Feedback Inhibitory Mechanisms in Skeletal Development

描述（由申请人提供）：成纤维细胞生长因子（FGF）和成纤维细胞生长因子受体（FGFR）在发育中发挥重要作用，包括骨骼和颅面发育。人FGFR1、FGFR2和FGFRS中的激活突变与骨骼发育不良相关，包括颅缝早闭和侏儒综合征。这些突变影响成骨细胞和软骨细胞的增殖和分化。这些数据表明FGFR信号传导在骨骼发育中的关键作用：FGFR的信号传导必须受到严格调控才能正常发育。重要的未解决的问题是识别骨骼发育过程中FGFR激活的途径，以及这些途径如何反馈调节FGF信号传导。在果蝇和脊椎动物中的研究已经证明Sprouty（Spry）基因家族的成员是FGFR信号传导的抑制剂。Spry1、Spry2和Spry4在小鼠发育过程中的肢芽、上颌和下颌弓以及其他部位表达。逆转录病毒介导的Spry1在鸡肢芽中的过度表达导致软骨发育不良我们已经建立了一个条件性转基因小鼠模型，以探讨Spry家族成员在骨骼发育中的作用。我们发现，在颅神经嵴细胞中条件性表达Spry1会导致严重的颅面缺陷，包括鼻骨和额骨的缺失。这些小鼠在颅面原基中还表现出转录因子Msx1、Msx2和AP 2的表达大大降低。我们假设Spry在骨骼发育中的功能是维持FGF介导的增殖、分化和凋亡之间的平衡，并且Spry的表达水平决定了这种平衡。因此，我们提出了以下3个具体目标：1）检验Spry过表达抑制骨原基中FGF活性，导致整体成骨细胞发育下降的假设; 2）使用功能丧失方法研究Spry在体内成骨细胞增殖、分化和凋亡中的作用;（3）通过体外培养的颅骨细胞研究Spry对成骨细胞增殖、分化和凋亡的影响。这些研究将提供显着的洞察信号通路的负调控颅面和骨骼发育，以及如何扰动在这个反馈信号通路导致骨骼发育不良。

DESCRIPTION (provided by applicant): The fibroblast growth factors (FGFs) and the fibroblast growth factor receptors (FGFRs) play important roles in development, including skeletal and craniofacial development. Activating mutations in human FGFR1, FGFR2, and FGFRS are associated with skeletal dysplasias including craniosynostosis and dwarfing syndromes. These mutations affect the proliferation and differentiation of osteoblasts and chondrocytes. This data indicates a pivotal role for FGFR signaling in skeletal development: signaling by FGFRs must be tightly regulated for normal development. Important unresolved issues are the identification of pathways activated by FGFRs during skeletal development, and how these pathways feedback to regulate FGF signaling. Studies in Drosophila and vertebrates have demonstrated that members of the Sprouty (Spry) gene family are inhibitors of FGFR signaling. Spry1, Spry2, and Spry4 are expressed in the limb buds, and maxillary and mandibular arches, as well as other sites during mouse development. Retroviral-mediated over-expression of Spry1 in chick limb buds results in chondrodysplasia. We have developed a conditional transgenic mouse model to explore the role of Spry family members in skeletal development. We show that conditional expression of Spry1 in cranial neural crest cells results in severe craniofacial defects including the absence of the nasal and frontal bones. These mice also exhibit greatly reduced expression of the transcription factors Msx1, Msx2, and AP2 in craniofacial primordia. We hypothesize that the function of Spry in skeletal development is to maintain a balance between FGF-mediated proliferation, differentiation, and apoptosis, and that the level of Spry expression determines this balance. Accordingly, we propose the following 3 specific aims: 1) to test the hypothesis that over- expression of Spry inhibits FGF activity in bone primordia, leading to decreased overall osteoblast development; 2) to use loss-of-function approaches to investigate the role of Spry in osteoblast proliferation, differentiation, and apoptosis in vivo; and 3) to characterize the mechanisms by which Spry affects osteoblast proliferation, differentiation, and apoptosis using calvarial cultures in vitro. These studies will provide significant insight into the negative regulation of signaling pathways in craniofacial and skeletal development and how perturbations in this feedback signaling pathway lead to skeletal dysplasias.