Copy Number Variation Analysis on a Non-Hodgkin Lymphoma Case-Control Study Identifies an 11q25 Duplication Associated with Diffuse Large B-Cell Lymphoma

Recent GWAS have identified several susceptibility loci for NHL. Despite these successes, much of the heritable variation in NHL risk remains to be explained. Common copy-number variants are important genomic sources of variability, and hence a potential source to explain part of this missing heritability. In this study, we carried out a CNV analysis using GWAS data from 681 NHL cases and 749 controls to explore the relationship between common structural variation and lymphoma susceptibility. Here we found a novel association with diffuse large B-cell lymphoma (DLBCL) risk involving a partial duplication of the C-terminus region of the LOC283177 long non-coding RNA that was further confirmed by quantitative PCR. For chronic lymphocytic leukemia/small lymphocytic lymphoma (CLL/SLL), known somatic deletions were identified on chromosomes 13q14, 11q22-23, 14q32 and 22q11.22. Our study shows that GWAS data can be used to identify germline CNVs associated with disease risk for DLBCL and somatic CNVs for CLL/SLL.     

Pooled sample-based GWAS: a cost-effective alternative for identifying colorectal and prostate cancer risk variants in the Polish population

Background

Prostate cancer (PCa) and colorectal cancer (CRC) are the most commonly diagnosed cancers and cancer-related causes of death in Poland. To date, numerous single nucleotide polymorphisms (SNPs) associated with susceptibility to both cancer types have been identified, but their effect on disease risk may differ among populations.

Methods

To identify new SNPs associated with PCa and CRC in the Polish population, a genome-wide association study (GWAS) was performed using DNA sample pools on Affymetrix Genome-Wide Human SNP 6.0 arrays. A total of 135 PCa patients and 270 healthy men (PCa sub-study) and 525 patients with adenoma (AD), 630 patients with CRC and 690 controls (AD/CRC sub-study) were included in the analysis. Allele frequency distributions were compared with t-tests and χ²-tests. Only those significantly associated SNPs with a proxy SNP (p<0.001; distance of 100 kb; r²>0.7) were selected. GWAS marker selection was conducted using PLINK. The study was replicated using extended cohorts of patients and controls. The association with previously reported PCa and CRC susceptibility variants was also examined. Individual patients were genotyped using TaqMan SNP Genotyping Assays.

Results

The GWAS selected six and 24 new candidate SNPs associated with PCa and CRC susceptibility, respectively. In the replication study, 17 of these associations were confirmed as significant in additive model of inheritance. Seven of them remained significant after correction for multiple hypothesis testing. Additionally, 17 previously reported risk variants have been identified, five of which remained significant after correction.

Conclusion

Pooled-DNA GWAS enabled the identification of new susceptibility loci for CRC in the Polish population. Previously reported CRC and PCa predisposition variants were also identified, validating the global nature of their associations. Further independent replication studies are required to confirm significance of the newly uncovered candidate susceptibility loci.     

Whole-Genome Sequencing of Individuals from a Founder Population Identifies Candidate Genes for Asthma

