Whole-genome methylation profiling of the retinal pigment epithelium of individuals with age-related macular degeneration reveals differential methylation of the SKI, GTF2H4, and TNXB genes

Louise F Porter, Neil Saptarshi, Yongxiang Fang, Sonika Rathi, Anneke I den Hollander, Eiko K de Jong, Simon J Clark, Paul N Bishop, Timothy W Olsen, Triantafillos Liloglou, Venkata R M Chavali, Luminita Paraoan

Research output: Contribution to journalArticle (journal)peer-review

47 Citations (Scopus)


BACKGROUND: Age-related macular degeneration (AMD) is a degenerative disorder of the central retina and the foremost cause of blindness. The retinal pigment epithelium (RPE) is a primary site of disease pathogenesis. The genetic basis of AMD is relatively well understood; however, this knowledge is yet to yield a treatment for the most prevalent non-neovascular disease forms. Therefore, tissue-specific epigenetic mechanisms of gene regulation are of considerable interest in AMD. We aimed to identify differentially methylated genes associated with AMD in the RPE and differentiate local DNA methylation aberrations from global DNA methylation changes, as local DNA methylation changes may be more amenable to therapeutic manipulation.

METHODS: Epigenome-wide association study and targeted gene expression profiling were carried out in RPE cells from eyes of human donors. We performed genome-wide DNA methylation profiling (Illumina 450k BeadChip array) on RPE cells from 44 human donor eyes (25 AMD and 19 normal controls). We validated the findings using bisulfite pyrosequencing in 55 RPE samples (30 AMD and 25 normal controls) including technical (n = 38) and independent replicate samples (n = 17). Long interspersed nucleotide element 1 (LINE-1) analysis was then applied to assess global DNA methylation changes in the RPE. RT-qPCR on independent donor RPE samples was performed to assess gene expression changes.

RESULTS: Genome-wide DNA methylation profiling identified differential methylation of multiple loci including the SKI proto-oncogene (SKI) (p = 1.18 × 10-9), general transcription factor IIH subunit H4 (GTF2H4) (p = 7.03 × 10-7), and Tenascin X (TNXB) (p = 6.30 × 10-6) genes in AMD. Bisulfite pyrosequencing validated the differentially methylated locus cg18934822 in SKI, and cg22508626 within GTF2H4, and excluded global DNA methylation changes in the RPE in AMD. We further demonstrated the differential expression of SKI, GTF2H4, and TNXB in the RPE of independent AMD donors.

CONCLUSIONS: We report the largest genome-wide methylation analysis of RPE in AMD along with associated gene expression changes to date, for the first-time reaching genome-wide significance, and identified novel targets for functional and future therapeutic intervention studies. The novel differentially methylated genes SKI and GTF2H4 have not been previously associated with AMD, and regulate disease pathways implicated in AMD, including TGF beta signaling (SKI) and transcription-dependent DNA repair mechanisms (GTF2H4).

Original languageEnglish
Article number6
Pages (from-to)6
JournalClinical Epigenetics
Issue number1
Publication statusPublished - 14 Jan 2019


  • Aged
  • Autopsy
  • Case-Control Studies
  • DNA Methylation
  • DNA-Binding Proteins/genetics
  • Epigenesis, Genetic
  • Female
  • Gene Expression Profiling
  • Gene Expression Regulation
  • Genetic Association Studies
  • High-Throughput Nucleotide Sequencing
  • Humans
  • Macular Degeneration/genetics
  • Male
  • Middle Aged
  • Organ Specificity
  • Proto-Oncogene Mas
  • Proto-Oncogene Proteins/genetics
  • Retinal Pigment Epithelium/chemistry
  • Tenascin/genetics
  • Transcription Factors, General/genetics
  • Transcription Factors, TFII/genetics
  • Whole Genome Sequencing/methods


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