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Anand Swaroop, Freekje Van Asten, Madeline Kwiklis, Kayode Sosina, Margaret Starostik, Nivedita Singh, Dena Hernandez, Emily Y Chew, Andrew Singleton, Alexis Battle, Deb A Ferrington, Nilanjan Chatterjee, Rinki Ratnapriya; Beyond GWAS: Functional genomics of age-related macular degeneration. Invest. Ophthalmol. Vis. Sci. 2019;60(9):4246. doi: https://doi.org/.
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© ARVO (1962-2015); The Authors (2016-present)
Genome-wide association studies (GWAS) have revealed 52 independent variants at 34 loci associated with age-related macular degeneration (AMD). However, we have little insight into their causal mechanisms as a majority of signals reside in the non-coding genome. We hypothesize that associated variants impact AMD progression through their effect on context- or tissue-specific gene expression.
Donor eyes were evaluated for AMD using the Minnesota Grading System. RNA and DNA from 128 controls, 197 early, 126 intermediate and 66 advanced AMD retina were used to generate RNA-Seq and for genotyping. eQTL analysis was performed using QTLtools. AMD-GWAS was integrated with eQTLs using eCAVIAR and TWAS. DNA methylation patterns of human retina samples were used to discover methylation-QTLs using QTLtools and integrated with AMD-GWAS and eQTL data using MOLOC. The effect of variants on gene expression was examined by dual-luciferase reporter assays in HEK293 cells.
The reference transcriptome of human retina included 13,662 protein-coding genes and 1,462 non-coding genes. Cis-eQTL analysis revealed 10,474 genes under genetic regulation, including 4,541 eQTLs detected only in the retina. Integrated analysis of AMD-GWAS with eQTLs ascertained likely target genes at six reported loci. Using transcriptome wide association analysis, we identified three new AMD genes after Bonferroni correction. Pathway analysis revealed upregulation of immune regulation and cholesterol metabolism pathways in early and advanced AMD. Several AMD-associated pathways operate through closely connected networks in the retina. mQTL analysis identified 52,030 genetic signals regulating 51,431 CpG sites. AMD GWAS and eQTL signals colocalized near three genes. The eQTL at PILRA/PILRB locus was significant as mQTL as well. Two of the six AMD-associated variants identified in both eQTL and mQTL analyses showed significant increase in the luciferase activity.
Our data represent the largest resource of retinal transcriptome and methylome and highlights the genes, pathways and regulatory mechanisms associated with AMD progression. This resource provides functional genomic foundation for post-GWAS analysis across multiple vision-related complex traits, especially glaucoma and diabetic retinopathy.
This abstract was presented at the 2019 ARVO Annual Meeting, held in Vancouver, Canada, April 28 - May 2, 2019.
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