Role of a TGF-? Regulated Gene in Human and Mouse Osteoblasts and Skeleton

描述（由申请人提供）：骨质疏松症是一种骨骼疾病，影响全球近三分之一的女性，超过55%的美国人，仅在美国每年的医疗保健费用就超过190亿美元。自从发现TGF？诱导早期基因-1（TIEG）的表达，我们已经证明小鼠中TIEG表达的缺失导致成骨细胞和破骨细胞的多种缺陷，并赋予仅影响雌性动物的性别特异性骨质减少表型。最近，TIEG已被鉴定为仅有的几个基因之一，其改变的表达水平或等位基因变异与人类骨量减少和骨质疏松症相关。在过去的资助期间，我们已经表明TIEG表达对于骨中的雌激素信号传导是必不可少的，并且已经确定了TIEG在介导骨骼中的经典Wnt信号传导中的新作用。这些观察结果具有重要意义，因为雌激素仍然是男性和女性骨稳态的最重要调节剂之一，并且因为Wnt信号传导对于正常骨发育和维持至关重要。由于这些原因，了解骨骼中这些通路的调节至关重要。在这个建议中，我们提出的证据表明，TIEG通过双重机制增强Wnt信号通路，从而直接抑制sclerostin的表达，并作为转录共激活剂？连环蛋白。此外，我们的数据表明，TIEG作为骨中雌激素和经典Wnt通路之间的直接联系。基于这些观察结果，我们的中心假设是TIEG是经典Wnt信号传导的重要组成部分，并在调节骨骼中雌激素和Wnt通路之间的串扰中发挥核心作用。为了验证这一假设，提出了以下具体目的：1）：表征TIEG对硬化蛋白基因表达的调节，并确定其对TIEG敲除小鼠观察到的骨质减少表型的贡献; 2）：表征TIEG通过共激活？连环蛋白;和3）确定TIEG KO小鼠的性别特异性（仅雌性）骨质减少表型是否由骨中雌激素和经典Wnt信号传导途径之间的缺陷性串扰引起。为了解决这些具体目标，我们将采用体外方法来确定TIEG抑制骨细胞中sclerostin表达并增强骨细胞中sclerostin表达的分子机制。连环蛋白在成骨细胞中的功能。此外，我们将利用多种体内模型和方法来剖析TIEG在调节经典Wnt通路活性和骨骼中雌激素与Wnt信号传导之间的串扰中的作用。考虑到雌激素和Wnt信号传导在骨中的基本作用，以及目前针对这些途径治疗骨质疏松症，了解TIEG在调节这两种途径中的作用至关重要。因此，预计完成拟议的研究将为开发治疗这种使人衰弱的疾病的新策略提供重要的知识。 公共卫生相关性：骨质疏松症是一种骨骼疾病，其特征是骨矿物质密度降低和骨微结构改变，最终导致骨折。骨质疏松症影响大约55%的美国人，导致每年近190亿美元的医疗保健费用，预计到2025年这一数字将超过250亿美元。该项目涉及确定TIEG在调节骨中经典Wnt和雌激素信号通路中的作用，TIEG是少数被鉴定为与人类骨质疏松症的发展临床相关的基因之一。这些研究的结果有望进一步加深我们对骨质疏松症的理解，并有助于开发治疗这种使人衰弱的疾病的新疗法。

DESCRIPTION (provided by applicant): Osteoporosis is a bone disease affecting nearly 1 in 3 women worldwide and over 55% of all Americans resulting in health care costs exceeding 19 billion dollars annually in the United States alone. Since the discovery of the TGF? inducible early gene-1 (TIEG) by our laboratory, we have demonstrated that loss of TIEG expression in mice results in multiple defects in osteoblasts and osteoclasts and confers a gender specific osteopenic phenotype affecting only female animals. Recently, TIEG has been identified as one of only a few genes whose altered expression levels or allelelic variations associate with decreased bone mass and osteoporosis in humans. During the past funding period, we have shown that TIEG expression is essential for estrogen signaling in bone and have identified a novel role for TIEG in mediating canonical Wnt signaling in the skeleton. These observations are of significant interest since estrogen remains one of the most important regulators of bone homeostasis in both men and women and since Wnt signaling is essential for normal bone development and maintenance. For these reasons, it is crucial to understand the regulation of these pathways in the skeleton. In this proposal, we present evidence that TIEG enhances the Wnt signaling pathway via dual mechanisms whereby it directly suppresses the expression of sclerostin and serves as a transcriptional co- activator for ?-catenin. Furthermore, our data suggest that TIEG serves as a direct link between the estrogen and canonical Wnt pathways in bone. Based on these observations, our central hypothesis is that TIEG is a crucial component of canonical Wnt signaling and serves a central role in regulating cross-talk between the estrogen and Wnt pathways in the skeleton. In order to test this hypothesis, the following Specific Aims are proposed: 1): Characterize the regulation of sclerostin gene expression by TIEG and determine its contribution to the observed osteopenic phenotype of TIEG knockout mice; 2): Characterize the ability of TIEG to enhance canonical Wnt signaling through co-activation of ?-catenin; and 3) Determine if the gender specific (female only) osteopenic phenotype of TIEG KO mice results from defective cross-talk between the estrogen and canonical Wnt signaling pathways in bone. To address these Specific Aims, we will employ in vitro approaches to identify the molecular mechanisms by which TIEG suppresses sclerostin expression in osteocytes and enhances ?-catenin function in osteoblasts. Additionally, we will utilize multiple in vivo models and approaches to dissect the role of TIEG in regulating canonical Wnt pathway activity and cross-talk between estrogen and Wnt signaling in the skeleton. Considering the fundamental roles of estrogen and Wnt signaling in bone, as well as the present day targeting of these pathways for the treatment of osteoporosis, it is essential to understand the role of TIEG in regulating these two pathways. Completion of the proposed studies is therefore expected to provide significant knowledge with regard to developing novel strategies to treat this debilitating disease. PUBLIC HEALTH RELEVANCE: Osteoporosis is a skeletal disease characterized by decreased bone mineral density and altered bone microarchitecture which ultimately results in bone fractures. Osteoporosis affects approximately 55% of all Americans resulting in health care costs of nearly 19 billion dollars annually, a figure that is expected to surpass 25 billion dollars by the year 2025. This project involves determining the role of TIEG, one of only a handful of genes identified to be clinically associated with the development of osteoporosis in humans, in regulating the canonical Wnt and estrogen signaling pathways in bone. Outcomes of the proposed studies are expected to further our understanding of osteoporosis and aid in the development of novel therapies to treat this debilitating disease.