DNA methylation changes and increased mRNA expression of coagulation proteins, Factor V and Thrombomodulin in Fuchs endothelial corneal dystrophy

Abstract

Late-onset Fuchs endothelial corneal dystrophy (FECD) is a disease affecting the corneal endothelium (CE) associated with a cytosine-thymine-guanine repeat expansion at the CTG18.1 locus in the transcription factor 4 ( TCF4 ) gene. It is unknown if CTG18.1 expansions affect global methylation including TCF4 gene in CE, or if global CE methylation changes at advanced age. Using genome-wide DNA methylation array, we investigated methylation in CE from FECD patients with CTG18.1 expansions and studied if the methylation in healthy CE is age dependent. The most significant DNA methylation findings were analyzed by gene expression and protein analysis. 3488 CpGs had significantly altered methylation pattern in FECD though no significant changes were found in TCF4 . The most hypermethylated site was in promoter of aquaporin 1 ( AQP1 ) gene, and the most hypomethylated site was in promoter of coagulation factor V ( F5 ). In FECD, AQP1 mRNA expression was variable while F5 gene expression showed a ~23-fold increase. FV protein was present in both healthy and affected CE. Further gene expression analysis of coagulation factors interacting with FV revealed a ~34-fold increase of thrombomodulin ( THBD ). THBD protein was detected only in CE from FECD patients. Additionally, we observed an age-dependent hypomethylation in elderly healthy CE. Thus, tissue-specific genome-wide and gene-specific methylation changes associated with altered gene expression were discovered in FECD. An age-associated hypomethylation was observed in healthy elderly CE, that could contribute to FECD’ late onset. TCF4 pathological methylation in FECD because of CTG18.1 expansion was ruled out.