Asthma is a complex genetic disease caused by a combination of genetic and environmental risk factors. We sought to test classes of genetic variants largely missed by genome-wide association studies (GWAS), including copy number variants (CNVs) and low-frequency variants, by performing whole-genome sequencing (WGS) on 16 individuals from asthma-enriched and asthma-depleted families. The samples were obtained from an extended 13-generation Hutterite pedigree with reduced genetic heterogeneity due to a small founding gene pool and reduced environmental heterogeneity as a result of a communal lifestyle. We sequenced each individual to an average depth of 13-fold, generated a comprehensive catalog of genetic variants, and tested the most severe mutations for association with asthma. We identified and validated 1960 CNVs, 19 nonsense or splice-site single nucleotide variants (SNVs), and 18 insertions or deletions that were out of frame. As follow-up, we performed targeted sequencing of 16 genes in 837 cases and 540 controls of Puerto Rican ancestry and found that controls carry a significantly higher burden of mutations in IL27RA (2.0% of controls; 0.23% of cases; nominal p = 0.004; Bonferroni p = 0.21). We also genotyped 593 CNVs in 1199 Hutterite individuals. We identified a nominally significant association (p = 0.03; Odds ratio (OR) = 3.13) between a 6 kbp deletion in an intron of NEDD4L and increased risk of asthma. We genotyped this deletion in an additional 4787 non-Hutterite individuals (nominal p = 0.056; OR = 1.69). NEDD4L is expressed in bronchial epithelial cells, and conditional knockout of this gene in the lung in mice leads to severe inflammation and mucus accumulation. Our study represents one of the early instances of applying WGS to complex disease with a large environmental component and demonstrates how WGS can identify risk variants, including CNVs and low-frequency variants, largely untested in GWAS.     

Association of polymorphisms in the Chr18q11.2 locus with tuberculosis in Chinese population

A GWAS study has reported that two single nucleotide polymorphisms (SNPs) were associated with predisposition to tuberculosis (TB) in African populations. These two loci represented the long-waited GWAS hits for TB susceptibility. To determine whether these two SNPs are associated with TB in Chinese population, we attempted an replication in a cohort of over one thousand Chinese TB patients and 1,280 healthy controls using melting temperature shift allele-specific genotyping analysis. We found that only SNP rs4331426 was significantly associated with TB in Chinese population (p = 0.011). However, the effect was opposite. The G allele of the SNP in Chinese population is a protective allele (OR = 0.62, 95 % CI 0.44–0.87), while it was the risk allele for African population (OR = 1.19, 95 % CI 1.12–1.26). No significance was found for SNP rs2335704. The results provided an independent support for a role in susceptibility to TB for SNP rs4331426. However, it also indicated that direct predisposition element to TB and the association effects may vary across ethnic groups.     

Identification of Rare Recurrent Copy Number Variants in High-Risk Autism Families and Their Prevalence in a Large ASD Population

Structural variation is thought to play a major etiological role in the development of autism spectrum disorders (ASDs), and numerous studies documenting the relevance of copy number variants (CNVs) in ASD have been published since 2006. To determine if large ASD families harbor high-impact CNVs that may have broader impact in the general ASD population, we used the Affymetrix genome-wide human SNP array 6.0 to identify 153 putative autism-specific CNVs present in 55 individuals with ASD from 9 multiplex ASD pedigrees. To evaluate the actual prevalence of these CNVs as well as 185 CNVs reportedly associated with ASD from published studies many of which are insufficiently powered, we designed a custom Illumina array and used it to interrogate these CNVs in 3,000 ASD cases and 6,000 controls. Additional single nucleotide variants (SNVs) on the array identified 25 CNVs that we did not detect in our family studies at the standard SNP array resolution. After molecular validation, our results demonstrated that 15 CNVs identified in high-risk ASD families also were found in two or more ASD cases with odds ratios greater than 2.0, strengthening their support as ASD risk variants. In addition, of the 25 CNVs identified using SNV probes on our custom array, 9 also had odds ratios greater than 2.0, suggesting that these CNVs also are ASD risk variants. Eighteen of the validated CNVs have not been reported previously in individuals with ASD and three have only been observed once. Finally, we confirmed the association of 31 of 185 published ASD-associated CNVs in our dataset with odds ratios greater than 2.0, suggesting they may be of clinical relevance in the evaluation of children with ASDs. Taken together, these data provide strong support for the existence and application of high-impact CNVs in the clinical genetic evaluation of children with ASD.     

