Genome-wide association study identifies novel loci associated with concentrations of four plasma phospholipid fatty acids in the de novo lipogenesis pathway: results from the Cohorts for Heart and Aging Research in Genomic Epidemiology (CHARGE) consortium

Abstract

BACKGROUND- Palmitic acid (16:0), stearic acid (18:0), palmitoleic acid (16:1n-7), and oleic acid (18:1n-9) are major saturated and monounsaturated fatty acids that affect cellular signaling and metabolic pathways. They are synthesized via de novo lipogenesis and are the main saturated and monounsaturated fatty acids in the diet. Levels of these fatty acids have been linked to diseases including type 2 diabetes mellitus and coronary heart disease. METHODS AND RESULTS- Genome-wide association studies were conducted in 5 population-based cohorts comprising 8961 participants of European ancestry to investigate the association of common genetic variation with plasma levels of these 4 fatty acids. We identified polymorphisms in 7 novel loci associated with circulating levels of ≥1 of these fatty acids. ALG14 (asparagine-linked glycosylation 14 homolog) polymorphisms were associated with higher 16:0 (P=2.7×10(-11)) and lower 18:0 (P=2.2×10(-18)). FADS1 and FADS2 (desaturases) polymorphisms were associated with higher 16:1n-7 (P=6.6×10(-13)) and 18:1n-9 (P=2.2×10(-32)) and lower 18:0 (P=1.3×10(-20)). LPGAT1 (lysophosphatidylglycerol acyltransferase) polymorphisms were associated with lower 18:0 (P=2.8×10(-9)). GCKR (glucokinase regulator; P=9.8×10(-10)) and HIF1AN (factor inhibiting hypoxia-inducible factor-1; P=5.7×10(-9)) polymorphisms were associated with higher 16:1n-7, whereas PKD2L1 (polycystic kidney disease 2-like 1; P=5.7×10(-15)) and a locus on chromosome 2 (not near known genes) were associated with lower 16:1n-7 (P=4.1×10(-8)). CONCLUSIONS- Our findings provide novel evidence that common variations in genes with diverse functions, including protein-glycosylation, polyunsaturated fatty acid metabolism, phospholipid modeling, and glucose- and oxygen-sensing pathways, are associated with circulating levels of 4 fatty acids in the de novo lipogenesis pathway. These results expand our knowledge of genetic factors relevant to de novo lipogenesis and fatty acid biology.

Figure 1

Major fatty acids in the de novo lipogenesis pathway. Palmitic acid (16:0), stearic acid (18:0), palmitoleic acid (16:1n-7), and oleic acid (18:1n-9) are the major saturated and mono-unsaturated fatty acids that are obtained via de novo lipogenesis (DNL) or the diet, and investigated in this study (the metabolic steps which link the fatty acids are in italics, and the key enzyme involved in these processes labeled in brackets). In the initial steps of DNL, fatty acid synthase (FAS) catalyze the polymerization of malonyl-CoA to form 16:0 as the major initial product. 16:0 can be elongated to 18:0, and cellular studies suggested elongase-6 (Elov6) is the enzyme primarily responsible for this conversion . Both 16:0 and 18:0 are substrates for stearoyl-CoA desaturase (SCD) to give rise to 16:1n-7 and 18:1n-9, respectively .

Figure 2

Meta-analysis of genome-wide associations with fatty acids in the de novo lipogenesis pathway. A: palmitic acid (16:0); B: stearic acid (18:0); C: palmitoleic acid (16:1n-7), and D: oleic acid (18:1n-9). Associations are demonstrated by chromosome location and −log₁₀ (P-value), up to P-values of 10^-10. Triangles indicate additional SNPs with P-value < 10^-10. Genes of interest in each locus with SNPs variants which reached genome-wide significance are shown.

Figure 3

SNP association plots for palmitic acid-associated region. Genetic coordinates are along the x-axis (as per NCI build 36) and genome-wide association significance level is plotted against the y-axis as −log₁₀(P-value). LD is indicated by color scale in relationship to marker rs2391388, with red for strong linkage disequilibrium (LD; r² ≥0.8) and fading color for lower LD.

Figure 4

SNP association plots for stearic acid-associated region. Genetic coordinates are along the x-axis (as per NCI build 36) and genome-wide association significance level is plotted against the y-axis as −log₁₀(P-value). (A) ALG14 cluster region. LD is indicated by color scale in relationship to marker rs6675668. (B) LPGAT1 cluster region. LD is indicated by color scale in relationship to marker rs11119805. (C) FADS cluster region. LD is indicated by color scale in relationship to marker rs102275. The color scheme is red for strong linkage disequilibrium (LD; r² ≥0.8) and fading color for lower LD.

