Gene expression and missplicing in the corneal endothelium of patients with a TCF4 trinucleotide repeat expansion without fuchs’ endothelial corneal dystrophy

PURPOSE. CTG trinucleotide repeat (TNR) expansion in an intron of the TCF4 gene is the most common genetic variant associated with Fuchs’ endothelial corneal dystrophy (FECD). Although several mechanisms have been implicated in the disease process, their exact pathophysiologic importance is unclear. To understand events leading from TCF4 TNR expansion to disease phenotype, we characterized splicing, gene expression, and exon sequence changes in a rare cohort of patients with TNR expansions but no phenotypic FECD (REþ/FECD-). METHODS. Corneal endothelium and blood were collected from patients undergoing endothelial keratoplasty for non-FECD corneal edema. Total RNA was isolated from corneal endothelial tissue (n ¼ 3) and used for RNASeq. Gene splicing and expression was assessed by Mixture of Isoforms (MISO) and MAP-RSeq software. Genomic DNA was isolated from blood mononuclear cells and used for whole genome exome sequencing. Base calling was performed using Illumina’s Real-Time Analysis. RESULTS. Three genes (MBNL1, KIF13A, AKAP13) that were previously identified as misspliced in patients with a CTG TNR expansion and FECD disease (REþ/FECDþ) were found normally spliced in REþ/FECD- samples. Gene expression differences in pathways associated with the innate immune response, cell signaling (e.g., TGFb, WNT), and cell senescence markers were also identified between REþ/FECD- and REþ/FECDþ groups. No consistent genetic variants were identified in REþ/FECD- patient exomes. CONCLUSIONS. Identification of novel splicing patterns and differential gene expression in REþ/ FECD- samples provides new insights and more relevant gene targets that may be protective against FECD disease in vulnerable patients with TCF4 CTG TNR expansions.