Unidirectional expression of enhancers with cell type-dependent direction of transcription

Enhancers are genomic regulatory elements that can affect expression of genes over megabases of genomic distances. As previous evidence suggested that transcription initiation at enhancers occurs in a bidirectional pattern, balanced expression of divergently transcribed capped RNAs has been used as a hallmark for enhancer detection from transcriptome data. By analyzing deep Cap Analysis Gene Expression (CAGE) data from FANTOM5, FANTOM6, and other studies, we show that enhancers are usually unidirectionally transcribed in a given cell type. However, since the preferred direction of transcription of an enhancer may switch between cell types, enhancers may appear to be bidirectionally expressed when using CAGE data aggregated over many cell types. By analyzing expression directionality of enhancers across cell types, we classified enhancers into bidirectionally expressed, unidirectionally expressed with a consistent direction of transcription, and unidirectionally expressed with a switching direction, which was the largest category. We conclude that requiring bidirectional expression during enhancer prediction from transcriptome data may lead to false negatives. Also, the preference of a given cell type for a specific direction of transcription at an enhancer suggests that enhancer RNAs are not transcriptional noise, but may be functional.