FUNCTION AND REGULATION OF CD-RAP

项目总结/摘要 到2020年，骨关节炎将影响5000万美国人疾病的过程是 其特征在于基因表达异常和软骨细胞不能 修复导致软骨退化的细胞外基质。CD-RAP是一种小型 软骨特征性蛋白，在软骨中与II型胶原共表达， 聚集蛋白聚糖。它是一个紧凑的基因，与IIB型胶原共表达， 软骨形成，使其成为研究软骨细胞特异性基因的良好模型 表情在过去的四年里，我们已经扩展了我们对一个基因的了解， 涉及软骨细胞表型的转录调节的分析。 计算和实验方法相结合，以分析组的共同- 表达的基因，并确定共同的调控结构域的基因表达， 关节软骨目前的建议将侧重于扩展这些分析，以确定 转录调控网络主要在关节炎中起作用并定义关节炎。 软骨发育（与导致骨形成的生长板相反）。 随着新的转录或信号传导因子的出现，特定的活性机制将被阐明。 使用我们的CD-RAP和COL 2A 1基因模型或细胞培养来解密。具体 该提案的目的是：（1）确定转录调控机制， 通过E-box蛋白表达CD-RAP基因 EF-1/USF1/USF2、C/EBPss和HMGA。（二） 明确转录因子网络在软骨干细胞分化中的作用 （3）定义转录调控和信号传导在早期细胞模型中的作用， 骨关节炎和（4）从我们在具体目标3中的初步实验中产生， 研究EGF和FGF信号转导的细胞内调节因子Sprouty-4在 软骨形成和骨关节炎。实验方法将集中在（1）使用 全基因组表达分析，以确定共调节基因，（2）筛选 使用1700寡核苷酸阵列表达转录因子，（3）计算 分析共调节基因启动子的转录因子结合域， 和（4）对特异性软骨基因CD-RAP的机制功能的确认 COL2A1。这些研究将采用强大的技术，结合实验和 计算方法可以超越候选基因来阐明调控 对于开发控制软骨形成策略的关键机制， 发展，在组织工程和修复，并最终帮助找到生物 控制骨关节炎的方法。

Project Summary/Abstract Osteoarthritis will affect 50 million Americans by 2020. The disease process is characterized by the aberrant gene expression and the inability of cartilage chondrocytes to repair the extracellular matrix leading to cartilage degeneration. CD-RAP is a small cartilage-characteristic protein that is co-expressed in cartilage with type II collagen and aggrecan. It is a compact gene and co-expressed with type IIB collagen in chondrogenesis, making it an excellent model for study of chondrocyte-specific gene expression. Over the past four years, we have extended our knowledge of a single gene to the analysis of the transcriptional regulation of the chondrocyte phenotype. Computational and experimental methods were combined to analyze groups of co- expressed genes and define common regulatory domains of genes expressed in articular cartilage. The current proposal will focus on extending these analyses to define transcriptional regulatory networks that predominantly operate in and define articular cartilage development (as opposed to the growth plate which results in bone formation). As new transcription or signaling factors arise, specific mechanisms of activity will be deciphered using our CD-RAP and COL2A1 gene models or cell culture. The specific aims of the proposal are: (1) Determine the mechanism of transcriptional regulation of the CD-RAP gene by the E-box proteins EF-1/USF1/USF2, C/EBPss, and HMGA. (2) Define the role of transcription factor networks in differentiation of cartilage from stem cells (3) Define the role of transcriptional regulation and signaling in a model for early osteoarthritis and (4) Arising from our preliminary experiments in Specific Aim 3, investigate the role of the intracellular regulator of EGF and FGF signaling, Sprouty-4, in chondrogenesis and osteoarthritis. The experimental approach will focus on (1) using genome-wide expression analyses to determine co-regulated genes, (2) screening for expression of transcription factors using a 1700 oligonucleotide array, (3) computational analyses for transcription factor binding domains of the promoters of co-regulated genes, and (4) confirmation of mechanistic function on the specific cartilage genes, CD-RAP and COL2A1. These studies will apply powerful techniques combining experimental and computational approaches to look beyond candidate genes to elucidate regulatory mechanisms critical for developing strategies to control chondrogenesis during development, in tissue engineering and repair, and ultimately to help find biological methods to control osteoarthritis.