BONE SIALOPROTEIN: EXPRESSION & ROLE IN MINERALIZATION

说明(申请人提供)：骨涎蛋白(BSP)是一种 骨的非胶原性细胞外基质成分 与初始基质矿化和骨重建有关。BSP基因 一直是项目实验室用来理解 组织特异性基因在骨骼中表达的机制。BSP也很有用 以了解生物矿化的机制。在之前的资助期间 期间，我们分离到一个2.5kb的小鼠BSP基因启动子，含有足够的 指导成骨细胞选择性表达的信息在细胞培养和 转基因小鼠。在这个启动子中，我们确定了一个同源结构域 成骨细胞选择性表达所需的蛋白结合位点 BSP在细胞培养中的表达。该位点及相关位点在体内的作用 BSP的表达尚不清楚。该元件的转录活性 可以被与骨骼相关的同源结构域蛋白刺激，如DLX-5， 但我们不知道与这个位点调控的蛋白/S的身份 活着。相当多的体外证据支持BSP是一种 羟基磷灰石晶体的成核剂。我们证明了它也能刺激 矿化时在非矿化前成骨细胞系中过表达。 相比之下，基质GLA蛋白(MGP)是一种强有力的矿化抑制因子。 PTH和PTHrP都是成骨细胞介导的生物矿化的有效抑制剂， 抑制BSP的合成，同时快速诱导MGP，表明这些 分子具有相反的作用，并且相互调节。 在这些重要发现的基础上，这次竞争性更新将实现 以下具体目标：目标1.确定一种病毒在体内的意义 小鼠BSP启动子中的同源结构域蛋白结合部位。目标2.确定 与近端同源域相互作用和调节的核蛋白(S) 成骨细胞中的位置。目的3.确定MGP调控的分子基础 通过甲状旁腺激素和甲状旁腺素rP，评估BSP和MGP在体内相互拮抗的能力。 细胞培养和体内实验。 该项目支持的研究将描述一种新的机制 调控基因在骨中的表达并解释PTH/PTHrP的重要作用 论成矿规律。这些知识对于设计是必不可少的。 骨再生治疗创伤和病理的新策略 例如骨质疏松症和牙周病。

DESCRIPTION (provided by applicant): Bone sialoprotein (BSP) is a noncollagenous extracellular matrix component of the bone that is intimately associated with initial matrix mineralization and bone remodeling. The Bsp gene has been an important tool used by the project laboratory to understand mechanisms of tissue-specific gene expression in bone. BSP has also been useful for understanding mechanisms of biomineralization. During the previous funding period, we isolated a 2.5 kb murine Bsp gene promoter containing sufficient information to direct osteoblast-selective expression both in cell culture and transgenic mice. Within this promoter, we identified a homeodomain protein-binding site that is required for the osteoblast-selective expression of Bsp in cell culture. The role of this and related sites in the in vivo expression of Bsp is not known. The transcriptional activity of this element can be stimulated by homeodomain proteins associated with bone such as Dlx-5, but we do not know the identity of the protein/s regulating with this site in vivo. Considerable in vitro evidence supports the hypothesis that BSP is a nucleator of hydroxyapatite crystals. We showed that it can also stimulate mineralization when overexpressed in non-mineralizing preosteoblast cell lines. Matrix Gla protein (MGP), in contrast, is a potent inhibitor of mineralization. PTH and PTHrP, both potent inhibitors of osteoblast-mediated biomineralization, inhibit BSP synthesis while rapidly inducing MGP, suggesting that these molecules have opposing actions and are reciprocally regulated. Building on these important findings, this competitive renewal will achieve the following specific aims: Aim 1. Define the in vivo significance of a homeodomain protein-binding site in the murine BSP promoter. Aim 2. Identify the nuclear protein(s) that interact with and regulate the proximal homeodomain site in osteoblast. Aim 3. Define the molecular basis for the regulation of MGP by PTH and PTHrP; assess the ability of BSP and MGP to antagonize each other in cell culture and in vivo. Studies supported by this project will describe a novel mechanism for controlling gene expression in bone and explain important actions of PTH/PTHrP on the regulation of mineralization. Such knowledge is essential for designing new strategies for bone regeneration to treat trauma and pathological conditions such as osteoporosis and periodontal disease.