Genetic Interaction Between SOX9 and RUNX2/CBFA1during chondrogenesis

软骨形成过程中 SOX9 和 RUNX2/CBFA1 之间的遗传相互作用

• 批准号: 7119034
• 负责人: Guang Zhou
• 金额: $ 12.71万
• 依托单位: CASE WESTERN RESERVE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2003
• 资助国家: 美国
• 起止时间: 2003-07-01 至 2008-11-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7119034
• 关键词: Genetic; Interaction; Between; SOX9; RUNX2

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.

描述（由申请人提供）：Zhou博士的NICDR-K22职业发展奖申请申请申请，提议了为期一年的学者发展阶段和四年的教职员工过渡阶段。他的学者发展阶段培训将在Brendan Lee博士和Gerard Karsenty博士的指导下在贝勒医学院进行。在此期间，提议的职业发展活动包括德克萨斯儿童医院的骨骼发育不良临床，UT-Dental Branch的颅面和牙科遗传学的特殊课程，M.D. Anderson癌症中心的鼠标遗传研讨会，现场杂交以及Lee的实验室的ES Cell Technique，以及Karsenty Blaberatory博士的Lee's Laboratory培训。周博士的直接职业目标是完成他的博士后培训，并在学术/医疗中心环境中获得独立的研究员职位。在他的长期职业生涯中，周博士将致力于骨骼和软骨病理生理学的生物医学研究。 骨骼生成涉及模式，细胞命运承诺，分化，生长和重塑的协调过程。 RUNX2/CBFA1是成骨细胞分化，软骨细胞肥大和形成牙齿的转录因子。然而，对于Runx2表达的调节，尤其是在软骨发生过程中，知之甚少。我们最近确定Sox9是软骨形成所必需的转录因子，包括颅神经rest细胞分化，是用于Runx2反式激活的有效转录阻遏物。我们建议利用体内和体外工具，以进一步了解骨骼修剪期间Sox9和Runx2之间的相互作用，尤其是软骨细胞肥大。我们将使用原位杂交技术在软骨发生过程中与SOX9和RUNX2的表达模式相似。将进行GST-Pulldown实验，以检查Sox9是否与Runx2进行了物理相互作用。在组织培养系统中，我们将过表达Sox9或其在软骨细胞系ATDC5和多能间质前体前体细胞系C2C12中过度表达的SOX9或其抑制域，以确定它们是否可以破坏软骨细胞肥大和体内生成。 Sox9对Runx2表达及其下游基因的影响将通过北部分析和酶试验分析。我们还将在肥厚的软骨细胞中产生转基因小鼠SOX9的转发，以确定体内对长骨生长的后果。这项研究将有助于我们了解如何实现适当的骨骼图案和细胞命运承诺。