Effect of Gli2 on BMP-2 gene expression & bone formation

描述(申请人提供)：骨形态发生蛋白-2(BMP-2)在控制骨骼形态发生和出生后骨形成方面都很重要。然而，人们对调控BMP-2表达的因素知之甚少。因此，迫切需要确定在成骨过程中BMP-2基因表达调控的分子机制，这是候选者的短期目标。这项建议的直接目标是验证锌指蛋白Gli2通过调节BMP-2表达来影响骨形成的假设。候选人的长期目标是发展独立的研究生涯，并建立一个坚实的多资金项目，研究骨形成激活的分子机制，特别是骨合成代谢信号和基因调控。应聘者将使用这款K01来获得骨分子生物学方面的额外技能。导师Mundy博士和Harris博士是骨生物学领域的知名专家，他们将为候选人在圣安东尼奥UTHSC的“最先进”机构设施中提供出色的培训经验。在第一阶段的奖励中，候选人将受益于两位导师的科学指导。到第二阶段，候选人将成为一名完全独立的调查员。遗传学研究表明，转录因子GN2的干扰会导致严重的骨缺损，我们的初步数据表明，GH2上调了BMP-2基因的转录。建议的研究试图证实这些初步观察结果，并阐明GH2对BMP-2表达以及随后的成骨细胞分化和骨形成的影响机制。我们提出了四个具体的目标：(1)确定GH2功能丧失对成骨细胞分化和BMP-2基因表达的影响；(2)确定Gli2反式激活BMP-2基因的分子机制；(3)检测GH2过表达对Gli2转基因小鼠体内骨形成的影响；(4)研究E3泛素连接酶β-TrCP对成骨细胞GH2加工的影响。充分研究Gli2蛋白在BMP-2基因表达调控和骨形成过程中的作用，有助于深入了解骨合成代谢调控的分子机制。

DESCRIPTION (provided by applicant): Bone morphogenetic protein-2 (BMP-2) is important in the control of both skeletal morphogenesis and postnatal bone formation. However, little is known of the factors that regulate BMP-2 expression. Therefore, there is a great need to identify molecular mechanisms that are involved in the regulation of BMP-2 gene expression in osteogenesis, which serves as the short term goal of the candidate. The immediate objective of this proposal is to test the hypothesis that the zinc finger protein Gli2 influences bone formation by its regulatory effects on BMP-2 expression. The candidate's long-term goal is to develop an independent research career and to establish a solid multi-funded program on the studies of molecular mechanisms underlying the activation of bone formation, specifically focusing on bone anabolic signaling and gene regulation. The candidate will use this K01 to acquire additional skills in bone molecular biology. The mentors, Dr. Mundy and Dr. Harris, renowned experts in the field of bone biology, will provide excellent training experience for the candidate in "state of the art" institutional facilities at UTHSC at San Antonio. During phase I of the award, the candidate will benefit from the scientific guidance of two mentors. By phase II, the candidate will become a fully independent investigator. Genetic studies have shown that disruption of transcription factor GN2 results in severe bone defects, and our preliminary data suggests that GH2 up-regulates BMP-2 gene transcription. The proposed studies seek to confirm these preliminary observations, and to elucidate the mechanisms mediating the effect of GH2 on BMP-2 expression as well as on subsequent osteoblast differentiation and bone formation. Four specific aims are proposed (1) to determine the effects of loss-of-function of GH2 on osteoblast differentiation and BMP-2 gene expression, (2) to determine the molecular mechanisms by which Gli2 transactivates BMP-2 gene, (3) to examine the effects of overexpression of GH2 on bone formation in vivo in Gli2 transgenic mice and (4) to investigate the effects of the E3 ubiquitin ligase beta-TrCP on GH2 processing in osteoblasts. Full characterization of the roles of Gli2 protein in the regulation of BMP-2 gene expression and skeletogenesis should lead to insights into the molecular mechanisms of bone anabolic regulation.