More specifically, Sox9 is highly expressed in a population of mesenchyme cells within the endocardial cushions that later give rise to mature valve structures . During early stages of endocardial cushion formation, Sox9 is required to expand the pool of proliferating mesenchyme precursor cells . Following endocardial cushion formation, Sox9 expression is maintained in valve primorida and studies in mice suggest that similar to functions in the skeletal system, Sox9 promotes cartilaginous matrix phenotypes . Remodeling heart Ponatinib valves from embryonic Col2a1cre;Sox9fl/fl mice express significantly reduced levels of Col2a1 and CLP , while valves from viable Col2a1cre;Sox9fl/+ adult mice are calcified and express high levels of osteogenic genes including Runx2 and Spp1 . This valvular phenotype in Sox9 mutant mice is consistent with human calcific valve disease, frequently 601514-19-6 described as a pathological ��bone-like�� process . However, the molecular mechanism by which Sox9 prevents pathological matrix mineralization in normal cartilaginous heart valve connective tissue has not been examined. In this study, we use a genome-wide microarray technique to screen for interactions with novel target gene DNA enriched by Sox9 chromatin immunoprecipitation in primary neonate limb tissue. ChIP-on-chip analysis revealed Sox9 binding to over 450 promoters, including a region approximately 5000 bp upstream of the osteogenic glycoprotein, Spp1. At the functional level, luciferase assays show that Sox9 significantly represses Spp1 transactivation through interaction with an SRY binding site. Loss of Sox9 function in primary heart valve explants and chondrocyte cultures increases Spp1 transcript levels, while expression of chondrogenic matrix genes including Col2a1 and CLP are decreased. Conversely, Sox9 gain of function promotes Col2a1 and CLP expression. Finally, we show that Spp1 function is required for matrix mineralization induced by Sox9 knockdown in these culture systems. Taken together, these data suggest that in maturing heart valves and chondrocytes, Sox9 negatively regulates matrix mineralization through repressive regulation of Spp1. Our previous work has shown that several molecular phenotypes and signaling pathways are common to the skeletal system and heart valves. Similarly, Sox9 positively promotes cartilage and negatively regulates matrix mineralization in both these systems .