Cell-specific alterations in Pitx1 regulatory landscape activation caused by the loss of a single enhancer.

Developmental genes are frequently controlled by multiple enhancers sharing similar specificities. As a result, deletions of such regulatory elements have often failed to reveal their full function. Here, we use the Pitx1 testbed locus to characterize in detail the regulatory and cellular identity alterations following the deletion of one of its enhancers (Pen). By combining single cell transcriptomics and an in-embryo cell tracing approach, we observe an increased fraction of Pitx1 non/low-expressing cells and a decreased fraction of Pitx1 high-expressing cells. We find that the over-representation of Pitx1 non/low-expressing cells originates from a failure of the Pitx1 locus to coordinate enhancer activities and 3D chromatin changes. This locus mis-activation induces a localized heterochrony and a concurrent loss of irregular connective tissue, eventually leading to a clubfoot phenotype. This data suggests that, in some cases, redundant enhancers may be used to locally enforce a robust activation of their host regulatory landscapes.

Chromatin topology in development and disease.

The discovery of domains of preferential interaction or Topologically Associating Domains (TADs) has provided a framework to understand the relation between enhancers and promoters within intricate regulatory landscapes. It has also enabled the conceptualization of the effect of non-coding structural variants on TADs structure and insulation and reveal new patho-mechanisms leading to disease. Here, we will review current knowledge on enhancer-promoter communication in relation to TAD structure. In particular, we will discuss how enhancer-promoter interaction dynamics is established within or outside of TADs. We will further provide an overview of how mutations affect the normal organization of the genome and how it impacts the normal ability of enhancers to induce transcription at their cognate promoters in disease. Finally, we will discuss the future directions to be explored to understand the mutual influences between 3D chromatin topology and gene regulation.