Role and Mechanism of Action of Ksr2, an Obesity and Type 2 Diabetes Gene, in Bone Metabolism

摘要 在日益老龄化和肥胖的人口中，2型糖尿病(T2 DM)和骨质疏松性骨折是主要的公共健康 担忧。在美国，超过三分之一的成年人受到肥胖影响，肥胖是2型糖尿病最重要的风险因素。它是 现在公认的是肥胖和T2 DM对骨折风险有影响，而T2 DM的骨折与更大的 发病率高于普通人群。尽管众所周知，肥胖和2型糖尿病患者的骨密度会增加 在患者中，导致T2 DM/肥胖受试者骨折风险增加的基础原因仍有待确定。 因此，了解肥胖、2型糖尿病和骨折之间的相互作用正成为减少社会负担的迫切需要。 以及骨折的个人成本。我们识别控制分子和它们的信号通路的努力描绘了一种联系 在T2 DM、骨质疏松症和肥胖症之间导致了一个令人兴奋的发现，靶向干扰RAS的激酶抑制物 肥胖和T2 DM基因2(KSR2)导致4个月大婴儿股骨远端骨小梁骨量增加2倍 除诱发肥胖和2型糖尿病外，还可给小鼠灌胃。相比之下，一个密切相关的Ksr1基因的中断没有明显的影响 骨骼或肥胖表型。KSR1和KSR2被发现是协调Raf组装的支架蛋白， 典型的Ras-Raf-MAPKs通路中的MEK和ERK。而KSR2在下丘脑中表达的核心作用是 与控制摄食行为和能量平衡有关，从而导致肥胖，我们有了新的兴奋 初步数据表明，在成骨细胞中表达的KSR2以细胞自主的方式在局部发挥作用，调节其 功能。基于我们的初步数据和已发表的数据，我们提出了一个新的假设，即KSR2调节成骨细胞 通过调节mTORC1信号及其下游的低氧信号，促进分化和骨形成。在这 竞争续订RO1申请，我们将对KSR2行动的这个模型进行如下检验：1)检验假设 KSR2以细胞自主的方式调节骨小梁的形成，我们将产生成骨细胞特异性的KSR2 条件性基因敲除小鼠及其骨骼表型的显微CT、组织学、力学测试和基因分析 表情。2)为了验证KSR2对成骨细胞的作用是通过其对mTORC1信号的调节而介导的假设， 我们将通过评估KSR2/mTORC1途径相互作用的功能结果来评估是否敲除 成骨细胞中的Raptor取消了KSR2基因敲除的成骨细胞增加的合成代谢功能。3)检验假设 KSR2/mTORC1对成骨细胞的影响是通过HIF1α信号转导的，我们将评估缺氧信号的变化。 在KSR2和/或Raptor条件中断的成骨细胞中。我们还将确定HIF1α表达的中断 在成骨细胞特异性KSR2条件性基因敲除小鼠中拯救骨骼表型。成功完成 将提供有关KSR2调节成骨细胞和骨的途径/S的重要信息 骨髓脂肪细胞的功能，并可能有助于更好地理解肥胖和高骨的联系机制 大量的表型。

Abstract In an increasingly aging and obese population, type 2 diabetes (T2DM) and osteoporotic fractures are major public health concerns. Affecting more than 1 in 3 adults in the United States, obesity is the most important risk factor for T2DM. It is now well established that obesity and T2DM have effects on fracture risk, and fractures in T2DM are associated with greater morbidity than in the general population. Although bone density is generally known to be increased in obese and T2DM patients, the underpinning causes that contribute to increased fracture risk in T2DM/obese subjects remain to be established. Therefore, understanding the interactions of obesity, T2DM and fracture is becoming a pressing need to reduce the societal and individual costs of fracture. Our efforts to identify control molecules and their signaling pathways that delineate a link between T2DM, osteoporosis and obesity led to the exciting discovery that targeted disruption of kinase suppressor of Ras 2 (Ksr2), an obesity and T2DM gene, led to a >2-fold increase in trabecular bone mass at the distal femur of 4-month-old mice besides inducing obesity and T2DM. By contrast, disruption of a closely related Ksr1 gene had no significant effect on bone or obese phenotypes. KSR1 and 2 were discovered as scaffolding proteins that orchestrate the assembly of Raf, MEK and ERK in the canonical Ras-Raf-MAPKs pathway. While a central role for Ksr2 expressed in hypothalamus has been implicated in the control of feeding behavior and energy balance and, thereby obesity, we have new exciting preliminary data that suggest that KSR2 expressed in osteoblasts acts locally in a cell autonomous fashion to regulate its functions. Based on our preliminary data and published data, we propose a novel hypothesis that KSR2 regulates osteoblast differentiation and bone formation via regulating mTORC1 signaling and its downstream hypoxia signaling. In this competitive renewal RO1 application, we will test this model of KSR2 action as follows: 1) To test the hypothesis that KSR2 acts in a cell autonomous fashion to regulate trabecular bone formation, we will generate osteoblast-specific Ksr2 conditional knockout mice and characterize its skeletal phenotype by micro-CT, histology, mechanical testing and gene expression. 2) To test the hypothesis that KSR2 effects on osteoblasts are mediated via its regulation of mTORC1 signaling, we will determine the functional consequence of KSR2/mTORC1 pathway interactions by evaluating if knockdown of Raptor in osteoblasts abolishes the increased anabolic functions of Ksr2 knockdown osteoblasts. 3) To test the hypothesis that KSR2/mTORC1 effects on osteoblasts are mediated via HIF1α signaling, we will evaluate changes in hypoxia signaling in osteoblasts with conditional disruption of Ksr2 and/or Raptor. We will also determine if disruption of Hif1α expression in osteoblasts rescues the skeletal phenotype in osteoblast-specific Ksr2 conditional knockout mice. Successful completion of our proposed studies will provide important information on the pathway/s by which KSR2 regulates osteoblast and bone marrow adipocyte functions and could lead to improved understanding of the mechanisms linking obesity and a high bone mass phenotype.