Identification of potential functional variants and genes at 18q21.1 associated with the carcinogenesis of colorectal cancer

Genome-wide association studies (GWAS) have identified more than 160 susceptibility loci for colorectal cancer (CRC). The effects of these variants, particularly their mechanisms, however, remain unclear. In this study, a comprehensive functional annotation of CRC-related GWAS signals was firstly conducted to identify the potential causal variants. We found that the SNP rs7229639 in intron 3 of SMAD7 at 18q21.1 might serve as a putative functional variant in CRC. The SNP rs7229639 is located in a region with evidence of regulatory potential. Dual-luciferase reporter assays revealed that three other SNPs (rs77544449, rs60385309 and rs72917785), in strong linkage disequilibrium (LD) with rs7229639, exhibited allele-specific enhancer activity, of which one of the target genes may conceivably be LIPG, as suggested by eQTL association data and Hi-C data. We also verified that LIPG promoted malignancy of CRC cells in vitro, with supporting clinical data indicating that LIPG is upregulated and correlated with a poor prognosis in CRC. Finally, pitavastatin was observed to exhibit an anti-CRC activity and modest inhibition of LIPG mRNA levels. Collectively, our data suggest that these functional variants at 18q21.1 are involved in the pathogenesis of CRC by modulating enhancer activity, and possibly LIPG expression, thus indicating a promising therapeutic target for CRC. The results of functional annotation in our investigation could also serve as an inventory for CRC susceptibility SNPs and offer guides for post-GWAS downstream functional studies.     

Genome-wide copy number variation study and gene expression analysis identify ABI3BP as a susceptibility gene for Kashin–Beck disease

Kashin–Beck disease (KBD) is a chronic osteochondropathy. In this study, we conducted the first genome-wide copy number variation study (GCNVS) of KBD totally involving 2,743 Chinese Han adults. GCNVS was first performed using Affymetrix Human SNP6.0 Arrays. The identified copy number variations (CNVs) were then replicated in an independent Chinese Han sample containing 1,026 subjects. SNP genotyping, CNV identification and quality control were implemented by Birdsuite. STRUCTURE and EIGENSTRAT were applied for controlling potential population stratification in the GCNVS. Association analysis was conducted using PLINK. Microarray and qRT-PCR were also conducted to compare the expression levels of the genes overlapping with identified CNVs between KBD patients and healthy controls. GCNVS found that CNV452 (P value = 7.78 × 10^?5) overlapping with ABI3BP gene was significantly associated with KBD. Replication association study observed that rs9850273 (P value = 0.008) and rs7613610 (P value = 0.021) in ABI3BP gene were significantly associated with KBD. Gene expression analysis also found that ABI3BP was up-regulated in KBD patients compared to healthy controls. Our results suggest that ABI3BP was a novel susceptibility gene for KBD.     

Identification of a Potential Regulatory Variant for Colorectal Cancer Risk Mapping to Chromosome 5q31.1: A Post-GWAS Study

Large-scale genome-wide association studies (GWAS) have established chromosome 5q31.1 as a susceptibility locus for colorectal cancer (CRC), which was still lack of causal genetic variants. We searched potentially regulatory single nucleotide polymorphisms (SNPs) in the overlap region between linkage disequilibrium (LD) block of 5q31.1 and regulatory elements predicted by histone modifications, then tested their association with CRC via a case-control study. Among three candidate common variants, we found rs17716310 conferred significantly (heterozygous model: OR = 1.273, 95% confidence interval (95%CI) = 1.016–1.595, P = 0.036) and marginally (dominant model: OR = 1.238, 95%CI = 1.000–1.532, P = 0.050) increase risk for CRC in a Chinese population including 695 cases and 709 controls. This variation was suggested to be regulatory altering the activity of enhancer that control PITX1 expression. Using epigenetic information such as chromatin immunoprecipitation-sequencing (ChIP-seq) data might help researchers to interpret the results of GWAS and locate causal variants for diseases in post-GWAS era.     

