Discovery of an Osteocalcin Sensing GPCR Regulating Beta-Cell Function

描述（由申请人提供）：新出现的证据支持骨骼也是一种内分泌器官的假设，通过释放成骨细胞源性激素骨钙素（OCN）来调节能量代谢。这种新的假设是有争议的，因为重要的差距仍然有待填补我们的知识的生理效应OCN在多个器官和复杂的改变，在其他激素网络引起的OCN管理。了解骨能量代谢的综合调节的关键步骤是识别和表征OCN受体的功能。我们已经发现了OcnR的主要候选者GPRC 6A，其是一种氨基酸传感GPCR，其在细胞中高度表达，并在体外和体内被重组Ocn激活。全局消融GPRC 6A以产生OcnR-/-小鼠导致类似于Ocn-/-小鼠的表型，包括葡萄糖耐受不良、循环胰岛素水平降低和胰岛素抵抗。从OcnR-/-小鼠分离的胰岛表现出葡萄糖刺激的胰岛素分泌和胰岛发育不全的异常，表明OcnR也调节胰岛素分泌和胰岛细胞质量。因此，我们建议测试以下假设：[a]该GPCR是生物学相关的OcnR，[B]它定义了将骨代谢与胰岛素分泌和细胞增殖的代谢调节联系起来的分子机制。此外，OcnR也被L-精氨酸和睾酮激活，重组Ocn调节睾丸的睾酮产生，以及肝脏、肌肉和脂肪的代谢功能，表明OcnR可能是连接多个内分泌网络的多配体受体。具体目标是：1）通过在转基因小鼠模型和离体胰岛中的组织特异性功能丧失实验来测试OcnR在胰腺细胞中的特异性功能; 2）确认OcnR介导重组Ocn在胰腺细胞中的直接作用;和3）确定Ocn的羧化和羧化不足形式对OcnR的结合动力学。总的来说，这些研究将定义OcnR的功能，以整合将骨、Ocn和其他潜在配体与骨细胞功能联系起来的生理网络，并提供有助于简化骨内分泌功能与影响骨和能量代谢的其他器官分泌的激素之间复杂的相互依赖关系的知识。

DESCRIPTION (provided by applicant): Emerging evidence supports the hypothesis that the skeleton is also an endocrine organ that regulates energy metabolism through the release of the osteoblast-derived hormone, osteocalcin (Ocn). This novel hypothesis is controversial, because important gaps remain to be filled in our knowledge of the physiological effects of Ocn in multiple organs and the complex alterations in other hormonal networks induced by Ocn administration. Key steps toward understanding the integrative regulation of energy metabolism by bone would be to identify and characterize the function of the receptor for Ocn. We have discovered a prime candidate for the OcnR, GPRC6A, an amino-acid sensing GPCR that is highly expressed in ¿-cells and is activated by recombinant Ocn in vitro and in vivo. Global ablation of GPRC6A to create OcnR-/- mice results in a phenotype that resembles Ocn-/- mice, including glucose intolerance, reductions in circulating insulin levels, and insulin resistance. Pancreatic islets isolated from OcnR-/- mice exhibit abnormalities of glucose-stimulated insulin secretion and pancreatic islet hypoplasia, suggesting that the OcnR also regulates insulin secretion and ¿-cell mass. Thus, we propose to test the hypotheses that [a] this GPCR is the biologically relevant OcnR and [b] it defines a molecular mechanism for linking bone metabolism with metabolic regulation of insulin secretion and ¿-cell proliferation. In addition, OcnR is also activated by L-arginine and testosterone, and recombinant Ocn regulates testosterone production by the testes, as well as metabolic functions of liver, muscle and fat, suggesting that OcnR may be a multi-liganded receptor connecting multiple endocrine networks. The Specific Aims are to: 1) Test the specific functions of OcnR in pancreatic ¿-cells by tissue-specific loss-of-function experiments in transgenic mouse models and in isolated pancreatic islets ex vivo; 2) Confirm that OcnR mediates the direct effects of recombinant Ocn in pancreatic ¿-cells ; and 3) Determine the binding kinetics of carboxylated and undercarboxylated forms of Ocn for OcnR. Collectively these studies will define the functions of OcnR to integrate physiological networks linking bone, Ocn and other potential ligands to ¿-cell functions and provide knowledge that will help to simplify the complex interdependency between the endocrine functions of bone and hormones secreted by other organs that affect bone and energy metabolism.