A genome-wide association study identifies two loci associated with heart failure due to dilated cardiomyopathy

Abstract

Aims: Dilated cardiomyopathy (DCM) is a major cause of heart failure with a high familial recurrence risk. So far, the genetics of DCM remains largely unresolved. We conducted the first genome-wide association study (GWAS) to identify loci contributing to sporadic DCM.

Methods and results: One thousand one hundred and seventy-nine DCM patients and 1108 controls contributed to the discovery phase. Pools of DNA stratified on disease status, population, age, and gender were constituted and used for testing association of DCM with 517 382 single nucleotide polymorphisms (SNPs). Three DCM-associated SNPs were confirmed by individual genotyping (P < 5.0 10(-7)), and two of them, rs10927875 and rs2234962, were replicated in independent samples (1165 DCM patients and 1302 controls), with P-values of 0.002 and 0.009, respectively. rs10927875 maps to a region on chromosome 1p36.13 which encompasses several genes among which HSPB7 has been formerly suggested to be implicated in DCM. The second identified locus involves rs2234962, a non-synonymous SNP (c.T757C, p. C151R) located within the sequence of BAG3 on chromosome 10q26. To assess whether coding mutations of BAG3 might cause monogenic forms of the disease, we sequenced BAG3 exons in 168 independent index cases diagnosed with familial DCM and identified four truncating and two missense mutations. Each mutation was heterozygous, present in all genotyped relatives affected by the disease and absent in a control group of 347 healthy individuals, strongly suggesting that these mutations are causing the disease.

Conclusion: This GWAS identified two loci involved in sporadic DCM, one of them probably implicates BAG3. Our results show that rare mutations in BAG3 contribute to monogenic forms of the disease, while common variant(s) in the same gene are implicated in sporadic DCM.

Figure 1

Allelic odds ratios for associations with dilated cardiomyopathy of the three identified lead SNPs in the discovery and replication samples. Meta-analysis summary effects: rs2234962 (BAG3) OR: 0.66 (95% CI 0.53–0.82), rs10927875 (ZBTB17) OR: 0.76 (0.70–0.84), rs16983785 (TAK1L) OR: 1.53 (1.25–1.89). The values used to draw the Forest plots are derived from Supplementary material online, Table S2. Replication samples are prefixed by ‘REP.'. The black squares are centred at the corresponding odds ratio and their surface is proportional to the sample size, the 95% confidence intervals are shown. For convenience, in this representation, statistics are shown separately for men and women; however, the global statistics for the discovery and replication samples reported in the Results section are adjusted on population, gender, and age as indicated in Table 2. The association of rs2234962 with DCM was replicated in men (P = 0.0016) but not in women (P = 0.59). This difference is largely the consequence of the low minor allele frequency (MAF) of this SNP in women included in the German replication control sample (0.138 compared with >0.188 in the other groups, Supplementary material online, Table S2). This low frequency may be due to chance as the MAF of rs2234962 in German men and women was 0.214 and 0.218, respectively, in a large population-based sample of 3149 individuals from the Gutenberg Heart Study.

Figure 2

Association plot of SNPs with dilated cardiomyopathy (DCM) around the lead SNP rs10927875, based on pools–GWAS results. Upper panel: pairwise r² (derived from HAPMAP) between the lead SNP and the SNPs in its flanking regions are shown together with the association P-value with DCM of each SNP. Annotations of SNPs: none (circle), non-synonymous (triangle), synonymous or UTR (square), transcription factor binding site consensus (asterisk). Lower panel: genes encompassing the associated region. Linkage disequilibrium may explain the presence of several DCM-associated SNPs in the region (see also Supplementary material online, Table S3), especially in the sequences of HSPB7 (rs1763601, pools–GWAS: P < 1.0 × 10⁻⁴, this SNP being a perfect proxy of rs1739843, the DCM-associated SNP identified by Stark et al. according to HapMap release 22) and CLCNKA (rs1805152, pools–GWAS: P = 5.6 × 10⁻⁶).

Figure 3

Association plot of SNPs with dilated cardiomyopathy around the lead SNP rs2234962, based on Pools–GWAS results. See legend of Figure 2.

Figure 4

Associations with two non-synonymous SNPs in BAG3. Odds ratios for DCM associated with the diplotypes formed by the rs2234962 and rs3858340 SNPs. As these two SNPs are in complete linkage disequilibrium (D' = −1), they generate three haplotypes, C757-C1526 (R-P), T757-C1526 (C-P), and T757- T1526 (C-L), and six diplotypes that could be unambiguously determined in all individuals. In this plot, the most frequent diplotype (C-P/C-P) was used as reference.

Figure 5

Variants in BAG3 found in index patients with familial dilated cardiomyopathy (DCM). (A) Genomic structure of the BAG3 gene. The four exons are presented as boxes (white for UTR, grey for coding). Upper horizontal lines indicate sequenced regions. (B) BAG3 transcript with all missense and frame shift variants positions identified in familial DCM cases indicated. The variants are classified as likely disease causing (red), possibly disease causing (black), or probably neutral (green) as explained in the Results section. *Indicates SNPs associated with sporadic DCM in the GWAS. All likely and possibly disease-causing variants were found each in a single independent individual at the heterozygous state. The electrophoregrams representative of heterozygous mutated (upper) and homozygous wild-type (lower) sequences are shown for each DCM mutation. The arrows indicate the modified nucleotides and the sequenced strand orientation. (C) Schematic representation of the BAG3 protein with referenced domain signature according to UniprotKB database and dark grey boxes. The consequences of the DNA variants in (B) are shown as resultant predicted amino acid changes with the same colour code. (D) The ClustalW multiple alignments of orthologous BAG3 sequences from different species restricted to the immediate vicinity of each missense variant (red boxes) with MAF > 5% is shown. Interspecies conservation is indicated as blue boxes (dark blue: identical; light blue: similar; white: not conserved amino acid).