Rare variation at the TNFAIP3 locus and susceptibility to rheumatoid arthritis

Genome-wide association studies (GWAS) conducted using commercial single nucleotide polymorphisms (SNP) arrays have proven to be a powerful tool for the detection of common disease susceptibility variants. However, their utility for the detection of lower frequency variants is yet to be practically investigated. Here we describe the application of a rare variant collapsing method to a large genome-wide SNP dataset, the Wellcome Trust Case Control Consortium rheumatoid arthritis (RA) GWAS. We partitioned the data into gene-centric bins and collapsed genotypes of low frequency variants (defined here as MAF ≤0.05) into a single count coupled with univariate analysis. We then prioritised gene regions for further investigation in an independent cohort of 3,355 cases and 2,427 controls based on rare variant signal p value and prior evidence to support involvement in RA. A total of 14,536 gene bins were investigated in the primary analysis and signals mapping to the TNFAIP3 and chr17q24 loci were selected for further investigation. We detected replicating association to low frequency variants in the TNFAIP3 gene (combined p = 6.6 × 10^?6). Even though rare variants are not well-represented and can be difficult to genotype in GWAS, our study supports the application of low frequency variant collapsing methods to genome-wide SNP datasets as a means of exploiting data that are routinely ignored.     

Copy number variations identified in the chicken using a 60K SNP BeadChip

Copy number variation (CNV) is considered an important genetic variation, contributing to many economically important traits in the chicken. Although CNVs can be detected using a comparative genomic hybridization array, the high‐density SNP array has provided an alternative way to identify CNVs in the chicken. In the current study, a chicken 60K SNP BeadChip was used to identify CNVs in two distinct chicken genetic lines (White Leghorn and dwarf) using the penncnv program. A total of 209 CNV regions were identified, distributing on chromosomes 1–22 and 24–28 and encompassing 13.55 Mb (1.42%) of chicken autosomal genome area. Three of seven selected CNVs (73.2% individuals) were completely validated by quantitative PCR. To our knowledge, this is the first report in the chicken identifying CNVs using a SNP array. Identification of 190 new identified CNVs illustrates the feasibility of the chicken 60K SNP BeadChip to detect CNVs in the chicken, which lays a solid foundation for future analyses of associations of CNVs with economically important phenotypes in chickens.     

Multiple common susceptibility variants near BMP pathway loci GREM1, BMP4, and BMP2 explain part of the missing heritability of colorectal cancer

Genome-wide association studies (GWAS) have identified 14 tagging single nucleotide polymorphisms (tagSNPs) that are associated with the risk of colorectal cancer (CRC), and several of these tagSNPs are near bone morphogenetic protein (BMP) pathway loci. The penalty of multiple testing implicit in GWAS increases the attraction of complementary approaches for disease gene discovery, including candidate gene- or pathway-based analyses. The strongest candidate loci for additional predisposition SNPs are arguably those already known both to have functional relevance and to be involved in disease risk. To investigate this proposition, we searched for novel CRC susceptibility variants close to the BMP pathway genes GREM1 (15q13.3), BMP4 (14q22.2), and BMP2 (20p12.3) using sample sets totalling 24,910 CRC cases and 26,275 controls. We identified new, independent CRC predisposition SNPs close to BMP4 (rs1957636, P = 3.93×10(-10)) and BMP2 (rs4813802, P = 4.65×10(-11)). Near GREM1, we found using fine-mapping that the previously-identified association between tagSNP rs4779584 and CRC actually resulted from two independent signals represented by rs16969681 (P = 5.33×10(-8)) and rs11632715 (P = 2.30×10(-10)). As low-penetrance predisposition variants become harder to identify-owing to small effect sizes and/or low risk allele frequencies-approaches based on informed candidate gene selection may become increasingly attractive. Our data emphasise that genetic fine-mapping studies can deconvolute associations that have arisen owing to independent correlation of a tagSNP with more than one functional SNP, thus explaining some of the apparently missing heritability of common diseases.     

