive to the dramatic decrease in H3K27ac at their enhancers. Transcription of genes associated with super enhancers, which harbor enormous amounts of H3K27ac, is also mostly unchanged.

Besides, the researchers profiled the distributions of acetylation at other histone lysine residues, including H3K9, H3K18, H3K122, H4K5, and H4K8. They found that acetylations at these sites are remained genome-widely. Acetylation can neutralize the positive charge on lysine residues, leading to attenuated interaction between histone tails and DNA. Therefore, it is the synergistic effect of multiple acetylation events that crucial for chromatin opening and maintaining enhancer activity.

The results showed that although H3K27ac is, and remains to be the best indicator of active enhancers in the mammalian genome, its depletion at enhancer regions does not affect chromatin accessibility and gene transcription. This study suggested that H3K27ac has to work in concert with acetylation events on other histone lysine residues.