Characterization of the structure and function of DMP2

骨骼和牙齿的适当矿化对包括肌肉骨骼功能在内的正常人体生长发育具有重要意义。这些组织中的矿物相有助于结构的硬度和综合强度，并且作为钙和镁的代谢储存库具有重要的生理作用。矿化过程中的问题在许多骨骼疾病中都很明显。矿化组织研究中最有趣的问题之一是，在生理环境中，循环的钙、磷酸盐和其他矿相离子如何集中在特定的、局部的器官或组织中。到目前为止，我们对这个级联是如何启动和调节的过程几乎没有定义。然而，有机基质在矿化过程中起着重要的调节作用。我们主要把精力集中在牙本质矿化上，因为它是一个比骨骼更简单的系统，但这两种机制可能密切相关。我们对基质介导的矿化的基本假设是，酸性大分子首先结合在胶原基质内，这些大分子负责成核并启动矿化级联。可能这些酸性大分子也调节碳酸羟基磷灰石晶体的大小。在寻找牙本质主要酸性非胶原蛋白（NCP）磷酸化蛋白（PP）的基因编码过程中，我们发现了2个克隆，一个代表磷酸化蛋白，现在被命名为DMP2（牙本质基质蛋白2），另一个代表DMP3（牙本质基质蛋白3），它是牙本质唾液蛋白和一个微型“磷酸化蛋白”样结构域的化合物。从部分DMP2 cDNA中推断出的氨基酸序列是特别有趣的，因为它清楚地代表了一种富含天冬氨酸和丝氨酸的酸性蛋白质，这种蛋白质是牙本质基质成分的预期类型。该基因紧密定位于小鼠染色体5q21，相当于人类染色体4Q21。这条染色体的位置特别有趣，因为人类染色体4q13-21与牙本质矿化障碍牙本质发育不全II型有联系。为了扩大我们对DMP2基因结构及其功能的认识，我们提出以下具体目标：(1)确定大鼠DMP2的完整初级结构(2)确定DMP2启动子序列并确定参与组织特异性调控的元件(3)克隆人类DMP2基因，最终确定牙本质发育不全II型患者的基因改变(4)研究DMP2在牙齿发育过程中的时空表达模式(5)确定DMP2的钙结合特性。长期目标是了解DMP2在牙本质矿化中的调控机制。

The proper mineralization of bones and teeth has great importance in normal human growth and development including musculo-skeletal functions. The mineral phase in these tissues contributes to the hardness and comprehensive strength of the structure and also has a major physiological role as the metabolic reservoir of calcium and magnesium. Problems in the mineralization process are evident in a number of skeletal pathologies. One of the most interesting questions in mineralized tissue research is how, within the physiological environments, circulating calcium and phosphate and other mineral phase ions can be concentrated in specific, localized organs or tissues. Up until this point we have defined little about the process by which this cascade is initiated and regulated. However, the organic matrix has been implicated to have a major role in regulating the mineralization process. We have focussed our efforts primarily on dentin mineralization because it is a simpler system than bone, but the two mechanisms are probably closely related. Our basic hypothesis for matrix-mediated mineralization, is that acidic macromolecules first bind within the collagen matrix and these are responsible for nucleating and starting the mineralization cascade. Probably these acidic macromolecules also regulate the size of carbonated hydroxyapatite crystals. In the search for the gene encoding for the principle acidic noncollagenous protein (NCP) of dentin, namely phosphophoryn (PP) we identified 2 clones one representing phosphophoryn now named DMP2 (dentin matrix protein 2) and the other DMP3 (dentin matrix protein 3) which is a compound of dentin sialoprotein and a mini "phosphophoryn"like domain. The amino acid sequence deduced from the partial DMP2 cDNA is of special interest because it clearly represents an aspartic acid and serine rich acidic protein of the type to be expected of a dentin matrix component. This gene has been tightly localized to mouse chromosome 5q21, equivalent to human chromosome 4Q21. This chromosome location is especially interesting because of the linkage of human chromosome 4q13-21 with the dentin mineralization disorder dentinogenesis imperfecta type II. In order to expand our understanding of the DMP2 gene structure and its function we propose the following specific aims: (1) To determine the complete primary structure of rat DMP2 (2) To delineate DMP2 promoter sequences and identify elements involved in tissue -specific regulation (3) To clone the human DMP2 gene in order to ultimately identify gene alterations in patients with Dentinogenesis Imperfecta Type II (4) To examine the temporal and spatial patterns of DMP2 expression during tooth development (5) To determine the calcium binding property of DMP2. The long-term goal is to understand the regulatory mechanism of DMP2 in dentin mineralization.