Genome-Wide Search for Gene-Gene Interactions in Colorectal Cancer

Genome-wide association studies (GWAS) have successfully identified a number of single-nucleotide polymorphisms (SNPs) associated with colorectal cancer (CRC) risk. However, these susceptibility loci known today explain only a small fraction of the genetic risk. Gene-gene interaction (GxG) is considered to be one source of the missing heritability. To address this, we performed a genome-wide search for pair-wise GxG associated with CRC risk using 8,380 cases and 10,558 controls in the discovery phase and 2,527 cases and 2,658 controls in the replication phase. We developed a simple, but powerful method for testing interaction, which we term the Average Risk Due to Interaction (ARDI). With this method, we conducted a genome-wide search to identify SNPs showing evidence for GxG with previously identified CRC susceptibility loci from 14 independent regions. We also conducted a genome-wide search for GxG using the marginal association screening and examining interaction among SNPs that pass the screening threshold (p<10⁻⁴). For the known locus rs10795668 (10p14), we found an interacting SNP rs367615 (5q21) with replication p = 0.01 and combined p = 4.19×10⁻⁸. Among the top marginal SNPs after LD pruning (n = 163), we identified an interaction between rs1571218 (20p12.3) and rs10879357 (12q21.1) (nominal combined p = 2.51×10⁻⁶; Bonferroni adjusted p = 0.03). Our study represents the first comprehensive search for GxG in CRC, and our results may provide new insight into the genetic etiology of CRC.     

Genome-wide Association Study in a High-Risk Isolate for Multiple Sclerosis Reveals Associated Variants in STAT3 Gene

Genetic risk for multiple sclerosis (MS) is thought to involve both common and rare risk alleles. Recent GWAS and subsequent meta-analysis have established the critical role of the HLA locus and identified new common variants associated to MS. These variants have small odds ratios (ORs) and explain only a fraction of the genetic risk. To expose potentially rare, high-impact alleles, we conducted a GWAS of 68 distantly related cases and 136 controls from a high-risk internal isolate of Finland with increased prevalence and familial occurrence of MS. The top 27 loci with p < 10⁻⁴ were tested in 711 cases and 1029 controls from Finland, and the top two findings were validated in 3859 cases and 9110 controls from more heterogeneous populations. SNP (rs744166) within the STAT3 gene was associated to MS (p = 2.75 × 10⁻¹⁰, OR 0.87, confidence interval 0.83–0.91). The protective haplotype for MS in STAT3 is a risk allele for Crohn disease, implying that STAT3 represents a shared risk locus for at least two autoimmune diseases. This study also demonstrates the potential of special isolated populations in search for variants contributing to complex traits.     

Replication Study in Chinese Population and Meta-Analysis Supports Association of the 11q23 Locus with Colorectal Cancer

Background

A common single nucleotide polymorphism (SNP), rs3802842, located at 11q23, was identified by genome-wide association studies (GWAS) to be significantly associated with the risk of colorectal cancer (CRC); however, the results of following replication studies were not always concordant. Thus, a case-control study and a meta-analysis were performed to clearly discern the effect of this variant in CRC.

Method and Findings

We determined the genotypes of rs3802842 in 641 unrelated Chinese patients with CRC and 1037 cancer-free controls. Additionally, a meta-analysis comprising current and previously published studies was conducted. In our case-control study, significant associations between the polymorphism and CRC risk were observed in all genetic models, with an additive OR being 1.45 (95% CI = 1.26–1.67). The meta-analysis of 38534 cases and 39446 controls further confirmed the significant associations in all genetic models but with obvious between-study heterogeneity. Nevertheless, ethnicity, study type and whether subjects affected by Lynch syndrome could synthetically accounted for the heterogeneity. Besides, the cumulative and sensitivity analyses indicated the robust stability of the results.

Conclusion

The results from our case-control study and meta-analysis provided convincing evidence that rs3802842 significantly contributed to CRC risk.     