Figure 5

SNP association plots for oleic acid-associated region. Genetic coordinates are along the x-axis (as per NCI build 36) and genome-wide association significance level is plotted against the y-axis as −log₁₀(P-value). LD is indicated by color scale in relationship to marker rs102275, with red for strong linkage disequilibrium (LD; r² ≥0.8) and fading color for lower LD.

Figure 6

SNP association plots for palmitoleic acid-associated region. Genetic coordinates are along the x-axis (as per NCI build 36) and genome-wide association significance level is plotted against the y-axis as −log₁₀(P-value). (A) FADS cluster region. LD is indicated by color scale in relationship to marker rs102275. (B) PKD2L1 cluster region. LD is indicated by color scale in relationship to marker rs603424. (C) HIF1AN cluster region. LD is indicated by color scale in relationship to marker rs11190604. (D) GCKR cluster region. LD is indicated by color scale in relationship to marker rs780093. The color scheme is red for strong linkage disequilibrium (LD; r² ≥0.8) and fading color for lower LD.

Figure 7

Forest plots for each of the top SNP-fatty acid associations: Within cohort effect size and 95% CI's were obtained from linear regression analysis using robust standard errors, and results were pooled using inverse-variance weighted meta-analysis. The size of the grey box around the central effect size estimate of each study is proportional to its inverse-variance weight in the meta-analysis. The vertical dashed line indicates the pooled meta-analysis effect size estimate. Chi-square test for heterogeneity P-values, and the I² statistic are also shown for each meta-analysis. The magnitude and direction of associations were generally consistent across all 5 cohorts. In cases where moderate heterogeneity were present, this was largely due to different findings in the InCHIANTI cohort. We note that all other cohorts assessed fatty acids in plasma phospholipids, whereas fatty acids were measured in total plasma in InCHIANTI; this difference could, at least in part, account for some of this heterogeneity. It is also important to note that this heterogeneity was due to differences in the magnitudes of the gene-fatty acid associations in InCHIANTI versus the other cohorts, rather than differing directions of associations; and that exclusion of InCHIANTI from each meta-analysis did not materially alter the top SNP-fatty acid associations (results not shown)

Figure 7

Forest plots for each of the top SNP-fatty acid associations: Within cohort effect size and 95% CI's were obtained from linear regression analysis using robust standard errors, and results were pooled using inverse-variance weighted meta-analysis. The size of the grey box around the central effect size estimate of each study is proportional to its inverse-variance weight in the meta-analysis. The vertical dashed line indicates the pooled meta-analysis effect size estimate. Chi-square test for heterogeneity P-values, and the I² statistic are also shown for each meta-analysis. The magnitude and direction of associations were generally consistent across all 5 cohorts. In cases where moderate heterogeneity were present, this was largely due to different findings in the InCHIANTI cohort. We note that all other cohorts assessed fatty acids in plasma phospholipids, whereas fatty acids were measured in total plasma in InCHIANTI; this difference could, at least in part, account for some of this heterogeneity. It is also important to note that this heterogeneity was due to differences in the magnitudes of the gene-fatty acid associations in InCHIANTI versus the other cohorts, rather than differing directions of associations; and that exclusion of InCHIANTI from each meta-analysis did not materially alter the top SNP-fatty acid associations (results not shown)

Figure 7

Forest plots for each of the top SNP-fatty acid associations: Within cohort effect size and 95% CI's were obtained from linear regression analysis using robust standard errors, and results were pooled using inverse-variance weighted meta-analysis. The size of the grey box around the central effect size estimate of each study is proportional to its inverse-variance weight in the meta-analysis. The vertical dashed line indicates the pooled meta-analysis effect size estimate. Chi-square test for heterogeneity P-values, and the I² statistic are also shown for each meta-analysis. The magnitude and direction of associations were generally consistent across all 5 cohorts. In cases where moderate heterogeneity were present, this was largely due to different findings in the InCHIANTI cohort. We note that all other cohorts assessed fatty acids in plasma phospholipids, whereas fatty acids were measured in total plasma in InCHIANTI; this difference could, at least in part, account for some of this heterogeneity. It is also important to note that this heterogeneity was due to differences in the magnitudes of the gene-fatty acid associations in InCHIANTI versus the other cohorts, rather than differing directions of associations; and that exclusion of InCHIANTI from each meta-analysis did not materially alter the top SNP-fatty acid associations (results not shown)

Figure 8

Summary of genome-wide association results. The genome-wide significant associations of identified loci (and genes of potential interest) with each fatty acid are shown with dashed arrows, and the +/- signs indicate the direction of the associations for the minor alleles at each loci.