Genetic Interaction Between SOX9 and RUNX2/CBFA1

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DESCRIPTION (provided by applicant): This NICDR-K22 career development award application for Dr. Guang Zhou proposes a one-year Scholar Development Phase and a four-year Faculty Transition Phase. His Scholar Development Phase training will be performed at Baylor College of Medicine under the mentoring of Dr. Brendan Lee and Dr. Gerard Karsenty. During this period the proposed career development activities include Skeletal Dysplasia Clinical at Texas Children Hospital, special courses on craniofacial and dental genetics at UT-Dental Branch, mouse genetic seminar at M.D. Anderson Cancer Center, in situ hybridization and ES cell technique at Dr. Lee's laboratory, and bone histology training at Dr. Karsenty's laboratory. Dr. Zhou's immediate career goal is to complete his postdoctoral training and obtain an independent investigator position in an academic/medical center environment. For his long-term career Dr. Zhou will be committed to the biomedical research on bone and cartilage pathophysiology. Skeletogenesis involves the coordinated process of patterning, cell fate commitment, differentiation, growth, and remodeling. RUNX2/CBFA1 is a transcription factor essential for osteoblast differentiation, chondrocyte hypertrophy, and tooth formation. However little is known about the regulation of RUNX2 expression, especially during chondrogenesis. We recently identified SOX9, a transcription factor essential for cartilage formation including cranial neural crest cell differentiation, as a potent transcription repressor for RUNX2 transactivation. We propose to utilize both in vivo and in vitro tools to further understand the interaction between SOX9 and RUNX2 during skeletongenesis, especially chondrocyte hypertrophy. We will use in situ hybridization technique to study in parallel the expression patterns of Sox9 and Runx2 during chondrogenesis. GST-pulldown experiments will be performed to examine whether SOX9 physically interacts with RUNX2. In tissue culture systems, we will over-express SOX9 or its repression domain in chondrogenic cell line ATDC5 and pluripotent mesenchymal precursor cell line C2C12 to determine whether they can disrupt chondrocyte hypertrophy and osteogenesis in vitro. SOX9 effects on Runx2 expression and its downstream genes will be analyzed by Northern analysis and enzymatic assays. We will also generate transgenic mice mis-expressing SOX9 in hypertrophic chondrocytes to determine the consequences in vivo on long bone growth. This study will help us understand how proper skeletal patterning and cell fate commitment are achieved.