Combination Testing Using a Single MSH5 Variant alongside HLA Haplotypes Improves the Sensitivity of Predicting Coeliac Disease Risk in the Polish Population

Assessment of non-HLA variants alongside standard HLA testing was previously shown to improve the identification of potential coeliac disease (CD) patients. We intended to identify new genetic variants associated with CD in the Polish population that would improve CD risk prediction when used alongside HLA haplotype analysis. DNA samples of 336 CD and 264 unrelated healthy controls were used to create DNA pools for a genome wide association study (GWAS). GWAS findings were validated with individual HLA tag single nucleotide polymorphism (SNP) typing of 473 patients and 714 healthy controls. Association analysis using four HLA-tagging SNPs showed that, as was found in other populations, positive predicting genotypes (HLA-DQ2.5/DQ2.5, HLA-DQ2.5/DQ2.2, and HLA-DQ2.5/DQ8) were found at higher frequencies in CD patients than in healthy control individuals in the Polish population. Both CD-associated SNPs discovered by GWAS were found in the CD susceptibility region, confirming the previously-determined association of the major histocompatibility (MHC) region with CD pathogenesis. The two most significant SNPs from the GWAS were rs9272346 (HLA-dependent; localized within 1 Kb of DQA1) and rs3130484 (HLA-independent; mapped to MSH5). Specificity of CD prediction using the four HLA-tagging SNPs achieved 92.9%, but sensitivity was only 45.5%. However, when a testing combination of the HLA-tagging SNPs and the MSH5 SNP was used, specificity decreased to 80%, and sensitivity increased to 74%. This study confirmed that improvement of CD risk prediction sensitivity could be achieved by including non-HLA SNPs alongside HLA SNPs in genetic testing.     

Rare Copy Number Variants Are a Common Cause of Short Stature

Human growth has an estimated heritability of about 80%–90%. Nevertheless, the underlying cause of shortness of stature remains unknown in the majority of individuals. Genome-wide association studies (GWAS) showed that both common single nucleotide polymorphisms and copy number variants (CNVs) contribute to height variation under a polygenic model, although explaining only a small fraction of overall genetic variability in the general population. Under the hypothesis that severe forms of growth retardation might also be caused by major gene effects, we searched for rare CNVs in 200 families, 92 sporadic and 108 familial, with idiopathic short stature compared to 820 control individuals. Although similar in number, patients had overall significantly larger CNVs (p-value<1×10⁻⁷). In a gene-based analysis of all non-polymorphic CNVs>50 kb for gene function, tissue expression, and murine knock-out phenotypes, we identified 10 duplications and 10 deletions ranging in size from 109 kb to 14 Mb, of which 7 were de novo (p<0.03) and 13 inherited from the likewise affected parent but absent in controls. Patients with these likely disease causing 20 CNVs were smaller than the remaining group (p<0.01). Eleven (55%) of these CNVs either overlapped with known microaberration syndromes associated with short stature or contained GWAS loci for height. Haploinsufficiency (HI) score and further expression profiling suggested dosage sensitivity of major growth-related genes at these loci. Overall 10% of patients carried a disease-causing CNV indicating that, like in neurodevelopmental disorders, rare CNVs are a frequent cause of severe growth retardation.     

Recurrent CNVs disrupt three candidate genes in schizophrenia patients

Schizophrenia is a severe psychiatric disease with complex etiology, affecting approximately 1% of the general population. Most genetics studies so far have focused on disease association with common genetic variation, such as single-nucleotide polymorphisms (SNPs), but it has recently become apparent that large-scale genomic copy-number variants (CNVs) are involved in disease development as well. To assess the role of rare CNVs in schizophrenia, we screened 54 patients with deficit schizophrenia using Affymetrix's GeneChip 250K SNP arrays. We identified 90 CNVs in total, 77 of which have been reported previously in unaffected control cohorts. Among the genes disrupted by the remaining rare CNVs are MYT1L, CTNND2, NRXN1, and ASTN2, genes that play an important role in neuronal functioning but--except for NRXN1--have not been associated with schizophrenia before. We studied the occurrence of CNVs at these four loci in an additional cohort of 752 patients and 706 normal controls from The Netherlands. We identified eight additional CNVs, of which the four that affect coding sequences were found only in the patient cohort. Our study supports a role for rare CNVs in schizophrenia susceptibility and identifies at least three candidate genes for this complex disorder.     

