Genetic and epigenetic factors fine-tune TGFB1 expression within the osteoarthritic articular joint.

OBJECTIVE Osteoarthritis (OA) is an age-related disease characterised by articular cartilage degeneration. It has a large heritability and genetic screens have identified single nucleotide polymorphisms (SNPs) marking genomic risk loci. One such locus is marked by G\u003eA SNP rs75621460, downstream of TGFB1. This gene encodes TGF-β1, the correct expression of which is essential for cartilage maintenance. We have used a combination of human patient samples (n=319) and a chondrocyte model to characterise the impact of rs75621460 in multiple articular joint tissues. METHODS Patient samples were genotyped and DNA methylation (DNAm) levels quantified by pyrosequencing. Gene reporter and electrophoretic mobility shift assays were used to determine differential nuclear protein binding to the region. The functional impact of DNAm upon TGFB1 expression was tested using targeted epigenome editing. RESULTS We identified that rs75621460 is located within a TGFB1 enhancer, and that the OA risk A-allele alters transcription factor binding, decreasing enhancer activity. Protein complexes binding to A (but not G) induced DNAm at flanking CG-dinucleotides. Strong correlations were observed between patient DNAm levels and TGFB1 expression, the direction of which was opposing between cartilage and synovium. This demonstrated biological pleiotropy in the impact of the SNP within different tissues of the articulating joint. CONCLUSION The OA risk SNP rs75621460 impacts TGFB1 expression by modulating the function of a gene enhancer. We propose a mechanism by which the SNP impacts enhancer function, providing novel biological insight into one mechanism of osteoarthritis genetic risk, which may facilitate the development of future pharmacological therapies.