CpG islands (CGIs) represent a widespread feature of vertebrate genomes, being associated with ~70% of all gene promoters. CGIs control transcription initiation by conferring nearby promoters with unique chromatin properties. In addition, there are thousands of distal or orphan CGIs (oCGIs) whose functional relevance is barely known. Here we show that oCGIs are an essential component of poised enhancers (PEs) that augment their long-range regulatory activity and control the responsiveness of their target genes. Using a knock-in strategy in mouse embryonic stem cells (ESCs), we introduced PEs with or without oCGIs within topologically associating domains (TADs) harboring genes with different types of promoters. Analysis of the resulting cell lines revealed that oCGIs act as tethering elements that promote the physical and functional communication between PEs and distally located genes, particularly those with large CGI clusters in their promoters. Therefore, by acting as genetic determinants of gene-enhancer compatibility, CGIs can contribute to gene expression control under both physiological and potentially pathological conditions.

In the last few years, the role of non-coding regulatory elements and their involvement in human disease have received great attention. Among the non-coding regulatory sequences, enhancers are particularly important for the proper establishment of cell type-specific gene-expression programs. Furthermore, the disruption of enhancers can lead to human disease through two main mechanisms: (i) Mutations or copy number variants can directly alter the enhancer sequences and thereby affect expression of their target genes; (ii) structural variants can provoke changes in 3-D chromatin organization that alter neither the enhancers nor their target genes, but rather the physical communication between them. In this review, these pathomechanisms are mostly discussed in the context of neurocristopathies, congenital disorders caused by defects that occur during neural crest development. We highlight why, due to its contribution to multiple tissues and organs, the neural crest represents an important, yet understudied, cell type involved in multiple congenital disorders. Moreover, we discuss currently available resources and experimental models for the study of human neurocristopathies. Last, we provide some practical guidelines that can be followed when investigating human neurocristopathies caused by structural variants. Importantly, these guidelines can be useful not only to uncover the etiology of human neurocristopathies, but also of other human congenital disorders in which enhancer disruption is involved.