Integration of Signaling Pathways Regulating Chondrocyte Differentiation

调节软骨细胞分化的信号通路整合

基本信息

批准号：
7577207
负责人：
SHERRILL L. ADAMS
金额：
$ 35.28万
依托单位：
UNIVERSITY OF PENNSYLVANIA
依托单位国家：
美国
项目类别：
财政年份：
2009
资助国家：
美国
起止时间：
2009-07-17 至 2011-06-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/7577207
关键词：
Affect Affinity Architecture Binding Binding Sites Biological Assay Bone Marrow Bone Morphogenetic Proteins Cartilage Cartilage Matrix Cells Chondrocytes Chromatin Clinical Collagen Complex Country DNA DNA analysis Defect Degenerative polyarthritis Disease Distal Electrophoretic Mobility Shift Assay Elements Embryo Enhancers Epiphysial cartilage Extracellular Matrix Proteins Failure Family member Funding Gene Expression Genes Genetic Transcription Grant Growth Growth Disorders Growth Factor Hormones Hypertrophy Hypothyroidism Malignant Neoplasms Mesenchymal Methods Molecular Mutation Osteogenesis Pathway interactions Play Process Production Proliferating Protein Binding Proteins Proteomics Regulation Retinoic Acid Receptor Retinoids Role Signal Pathway Signal Transduction Signaling Molecule Site Skeleton Sternum Study models Testing Thyroid Diseases Thyroid Hormone Receptor Thyroid Hormones Transcription Factor AP-1 Transcription Initiation Site Transcriptional Activation Tretinoin Triiodothyronine Vitamin A Vitamin A Deficiency activating transcription factor articular cartilage bone chromatin immunoprecipitation in vivo insight mineralization mouse model premature prevent promoter protein protein interaction public health relevance research study response skeletal skin disorder therapeutic target transcription factor

项目摘要

DESCRIPTION (provided by applicant): The major focus of this grant is to identify the molecular mechanisms that regulate chondrocyte maturation during endochondral bone formation. During this process, chondrocytes undergo a differentiation process characterized by hypertrophy and production of a unique protein, collagen X (Col10). Thyroid hormone (T3), bone morphogenetic proteins (BMPs) and retinoic acid (RA, the major active metabolite of vitamin A) exert profound effects on this process. Each of these signaling pathways is essential for endochondral bone formation and cannot compensate for the others; abrogation of any one pathway (e.g. hypothyroidism, which blocks T3 signaling) prevents differentiation, resulting in shortened bones. Regulation of the Col10 gene provides a window into the molecular mechanisms by which T3, RA and BMPs control chondrocyte maturation. During the current funding period, we have shown that the T3, RA and BMP signaling pathways converge on a small distal region of the chick Col10 promoter that contains binding sites for multiple transcription factors, including RA receptors, Runx2, an Ets family member and two DNA-bending factors, Lef1 and AP1. All of these sites play a important roles in both basal and RA/BMP-stimulated transcriptional activity; thus this enhancer may function as an integrator of the various signaling pathways that activate Col10 gene transcription. We have demonstrated a hierarchical network of these effectors, in which T3 acts indirectly on Col10 gene expression by stimulating both retinoid and BMP signaling. In addition, we have shown that RA has both direct and indirect effects, stimulating Col10 transcription directly, but also further stimulating BMP signaling. In this renewal application, we propose the following specific aims: 1) We will test the hypothesis that alterations in binding of transcription factors and comodulators that alter promoter and chromatin architecture are responsible for the dramatic increase in Col10 that occur during chondrocyte maturation in vivo and in response to T3, BMPs and RA in culture; and 2) We will test the hypothesis that T3 activates Col10 transcription through stimulation of BMP and RA signaling and this stimulation is essential for T3 effects. The in vivo experiments will utilize two unique mouse models with targeted inactivation of thyroid hormone receptors. One strain displays skeletal hypothyroidism, resulting in delayed endochondral bone formation, while the other displays skeletal thyrotoxicotis, resulting in accelered endochondral ossification; both result in short stature. These experiments will provide significant mechanistic insights into the problems of short stature due to failure of growth plate differentiation or premature growth plate closure. PUBLIC HEALTH RELEVANCE: Skeletal growth is profoundly affected by thyroid hormones, vitamin A and bone morphogenetic proteins (BMPs). There are millions of people in this country with undiagnosed thyroid disease and many more millions worldwide with vitamin A deficiency; furthermore, vitamin A derivatives and BMPs are in widespread clinical use, and the impact of their use on skeletal growth is not well understood. The proposed experiments will provide significant insights into the roles these growth factors and hormones play in failure of normal skeletal growth, resulting in short stature.

描述（由申请人提供）：该赠款的主要重点是确定在内向骨骨形成过程中调节软骨细胞成熟的分子机制。在此过程中，软骨细胞经历了以肥大和产生独特蛋白质胶原蛋白X（Col10）的生产为特征的分化过程。甲状腺激素（T3），骨形态发生蛋白（BMP）和视黄酸（RA，维生素A的主要活性代谢产物）对这一过程产生深远的影响。这些信号通路中的每一个对于内侧软骨骨形成都是必不可少的，无法补偿其他信号。废除任何一种途径（例如，甲状腺功能减退症，阻断T3信号传导）可阻止分化，从而导致骨骼缩短。 Col10基因的调节为T3，RA和BMPS控制软骨细胞成熟的分子机制提供了一个窗口。在当前的融资期间，我们已经表明，T3，RA和BMP信号通路在Chick Col10启动子的一个小远端区域汇聚，该区域包含多种转录因子的结合位点，包括RA受体，Runx2，ETS家族成员和两个DNA弯曲因子LEF1和AP1。所有这些位点在基底和RA/BMP刺激的转录活性中都起着重要作用。因此，该增强子可以充当激活Col10基因转录的各种信号通路的集成剂。我们已经证明了这些效应子的分层网络，其中T3通过刺激性类维生素类和BMP信号传导间接作用于Col10基因表达。此外，我们已经表明RA具有直接和间接效应，直接刺激Col10转录，但也进一步刺激了BMP信号传导。在此续签应用中，我们提出以下具体目的：1）我们将测试以下假设：转录因子和更改启动子和染色质结构的结合变化是导致COL10的急剧增加，而在软骨细胞成熟过程中发生了急剧增加，并且对T3，BMP和RA的培养中的响应，在培养中响应T3，BMP和RA； 2）我们将检验以下假设：T3通过刺激BMP和RA信号传导激活COL10转录，而这种刺激对于T3效应至关重要。体内实验将利用两种独特的小鼠模型，具有针对性的甲状腺激素受体的灭活。一种菌株显示骨骼甲状腺功能减退症，导致延迟的内软骨骨形成，而另一个则显示骨骼甲状腺毒素，从而导致加速的内软骨骨化；两者都导致身材矮小。这些实验将为由于生长板分化或过早生长板闭合而导致的矮小身材问题提供重要的机械见解。公共卫生相关性：骨骼生长受到甲状腺激素，维生素A和骨形态发生蛋白（BMP）的深刻影响。这个国家有数百万的人患有未诊断的甲状腺疾病，全世界有数百万人患有维生素A缺乏症。此外，维生素A衍生物和BMP在广泛的临床上使用，其使用对骨骼生长的影响尚不清楚。提出的实验将为这些生长因子和激素在正常骨骼生长失败中的作用提供重大见解，从而导致身材矮小。