In a recent study posted to the medRxiv* preprint server, a group of researchers identified large deletions in individuals with unexplained hyperinsulinism (HI), thereby elucidating the potential dysregulation of Forkhead box protein A2 (FOXA2) expression and its implications for syndromic HI.
Study: Chromosome 20p11.2 deletions cause congenital hyperinsulinism via the likely disruption of FOXA2. Image Credit: angellodeco/Shutterstock.com
*Important notice: medRxiv publishes preliminary scientific reports that are not peer-reviewed and, therefore, should not be regarded as conclusive, guide clinical practice/health-related behavior, or treated as established information.
Background
Deletions in large genomic deoxyribonucleic acid (DNA) regions can influence both rare and common diseases. For instance, removing portions or entire coding sequences of a single gene can lead to monogenic diseases.
An example is the deletion in ATP-binding cassette sub-family c member 8 (ABCC8) or hydroxyacyl-coenzyme a dehydrogenase (HADH) gene, causing congenital HI – a condition marked by inappropriate insulin secretion causing severe hypoglycaemia.
Large deletions affecting multiple genes lead to syndromic diseases, but sometimes, these deletions highlight critical genes or regions, as observed in the HK1 gene.
As deletions significantly affect HI's etiology and conventional tests only detect 45-79% of causative variants, more studies identifying key deletions in unsolved HI cases are necessary.
About the study
In the present study, researchers investigated 1,063 individuals undergoing routine genetic testing for HI. They collected clinical data using a standardized form and when a significant deletion was found, additional follow-up data was gathered.
Parents or guardians gave informed consent, and the North Wales Research Ethics Committee approved the study.
To exclude disease-causing variants in known HI genes, the team performed targeted next-generation sequencing. They analyzed the data to identify any partial or complete gene deletions and checked for specific known deletions related to HI.
For discovering new disease-causing deletions, the team examined whole genome sequencing data from 180 individuals.
They searched for large deletions and verified their findings against control samples and external datasets. If a deletion was found, they further refined its boundaries and checked its presence in parental samples.
To understand the mechanism of the disease, they examined the genomic sequencing data, looking for any recessive variants that could be revealed by the deletion.
They also looked for new variants in a subset of individuals with unresolved HI genetic conditions. The team also explored publicly available datasets related to pancreatic differentiation and gene expression.
The researchers further delved into various sequencing datasets to assess the potential impact of the deletion on FOXA2 gene regulation. They utilized multiple methods and datasets to evaluate the regulatory regions within the FOXA2 control area.
Study results
By analyzing 180 HI probands with genome sequencing data, researchers identified large overlapping deletions on chromosome 20p11.2 in three unrelated participants.
These were the sole overlapping deletions in three or more HI probands absent in in-house controls. Expanding this to a larger cohort using off-target sequencing, the team scrutinized 883 more individuals, discovering two more probands with overlapping deletions in this region.
Parental testing showed four of these deletions arose spontaneously (de novo), while one was inherited from an unaffected, heterozygous mother. The maternal grandparents' samples for this case were unobtainable.
The five detected deletions encompassed a non-imprinted region on chromosome 20p11.2. Notably, one patient exhibited a particularly complex variant involving sequential deletions and inversions.
The sizes of the deletions in these patients varied, but they shared a minimal overlap of roughly 2.4Mbp.
This specific region was absent in over 6,500 internal controls and more than 600,000 population controls from sources like the United Kingdom (UK) Biobank and genome aggregation database structural variants (gnomAD SV).
These five probands, diagnosed with HI between one day and 52 weeks of age, displayed varied medical histories. Three were treated with diazoxide, with one requiring near-total pancreatectomy due to poor response.
By the last update, the condition persisted in three of the patients, and all displayed additional symptoms, including developmental delay and facial dysmorphism.
Inside the shared 2.4Mb deletion, seven genes span the entire coding region. Interestingly, in two participants, the deletion covered the entire FOXA2 gene.
Analysis of genome sequencing from three probands aimed to determine if the deletions revealed a recessive coding variant, but none were found. Moreover, a search for de novo variants in 103 additional individuals did not identify a likely disease-causing gene or regulatory area.
In exploring the deletions' functional impact, researchers found that most of the genes within the deleted area were expressed in pancreatic beta-cells, with FOXA2 showing the highest expression levels.
Given that FOXA2 heterozygous loss-of-function variants can cause HI and that two participants had the complete FOXA2 gene deleted, further analysis into the gene's regulatory potential was conducted.
Data uncovered multiple chromatin-accessible regions in insulin-producing cells, suggesting the area plays a vital role in controlling FOXA2. This notion was bolstered by evidence of the region's activity varying across cell types and stages of differentiation.
Furthermore, a connection was identified between FOXA2 and NK2 Homeobox 2 (NKX2)-2 genes, hinting at a co-regulation mechanism that is disrupted in the minimal deleted area.
*Important notice: medRxiv publishes preliminary scientific reports that are not peer-reviewed and, therefore, should not be regarded as conclusive, guide clinical practice/health-related behavior, or treated as established information.
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