Role of a TGF-beta 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质疾病，全世界近三分之一的女性和超过55%的美国人患有骨质疏松症，导致仅在美国每年的医疗费用就超过190亿美元。自从本实验室发现TIEG基因以来，我们已经证实TIEG基因在小鼠中的表达缺失导致成骨细胞和破骨细胞的多种缺陷，并使性别特异性的骨量减少表型仅影响雌性动物。最近，TIEG被确认为少数几个表达水平改变或等位基因变异与人类骨量减少和骨质疏松相关的基因之一。在过去的资助期间，我们已经证明了TIEG在骨骼中的表达是雌激素信号转导所必需的，并已经确定了TIEG在骨骼中介导典型的Wnt信号的新角色。这些观察结果很有意义，因为雌激素仍然是男性和女性骨骼稳态最重要的调节因素之一，而且Wnt信号对正常的骨骼发育和维护至关重要。出于这些原因，了解这些通路在骨架中的调节是至关重要的。在这个提案中，我们提出了TIEG通过双重机制增强Wnt信号通路的证据，通过这种机制，TIEG直接抑制skerostin的表达，并作为？-catenin的转录辅助激活因子。此外，我们的数据表明，TIEG是雌激素和骨骼中典型的Wnt通路之间的直接联系。基于这些观察，我们的中心假设是TIEG是典型的Wnt信号的关键组成部分，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。为了验证这一假说，当局提出了下列具体目标：1)鉴定TIEG对硬化素基因表达的调节，并确定其对观察到的TIEG基因敲除小鼠的成骨表型的贡献；2)表征TIEG通过共激活-catenin来增强规范Wnt信号的能力；以及3)确定TIEG KO小鼠的性别特异性(仅限雌性)成骨表型是否源于雌激素和骨中典型Wnt信号通路之间的缺陷信号传导。为了解决这些特定的目标，我们将使用体外方法来确定TIEG抑制骨细胞中的硬化素表达和增强成骨细胞中的ç-catenin功能的分子机制。此外，我们将利用多种体内模型和方法来剖析TIEG在调节典型的Wnt途径活性以及骨骼中雌激素和Wnt信号之间的串扰中的作用。考虑到雌激素和Wnt信号在骨骼中的基础作用，以及目前针对这些途径治疗骨质疏松症的靶向，有必要了解TIEG在调节这两条途径中的作用。因此，拟议研究的完成有望为制定治疗这种衰弱疾病的新战略提供重要知识。

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 TGFß 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 ß-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 ß-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 ß-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.