Role of Gsalpha Signaling in Osteoblasts

描述(申请人提供)：GSA是一种普遍存在的G蛋白，是一种亚单位，在包括PTH/PTHrP受体(PPR)在内的各种受体的刺激下激活腺苷环化酶。GSA的失活和激活突变都会对骨骼发育产生影响。为了确定GSA信号在体内成骨细胞发育中的作用，我们通过将GSA诱导的小鼠与Osterix启动子-Cre重组酶转基因小鼠交配，有条件地删除了早期成骨细胞前体中的GSA。早期成骨细胞特异性失活的GSA(BGsaKO)小鼠在出生后第1天出现明显的骨骼脆性并出现大量骨折。BGsaKO小鼠的长骨表现出严重的骨小梁和皮质骨减少。具体目的：在体内评价成骨细胞前体GSA消融对成骨细胞分化、骨形成和骨吸收的生理影响。在特定目标II中，重点将是从BGsaKO小鼠中分离骨髓基质细胞。这些细胞可以用来探索调控成骨细胞发育的信号通路，特别是研究甲状旁腺素是否需要GSA来依赖PKA刺激成骨细胞特异性基因的表达。

DESCRIPTION (provided by applicant): Gsa is a ubiquitous G protein a subunit which activates adenylyl cyclase upon stimulation of a variety of receptors including the PTH/PTHrP receptor (PPR). Both inactivating and activating mutations in Gsa have consequences in skeletal development. To determine the role of Gsa signaling during osteoblast development in vivo, we have conditionally deleted Gsa in early osteoblast precursors by mating Gsa-floxed mice with osterix promoter-Cre recombinase transgenic mice. Mice with early osteoblast-specific inactivation of Gsa (BGsaKO) have marked skeletal fragility and exhibit numerous fractures by postnatal day 1. Long bones from BGsaKO mice demonstrate severe reduction in trabecular and cortical bone. Specific Aim I will evaluate the physiologic effects of Gsa ablation in osteoblast precursors on osteoblast differentiation, bone formation and bone resorption in vivo. In Specific Aim II the focus will be to isolate bone marrow stromal cells from BGsaKO mice. These cells can then be used to probe the signaling pathways regulating osteoblast development, in particular to examine whether Gsa is required for the PKA-dependent stimulation of osteoblast-specific gene expression by PTH.