Role of FoxOs in Skeletal Homeostasis

描述（由申请人提供）：Wnt/2-catenin信号是成骨细胞发生和骨形成不可或缺的。在该应用的研究中，阐明了Wnt/2-catenin拮抗的新机制，即FoxO转录因子的激活将有限的2-catenin库从Wnt诱导的TCF/ left转录转移到FoxO介导的转录。骨中的这种拮抗级联反应是由活性氧（ROS）启动的，它既是氧化应激的中介，也是氧化应激的防御机制。FoxOs通过调节参与细胞周期、DNA修复和寿命的基因表达介导氧化应激反应。组蛋白去乙酰化酶Sirt1参与ROS对Wnt/2-catenin的抑制2-catenin共同激活foxo靶基因的表达，促进成骨细胞在应激条件下的存活。此外，有证据表明糖皮质激素或TNF1对骨的作用至少部分是由ros诱导的FoxO激活介导的。上述观察结果为以下假设奠定了基础：氧化应激激活FoxO转录因子代表了一种以前未被认识到的Wnt/2-catenin拮抗的细胞自主机制，该机制通过将2-catenin从TCF-转移到FoxO介导的转录，从而导致衰老、糖皮质激素过量和炎症细胞因子对骨骼的不良影响。为了验证这一假设，我们将研究ROS诱导的FoxO翻译后修饰在成骨细胞中FoxO与2-catenin结合中的作用，以及ROS是否导致2-catenin募集到FoxO靶基因的启动子上（Aim 1）。此外，将进行体外研究，以确定FoxO激活对成骨细胞寿命和分化的影响（目的2）。最后，在成骨细胞前体及其子代（成骨细胞、骨细胞）中过表达FoxO3a的小鼠；以及FoxO三种主要亚型FoxO1、3a和4有条件缺失的小鼠，将被用来研究FoxO在骨骼稳态中的作用（目的3）。这项工作将促进对衰老、糖皮质激素过量或炎症细胞因子如何降低骨量的认识。此外，它应该提供一个更好的理解如何优化治疗这种情况。

DESCRIPTION (provided by applicant): Wnt/2-catenin signaling is indispensable for osteoblastogenesis and bone formation. In studies leading to this application, a novel mechanism of Wnt/2-catenin antagonism, whereby activation of FoxO transcription factors diverts the limited pool of 2-catenin from Wnt induced TCF/Lef- transcription to FoxO-mediated transcription was elucidated. This antagonistic cascade in bone is initiated by reactive oxygen species (ROS) and represents both a mediator of, and a defense mechanism against, oxidative stress. FoxOs mediate oxidative stress responses by regulating the expression of genes involved in cell cycle, DNA repair, and lifespan. The involvement of the histone deacetylase Sirt1 in the inhibition of Wnt/2-catenin by ROS; and co- activation by 2-catenin of the expression of FoxO-target genes that promote osteoblast survival under stress conditions was also established. Moreover, evidence was obtained that the actions of glucocorticoids or TNF1 on bone are mediated, at least in part, by ROS-induced FoxO activation. The above observations form the foundation of the hypothesis that activation of FoxO transcription factors by oxidative stress represents a previously unappreciated cell-autonomous mechanism of Wnt/2-catenin antagonism which contributes to the adverse effects of aging, glucocorticoid excess and inflammatory cytokines on bone, by diverting 2-catenin from TCF- to FoxO-mediated transcription. To test this hypothesis, the role of ROS-induced FoxO post- translational modifications on the binding of FoxOs to 2-catenin, in osteoblastic cells, and whether ROS lead to recruitment of 2-catenin to the promoter of FoxO-target genes will be investigated (Aim 1). In addition, in vitro studies will be done to establish the consequences of FoxO activation in osteoblast lifespan and differentiation (Aim 2). Finally, mice overexpressing FoxO3a in osteoblast precursors and their progeny (osteoblasts, osteocytes); and mice in which the three main FoxO isoforms FoxO1, 3a, and 4 are conditionally deleted will be used to examine the role of FoxOs in skeletal homeostasis (Aim 3). This work should advance knowledge of how aging, glucocorticoid excess or inflammatory cytokines decrease bone mass. Furthermore, it should provide a better understanding of how to optimize the treatment of this condition. PUBLIC HEALTH RELEVANCE: The proposed studies seek to identify the means by which aging, glucocorticoid excess or inflammatory cytokines cause bone loss. This will be accomplished by studying changes in the function of proteins that control bone-forming cells. Increased understanding of the mechanisms that control bone formation will provide important information for the development of therapies to maintain or increase bone mass and strength, thereby reducing the risk of osteoporotic fractures