Cindy Boer

82 | Chapter 2.1 involved in endochondral bone formation in mice, specifically the transition from hy- pertrophic cartilage to bone[29]. Recent, TGFA has also been implicated in the degener- ation of articular cartilage during OA in rats[30]. Our results now imply a relationship between TGFA and human OA. In addition to the genetic association, we also show that TGFA expression is higher in human OA affected versus non-lesioned cartilage, possibly indicating that TGFA has a role in cartilage remodelling. Functional characterization of the TGFA -associated locus by an examination of the histone methylation marks representing promoter or enhancer activity, did not re- veal an obvious explanation for the functional impact of the SNP. However, the examined histone mark data represent unstimulated cells, and it is anticipated that the promoter and enhancer activity change upon stimulation of the cells. It is becoming more clear that effects of SNPs can be stimulus and context dependent, as has recently been shown for human monocytes, where many regulatory variants display functionality only after pathophysiological relevant immune stimuli[31]. The identified SUPT3H-RUNX2 locus contains two variants, rs12206662 and rs10948155, which are partially independent of each other. Where rs12206662 is lo- cated in the first intron of the RUNX2 gene near the second transcription start site (the so-called P2 promoter), rs10948155 is located more than 500kb away from RUNX2 between CDC5L and SUPT3H . However, rs10948155 is in high linkage disequilibrium with SNPs near in the P2 promoter and SNPs located in chondroblast specific enhancer regions. Possibly, these enhancer regions regulate RUNX2 gene expression during en- dochondral differentiation. RUNX2 (Runt-related transcription factor 2) is a master transcription factor for controlling chondrocyte hypertrophy and osteoblast differen- tiation[32]. Previous genome-wide association studies have identified variants in the SUPT3H-RUNX2 locus associated with other bone and cartilage related phenotypes in- cluding height[14], bone mineral density[10] and ossification of the posterior longitu- dinal ligament of the spine[12]. All these previously published loci are independent of the two mJSW SNPs identified in the current study. We hypothesize that the SNPs are located in long-range enhancers, which regulate RUNX2 gene expression during endo- chondral differentiation via a chromatin-loop mediating protein. We have also identified rs10471753, with PIK3R1 (Phosphoinositide-3-Kinase, Regulatory subunit 1 alpha) as the closest and strongest prioritized gene, related to rs10471753 associated with mJSW. Mutations in this gene are known to cause the SHORT syndrome, which is a rare multisystem disease with several manifestations in- cluding short stature, hernias, hyper extensibility and delayed dentition [33]. Taken together with the fact that PIK3R1 is differentially expressed in OA affected cartilage, these results identify PIK3R1 as the most likely causal gene. Another possibility is that