Contribution of global rare copy-number variants to the risk of sporadic congenital heart disease

Abstract

Previous studies have shown that copy-number variants (CNVs) contribute to the risk of complex developmental phenotypes. However, the contribution of global CNV burden to the risk of sporadic congenital heart disease (CHD) remains incompletely defined. We generated genome-wide CNV data by using Illumina 660W-Quad SNP arrays in 2,256 individuals with CHD, 283 trio CHD-affected families, and 1,538 controls. We found association of rare genic deletions with CHD risk (odds ratio [OR] = 1.8, p = 0.0008). Rare deletions in study participants with CHD had higher gene content (p = 0.001) with higher haploinsufficiency scores (p = 0.03) than they did in controls, and they were enriched with Wnt-signaling genes (p = 1 × 10(-5)). Recurrent 15q11.2 deletions were associated with CHD risk (OR = 8.2, p = 0.02). Rare de novo CNVs were observed in ~5% of CHD trios; 10 out of 11 occurred on the paternally transmitted chromosome (p = 0.01). Some of the rare de novo CNVs spanned genes known to be involved in heart development (e.g., HAND2 and GJA5). Rare genic deletions contribute ~4% of the population-attributable risk of sporadic CHD. Second to previously described CNVs at 1q21.1, deletions at 15q11.2 and those implicating Wnt signaling are the most significant contributors to the risk of sporadic CHD. Rare de novo CNVs identified in CHD trios exhibit paternal origin bias.

Figure 1

Recurrent Rare Deletions in 15q11.2 Twelve deletions (shown as red bars in the UCSC Genome Browser) were identified in three individuals with complex left-sided malformations (L-sided), three with coarctation of the aorta (CoA), two with ventricular septal defects (VSDs), two with atrial septal defects (ASDs), one with total anomalous pulmonary venous drainage (TAPVD), and one with TOF. RefSeq genes, segmental duplications, and coverage of the Illumina 660W platform in the region are shown. The smallest deletions encompass four RefSeq genes: TUBGCP5, CYFIP1, NIPA2, and NIPA1.

Figure 2

Recurrent Rare CNVs in the Candidate 8p23.1 Locus (A and B) Four deletions were identified in one individual with an atrioventricular septal defect (AVSD), one with a ventricular septal defect (VSD), and two with TOF (all shown as red bars). The smallest deletion encompasses the last five exons of GATA4 and the whole coding regions of NEIL2 and FDFT1 (shown in B). In addition, an overlapping duplication was identified in one participant with a bicuspid aortic valve with aortic regurgitation (AR) (blue bar). The parental samples of these people are not available for analysis.

Figure 3

Rare CNVs Overlapping Rare De Novo CNVs Identified in TOF Trios We examined the remaining 1,978 CHD cases for recurrent CNVs that overlap rare de novo findings in 283 TOF trios. We found overlaps in 4q34 (a rare deletion in the highly conserved region upstream of HAND2, as shown in A), 5q14.2 (one rare deletion overlapping EDIL3 and two others overlapping a conserved region ∼100 kb upstream of EDIL3, as shown in B), and 5q35 (an deletion overlapping CNOT6, as shown in C). Deletions and duplications are shown in the UCSC Genome Browser as red and blue bars, respectively. The following abbreviations are used: PS, pulmonary stenosis; and VSD, ventricular septal defect.

Figure 4

Rare CNVs Overlapping CNVs Reported by Greenway et al., 2009 We searched in 1,987 CHD cases for CNVs overlapping the rare de novo CNVs previously reported by Greenway et al. We found recurrent rare deletions in the 7p21.3 locus (A) in two TOF probands, both of whom inherited the deletions from their respective unaffected fathers. Both of these findings had been confirmed on the Affymetrix 6.0 platform and by MLPA. We did not find such variants in 841 unrelated controls or other unaffected family members (n = 695). These rare CNVs did not overlap any known RefSeq genes, although there are some overlaps with transcription-factor binding-site conservation (shown). The nearest gene is NXPH1. (B) On the Illumina 660W platform (shown), there is insufficient coverage overlapping the 4q22.1 de novo variant reported by Greenway et al. Therefore, in addition to examining this locus in the 1,987 CHD probands who had been typed on the Illumina 660W, we also screened this locus with MLPA in 1,007 CHD cases, 866 of whom were also typed on the Illumina 660W. In a TOF proband, we detected a duplication that encompassed PPM1K (as shown). No overlapping duplication was found in 841 unrelated controls or 697 unaffected family members. The parental DNA samples of this proband were not available for analysis. Deletions and duplications are shown in the UCSC Genome Browser as red and blue bars, respectively.