All SNPs are not created equal: genome-wide association studies reveal a consistent pattern of enrichment among functionally annotated SNPs

Abstract

Recent results indicate that genome-wide association studies (GWAS) have the potential to explain much of the heritability of common complex phenotypes, but methods are lacking to reliably identify the remaining associated single nucleotide polymorphisms (SNPs). We applied stratified False Discovery Rate (sFDR) methods to leverage genic enrichment in GWAS summary statistics data to uncover new loci likely to replicate in independent samples. Specifically, we use linkage disequilibrium-weighted annotations for each SNP in combination with nominal p-values to estimate the True Discovery Rate (TDR = 1-FDR) for strata determined by different genic categories. We show a consistent pattern of enrichment of polygenic effects in specific annotation categories across diverse phenotypes, with the greatest enrichment for SNPs tagging regulatory and coding genic elements, little enrichment in introns, and negative enrichment for intergenic SNPs. Stratified enrichment directly leads to increased TDR for a given p-value, mirrored by increased replication rates in independent samples. We show this in independent Crohn's disease GWAS, where we find a hundredfold variation in replication rate across genic categories. Applying a well-established sFDR methodology we demonstrate the utility of stratification for improving power of GWAS in complex phenotypes, with increased rejection rates from 20% in height to 300% in schizophrenia with traditional FDR and sFDR both fixed at 0.05. Our analyses demonstrate an inherent stratification among GWAS SNPs with important conceptual implications that can be leveraged by statistical methods to improve the discovery of loci.

Figure 1. Stratified Q-Q plot for height shows enrichment by annotation categories using Linkage-Disequilibrium (LD)-weighted scores.

Genic annotation categories were: 1) 10,000 to 1,001 base pairs upstream (10 k Up); 2) 1,000 to 1 base pair upstream (1 k Up); 3) 5′ untranslated region (5′UTR); 4) Exon; 5) Intron; 6) 3′ untranslated region (3′UTR); 7) 1 to 1,000 base pairs downstream (1 k Down); 8) 1,001 to 10,000 base pairs downstream (10 k Down). Q-Q plot of height with non-LD weighted category scores are shown in Figure S3.

Figure 2. Stratified Q-Q plots and true discovery rates show consistency of enrichment.

Upper panel: Stratified Q-Q plots illustrating consistent enrichment of genic annotation categories across diverse phenotypes: (A) Height, (B) Schizophrenia (SCZ), and (C) Cigarettes per Day (CPD). All figures are corrected for inflation using intergenic inflation control. Only nominal p-values below the standard genome-wide significance threshold (p<5×10⁻⁸) are shown. Lower panel: Stratified True Discovery Rate (TDR) plots illustrating the increase in TDR associated with increased enrichment in (D) Height, (E) SCZ and (F) CPD. Genic annotation categories were: 5′ untranslated region (5′UTR), Exon, Intron, 3′ untranslated region (3′UTR), All SNPs, in addition to Intergenic.

Figure 3. Categorical enrichment for seven diverse phenotypes.

The relative pattern of enrichment, as measured by the mean (z-score²−1) after intergenic inflation control, of LD-weighted genic annotation categories remain consistent. Results for all phenotypes are shown in Figure S5, Table S6.

Figure 4. Independent study replication confirms enrichment in Crohn's disease.

(A). Stratified True Discovery Rate (TDR) plots illustrating the increase in TDR associated with increased enrichment. (B) Cumulative replication plot showing the average rate of replication (p<.05) within sub-studies for a given p-value threshold shows enriched categories replicate at a higher rate in independent samples. The vertical intercept is the overall replication rate per category.

Figure 5. Enrichment improves discovery using established methods.

Among three phenotypes, (A) Height, (B) Crohn's Disease, (C) and Schizophrenia, we demonstrate an increased discovery of SNPs at a given FDR when incorporating the enriched genic annotation information into an established stratified false discovery rate (sFDR; red) framework. SNPs declared significant by sFDR also replicate at a higher rate (Figure S12).