AbstractMost cell type-specific genes are regulated by the interaction of enhancers with their promoters. The identification of enhancers is not trivial as enhancers are diverse in their characteristics and dynamic in their interaction partners. We present Esearch3D, a new method that exploits network theory approaches to identify active enhancers. Our work is based on the fact that enhancers act as a source of regulatory information to increase the rate of transcription of their target genes and that the flow of this information is mediated by the folding of chromatin in the three-dimensional (3D) nuclear space between the enhancer and the target gene promoter. Esearch3D reverse engineers this flow of information to calculate the likelihood of enhancer activity in intergenic regions by propagating the transcription levels of genes across 3D genome networks. Regions predicted to have high enhancer activity are shown to be enriched in annotations indicative of enhancer activity. These include: enhancer-associated histone marks, bidirectional CAGE-seq, STARR-seq, P300, RNA polymerase II and expression quantitative trait loci (eQTLs). Esearch3D leverages the relationship between chromatin architecture and transcription, allowing the prediction of active enhancers and an understanding of the complex underpinnings of regulatory networks. The method is available at:https://github.com/InfOmics/Esearch3Dandhttps://doi.org/10.5281/zenodo.7737123.