Genome-wide association study of aspirin-exacerbated respiratory disease in a Korean population

Aspirin-exacerbated respiratory disease (AERD) is a nonallergic clinical syndrome characterized by a severe decline in forced expiratory volume in one second (FEV1) following the ingestion of non-steroidal anti-inflammatory drugs (NSAIDs) such as aspirin. The effects of genetic variants have not fully explained all of the observed individual differences to an aspirin challenge despite previous attempts to identify AERD-related genes. In the present study, we performed genome-wide association study (GWAS) and targeted association study in Korean asthmatics to identify new genetic factors associated with AERD. A total of 685 asthmatic patients without AERD and 117 subjects with AERD were used for the GWAS of the first stage, and 996 asthmatics without AERD and 142 subjects with AERD were used for a follow-up study. A total of 702 SNPs were genotyped using the GoldenGate assay with the VeraCode microbead. GWAS revealed the top-ranked variants in 3′ regions of the HLA-DPB1 gene. To investigate the detailed genetic effects of an associated region with the risk of AERD, a follow-up targeted association study with the 702 single nucleotide polymorphisms (SNPs) of 14 genes was performed on 802 Korean subjects. In a case–control analysis, HLA-DPB1 rs1042151 (Met105Val) shows the most significant association with the susceptibility of AERD (p = 5.11 × 10^?7; OR = 2.40). Moreover, rs1042151 also shows a gene dose for the percent decline of FEV1 after an aspirin challenge (p = 2.82 × 10^?7). Our findings show that the HLA-DPB1 gene polymorphism may be the most susceptible genetic factor for the risk of AERD in Korean asthmatics and confirm the importance of HLA-DPB1 in the genetic etiology of AERD.     

Association of growth/differentiation factor 1 gene polymorphisms with the risk of congenital heart disease in the Chinese Han population

There is evidence suggesting that genetic variants of Nodal signaling may be associated with risk of congenital heart diseases (CHDs), in which several polymorphisms, such as Nodal rs1904589, have been considered to be implicated in the accumulation of the genetic burden of CHD risk with interacting genes. We hypothesized that genetic variants of GDF1, a protein that heterodimerizes with Nodal, may be related to increased CHD susceptibility. In this study, four tagSNPs of GDF1 were genotyped in 310 non-syndromic CHD patients and 320 healthy controls by using PCR-based DHPLC and RFLP. The results showed no statistically significant differences in genotype and allele frequencies between CHDs and controls with any of the analyzed variants of GDF1. However, a weak statistical association existed between GDF1 rs4808870 and conotruncal defects (CTDs) (uncorrected P = 0.027). Further stratified analysis for subtype revealed the SNP AA genotype and A allele have statistical significance in pulmonary atresia (PA) (corrected P = 1.01 × 10^?3 and 0.015, respectively), especially in pulmonary atresia with intact ventricular septum (PA + IVS) (corrected P = 1.67 × 10^?3 and 0.034, respectively). Furthermore, two haplotypes, TGGT and CAGT, were found to be significantly associated with increased CHD susceptibility (corrected P = 3.20 × 10^?3 and 2.73 × 10^?7, respectively). In summary, our results provide evidence that genetic variations of the Nodal-like factor, GDF1 may be associated with CHD risk, and these variations contribute at least in part to the development of some subtypes of CTD in the Chinese Han population.