Fuchs Endothelial Corneal Dystrophy associated risk variant, rs3768617 in LAMC1 shows allele specific binding of GFI1B.

AIMS: To investigate the genetic and functional association of an intronic variant of LAMC1, rs3768617 with Fuchs endothelial corneal dystrophy (FECD) in the Indian population. METHODS: Blood samples were collected from age and sex matched 356 controls and 120 FECD patients after a detailed assessment via specular microscopy. Genomic DNA was extracted and genotyping was done by fluorescence based capillary electrophoresis. The genetic association of rs3768617 polymorphisms was computed by the chi-square (χ2) test. Bioinformatics studies were performed to find the allele specific binding of different transcription factors in the region of rs3768617 and functional evaluation assessed by luciferase assay followed by Electrophoretic Mobility Shift Assay (EMSA) and Chromatin Immunoprecipitation assay (ChIP). Immunofluorescence assay was carried out to check for any differential expression of GFI1B between control and FECD endothelium samples. RESULTS: SNP rs3768617 {chr1:183123365 (GRCh38.p13)} was found to be genetically associated with FECD in Indian population (p = 2.646 × 10-8). Luciferase assay suggested that the rs3768617 locus has a regulatory role. In silico analysis showed that the transcription factor, GFI1B binds to the risk allele 'G' of rs3768617, but not to the protective allele 'A' which was also experimentally validated by EMSA. High enrichment of DNA flanking the surrounding region of rs3768617 was also found in presence of GFI1B specific antibody in ChIP assay. There was a 0.63 fold decrease in GFI1B expression in FECD affected corneal endothelium compared to control endothelium. CONCLUSIONS: The genetic association of rs3768617 in LAMC1 with FECD pathogenesis is mediated by GFI1B, thus finding the functional role of LAMC1 in FECD pathogenesis.

Genetic association of TCF4 intronic polymorphisms, CTG18.1 and rs17089887, with Fuchs' endothelial corneal dystrophy in an Indian population.

PURPOSE: To assess the genetic association of transcription factor 4 (TCF4) intronic polymorphisms and CTG18.1 allele in individuals with Fuchs' endothelial corneal dystrophy (FECD) individuals from a sample Indian population. METHODS: Forty-four FECD patients and 108 unrelated age-matched controls were recruited with informed consent for this study. Three, single nucleotide polymorphisms (SNPs) spanning the third intronic region of TCF4 (rs613872, rs17089887, and rs17089925) and an unstable trinucleotide repeat CTG18.1 allele were genotyped by direct sequencing using Sanger's method. The association of polymorphisms was analyzed using χ(2) test and logistic regression. RESULTS: SNP rs17089887 (P = 0.013) and CTG18.1 (P = 2 × 10(-4)) alleles were found to be significantly associated with FECD in the sample Indian population. However, the other two SNPs, rs613872 and rs17089925, were not likewise associated. Thirty-four percent of FECD subjects and 5% of control individuals harbor more than 50 trinucleotide repeats, which was considered as the disease threshold. CONCLUSIONS: TCF4 poses a major contributor to FECD manifestation globally, with a significant association of rs17089887 and CTG18.1 allele in the Indian population.