Molecular Mechanism of Matrix GLA Protein (MGP)

钙化在血管疾病中普遍存在，并且对发病率和死亡率有显着影响。矩阵 GLA 蛋白 (MGP) 是一种所谓的钙化抑制剂，在动脉粥样硬化病变中表达上调。删除 MGP 基因的缺失会导致小鼠和人类的动脉钙化和形态异常。 MGP 是一种分泌蛋白，可通过维生素 K 依赖性 g-羧化作用进行修饰。当被隔离时 骨头经过加工，缺乏 7 个 C 端氨基酸。 MGP 的机制尚不清楚。然而，我们的实验数据表明， 转化生长因子-fi (TGF-fi) 和骨形态发生蛋白 (BMP) 信号传导。数据表明 BMP 和 TGF-li 之间的潜在联系在于 BMP2 和 BMP-4 诱导 TGF-R 受体的表达 激活素样激酶受体 1 (ALK1)。通过 ALK1 的信号传导可诱导 MGP，从而提供负作用 BMP 的反馈调节。还观察到 MGP 和特定受体之间的相互作用。 本次拨款将测试三个假设。第一个假设是 MGP 调节 TGF-fi 和 BMP 受体水平的信号传导，导致 SMAD 信号传导和基因表达改变。我们将表征 TGF-li1、BMP-2 和 BMP-4 诱导的 SMAD 信号和相关报告基因的激活 MGP 的存在，并鉴定受 MGP 调节的 TGF-li/BMP 受体。第二个 假设 MGP 参与特定的蛋白质-蛋白质相互作用，该相互作用可能被 C 末端改变 处理。我们将使用特定的方法来表征相互作用，其中包括 TGF-ft 受体 免疫沉淀与一般蛋白质组学方法相结合。第三个假设是 MGP 在动脉发育和动脉粥样硬化病变中充当 BMP 抑制剂。我们将生成一个 用于 BMP 抑制剂 Noggin 靶向、条件表达的转基因小鼠。 Noggin 表达将是 在 MGP 缺失小鼠中诱导，试图替代 MGP 并消除钙化。 Noggin 的效果也会 在 LDL 受体缺失小鼠的动脉粥样硬化病变中进行测定。 我们的结果将增加对血管和心脏瓣膜钙化（心脏病的常见问题）的了解。 此外，它还可以提供有关如何设计针对不需要的钙化的新治疗方法的信息。

Calcification is ubiquitous in vascular disease, and contributes significantly to morbidity and mortality. Matrix GLA protein (MGP) is an alleged calcification inhibitor that is up-regulated in atherosclerotic lesions. Deletion of the MGP gene results in arterial calcification and in morphological abnormalities in mice and humans. MGP is a secreted protein that is modified by vitamin K-dependent g-carboxylation. When isolated from bone, it is processed and lacks seven C-terminal amino acids. The mechanism of MGP is poorly understood. However, our experimental data points to a role in transforming growth factor-fi (TGF-fi) and bone morphogenetic protein (BMP) signaling. The data suggest a potential link between BMP and TGF-li in that BMP2 and BMP-4 induce expression of the TGF-R, receptor Activin-like Kinase receptor 1 (ALK1). Signaling through ALK1 induces MGP, which provides negative feedback regulation for BMP. Interactions between MGP and specific receptors were also observed. Three hypotheses will be tested in this grant. The first hypothesis is that MGP regulates TGF-fi and BMP signaling at the receptor level, resulting in altered SMAD signaling and gene expression. We will characterize activation of SMAD signaling and relevant reporter genes induced by TGF-li1, BMP-2 and BMP-4 in presence of MGP, and identify the TGF-li / BMP receptors that are regulated by MGP. The second hypothesis is that MGP participates in specific protein-protein interactions that may be altered by C-terminal processing. We will characterize the interactions, which include the TGF-ft receptors, using specific immunoprecipitations in combination with a general proteomic approach. The third hypothesis is that MGP functions as a BMP inhibitor in developing arteries and in atherosclerotic lesions. We will generate a transgenic mouse for targeted, conditional expression of the BMP-inhibitor Noggin. Noggin expression will be induced in MGP null mice in attempt to replace MGP and abolish calcification. The effect of Noggin will also be determined in atherosclerotic lesions in LDL-receptor null mice. Our results will add insight to calcification of vessels and heart valves, a common problem in heart disease. In addition, it may provide information on how to design new treatments for unwanted calcification.