Complete lung agenesis caused by complex genomic rearrangements with neo-TAD formation at the SHH locus

Abstract

During human organogenesis, lung development is a timely and tightly regulated developmental process under the control of a large number of signaling molecules. Understanding how genetic variants can disturb normal lung development causing different lung malformations is a major goal for dissecting molecular mechanisms during embryogenesis. Here, through exome sequencing (ES), array CGH, genome sequencing (GS) and Hi-C, we aimed at elucidating the molecular basis of bilateral isolated lung agenesis in three fetuses born to a non-consanguineous family. We detected a complex genomic rearrangement containing duplicated, triplicated and deleted fragments involving the SHH locus in fetuses presenting complete agenesis of both lungs and near-complete agenesis of the trachea, diagnosed by ultrasound screening and confirmed at autopsy following termination. The rearrangement did not include SHH itself, but several regulatory elements for lung development, such as MACS1, a major SHH lung enhancer, and the neighboring genes MNX1 and NOM1. The rearrangement incorporated parts of two topologically associating domains (TADs) including their boundaries. Hi-C of cells from one of the affected fetuses showed the formation of two novel TADs each containing SHH enhancers and the MNX1 and NOM1 genes. Hi-C together with GS indicate that the new 3D conformation is likely causative for this condition by an inappropriate activation of MNX1 included in the neo-TADs by MACS1 enhancer, further highlighting the importance of the 3D chromatin conformation in human disease.

Three‐dimensional genome structure and function

“…Duplication of DNA sequences containing boundary elements can result in the formation of “neo‐TADs”, 300 , 301 , 302 which are located between the original TADs. The enhancers and promoters within these “neo‐TADs” originate from different initial TADs, but their interactions do not interfere with the enhancer–promoter interactions in the original TADs (Figure 4B ).…”

HYENA detects oncogenes activated by distal enhancers in cancer

“…For this reason, one can reliably expect that upon chromosomal rearrangements, normal TADs can be disrupted, and new TADs can form by relocations of TAD boundaries. This assumption has been validated with direct experimental evidence from examining the "neo-TADs" associated with SVs at different loci [41][42][43] . The wildtype TOB1-AS1 locus had a TAD between a CTCF binding site in RSAD1 and another one upstream of SPAG9 (Fig.…”

A distant global control region is essential for normal expression of anterior HOXA genes during mouse and human craniofacial development

“…TAD organization has been shown to be important for gene regulation. Breaking or inverting TAD boundaries to affect their enhancer-gene content can result in developmental abnormalities (Lupiáñez et al, 2015;Melo et al, 2020Melo et al, , 2021Symmons et al, 2016). In an attempt to separate the function of the novel superenhancer region from larger scale TAD organization, we attempted to create a "scarless" edit with respect to TAD boundaries.…”