Novel Loci for Non-Syndromic Coarctation of the Aorta in Sporadic and Familial Cases

<div><p>Backround</p><p>Coarctation of the aorta (CoA) accounts for 5-8% of all congenital heart defects. CoA can be detected in up to 20% of patients with Ullrich-Turner syndrome (UTS), in which a part or all of one of the X chromosomes is absent. The etiology of non-syndromic CoA is poorly understood. In the present work, we test the hypothesis that rare copy number variation (CNV) especially on the gonosomes, contribute to the etiology of non-syndromic CoA.</p><p>Methods</p><p>We performed high-resolution genome-wide CNV analysis using the Affymetrix SNP 6.0 microarray platform for 70 individuals with sporadic CoA, 3 families with inherited CoA (n=13) and 605 controls. Our analysis comprised genome wide association, CNV burden and linkage. CNV was validated by multiplex ligation-dependent probe amplification.</p><p>Results</p><p>We identified a significant abundance of large (>100 kb) CNVs on the X chromosome in males with CoA (p=0.005). 11 out of 51 (~ 22%) male cases had these large CNVs. Association analysis in the sporadic cohort revealed 14 novel loci for CoA. The locus on 21q22.3 in the sporadic CoA cohort overlapped with a gene locus identified in all familial cases of CoA (candidate gene <i>TRPM2</i>). We identified one CNV locus within a locus with high multipoint LOD score from a linkage analysis of the familial cases (<i>SEPT9</i>); another locus overlapped with a region implicated in Kabuki syndrome. In the familial cases, we identified a total of 7 CNV loci that were exclusively present in cases but not in unaffected family members.</p><p>Conclusion</p><p>Of all candidate loci identified, the TRPM2 locus was the most frequently implicated autosomal locus in sporadic and familial cases. However, the abundance of large CNVs on the X chromosome of affected males suggests that gonosomal aberrations are not only responsible for syndromic CoA but also involved in the development of sporadic and non-syndromic CoA and their male dominance.</p></div

CNV Loci in the sporadic CoA cohort.

<p>14 loci exceeded the significance threshold of 5 E-05, including 5 deletions and 9 duplications. The first locus on chromosome 17 did not quite pass the significance threshold, but was retained due to overlap with a linkage region.</p><p>CNV Loci in the sporadic CoA cohort.</p

CNV burden analysis of large (>100kb) CNVs on gonosomes and autosomes in familial cases versus healthy individuals.

<p>CNV burden in familial CoA was significantly higher for deletions >100kb (p<0.01) and significantly lower for duplications > 100 kb (p<0.01). The total number of CNVs were higher in controls. No significant difference was observed between males and females. Fisher ‘s exact test was calculated from a contingency table with the total number of CNV segments, with a size between 10 and 100 kb and those larger than 100 kb in both cases and controls.</p><p>CNV burden analysis of large (>100kb) CNVs on gonosomes and autosomes in familial cases versus healthy individuals.</p

Overlapping CNV segments in familial CoA with candidate gene.

<p>Seven overlapping CNVs shared in >4 individuals with CoA were identified. The chromosome 21 CNV locus was present in all familial CoA cases. It contains <i>TRPM2</i>, which was also identified in the sporadic CoA cohort.</p><p>Overlapping CNV segments in familial CoA with candidate gene.</p

CNV burden analysis of large (>100 kb) CNVs on gonosomes and autosomes in the sporadic CoA cohort.

<p>Total number of CNVs on gonosomes and autosomes. CNV burden was significantly higher for large (>100kb) CNVs in male patients (p<0.01) on the X-chromosome. No changes were seen for the X-chromosome in females and on the autosomes. A one-sided Fisher‘s exact test was calculated from a contingency table with the total number of CNV segments, with a size between 10 and 100 kb and those larger than 100 kb in both cases and controls.</p><p>CNV burden analysis of large (>100 kb) CNVs on gonosomes and autosomes in the sporadic CoA cohort.</p

Additional file 2 of Integrative bioinformatics analysis characterizing the role of EDC3 in mRNA decay and its association to intellectual disability

Figure S1 Validation of SKNBE transcriptome data via qPCR. Comparison between fold changes obtained with RNA sequencing and with real time qPCR of selected genes. Direction of fold change was confirmed for 9 out of 10 assayed genes. (PDF 111 kb

LOD-Plot showing genome wide linkage analysis in all three families.

<p>19 linkage regions with a maximum LOD-score of 1.62 were identified, which are illustrated by multipoint LOD scores above background.</p

Overview of workflow of CNV analysis in sporadic CoA patients and families with CoA.

<p>Genotyping was performed in 70 individuals with sporadic CoA, 605 healthy controls, and 3 families with 6 CoA cases and 7 healthy individuals. CNV calling was performed using Birdseye and Birdseed software. CNV burden analysis in sporadic cohort and familial CoA samples was performed for autosomes and gonosomes separately. Association analysis was performed in the sporadic CoA cohort and revealed 14 CNV loci. Overlap of CNVs in familial patients was assessed and revealed 7 CNVs shared in >4 patients, overlapping with one CNV locus in the sporadic CoA cohort. Linkage analysis in familial samples revealed 19 linked regions, one overlapping with the CNV locus on chromosome 17, which did not pass the threshold of significance in the association analysis.</p

Match between linkage loci and genes of cardiac gene list.

<p>Of the significant 19 linkage loci of familial cases, 9 loci contained genes (n = 22) of the cardiac gene list.</p><p>Match between linkage loci and genes of cardiac gene list.</p

Distribution of CNV on 8p23.1 in all affected patients according to their phenotype.

<p>The 20 patients with CNV on 8p23.1 represent a typical distribution of concomitant malformation in CoA. 10 patients with isolated CoA, 7 with additional BAV and 3 with additional VSD.</p><p>Distribution of CNV on 8p23.1 in all affected patients according to their phenotype.</p