Noncoding copy-number variations are associated with congenital limb malformation

Abstract

PurposeCopy-number variants (CNVs) are generally interpreted by linking the effects of gene dosage with phenotypes. The clinical interpretation of noncoding CNVs remains challenging. We investigated the percentage of disease-associated CNVs in patients with congenital limb malformations that affect noncoding cis-regulatory sequences versus genes sensitive to gene dosage effects.MethodsWe applied high-resolution copy-number analysis to 340 unrelated individuals with isolated limb malformation. To investigate novel candidate CNVs, we re-engineered human CNVs in mice using clustered regularly interspaced short palindromic repeats (CRISPR)-based genome editing.ResultsOf the individuals studied, 10% harbored CNVs segregating with the phenotype in the affected families. We identified 31 CNVs previously associated with congenital limb malformations and four novel candidate CNVs. Most of the disease-associated CNVs (57%) affected the noncoding cis-regulatory genome, while only 43% included a known disease gene and were likely to result from gene dosage effects. In transgenic mice harboring four novel candidate CNVs, we observed altered gene expression in all cases, indicating that the CNVs had a regulatory effect either by changing the enhancer dosage or altering the topological associating domain architecture of the genome.ConclusionOur findings suggest that CNVs affecting noncoding regulatory elements are a major cause of congenital limb malformations.

Multiplex targeted high‐throughput sequencing in a series of 352 patients with congenital limb malformations

“…With bioinformatic algorithms, CNV detection with NGS is easy and can replace traditional methods such as quantitative PCR or MLPA (Yao, Yu, Qing, Wang, & Shen, ). The importance of CNV detection in CLM has been confirmed by numerous studies (Carter et al, ; Flottmann et al, ). Our strategy allows the detection of CNVs in all 52 targeted regions, when formerly CNV determination was possible in only a very limited number of genes.…”

“…In most cases, the responsible genetic anomaly is a single nucleotide variation (SNV) (Manouvrier‐Hanu et al, ). However, copy‐number variations (CNVs) involving genes or noncoding regulatory elements are also a major cause of CLM (Flottmann et al, ).…”

Chromatin features constrain structural variation across evolutionary timescales

“…One likely mechanism is 'enhancer hijacking' (Beroukhim et al, 2016;Northcott et al, 2014), also previously termed 'enhancer adoption' (Lettice et al, 2011), whereby a structural variant removes or moves a TAD boundary to expose TSSs to regulatory enhancers from which they would normally be insulated. While there have been intriguing examples of TAD boundary disruptions in developmental diseases (Franke et al, 2016;Kraft et al, 2015;Lupiáñez et al, 2015;Symmons et al, 2016), the effect of structural variants on chromatin features like TAD boundaries has received relatively little systematic attention outside of cancer (Ibn-Salem et al, 2014), until the past year (Flöttmann et al, 2017;Huynh and Hormozdiari, 2018;Krefting et al, 2017;Lazar et al, 2017;Zepeda-Mendoza et al, 2017).…”

Disruption of the PTHLH regulatory landscape results in features consistent with hyperparathyroid disease

“…Deletions resulting in changes in chromatin architecture that drive genetic disease have been previously demonstrated (Flöttmann et al, ; Lupiáñez et al, ). Deletions found in patients with limb anomalies were found to result in ectopic interactions between gene promoters and noncoding DNA.…”