NUCLEAR EVENTS IN PTH ACTION ON BONE CELLS

甲状旁腺激素 (PTH) 是钙稳态的重要调节剂，也具有调节钙稳态的作用。 骨骼合成代谢激素。 PTH 诱导基质金属蛋白酶-13 (MMP-13) 基因转录 成骨细胞通过 cAMP 依赖性途径需要从头合成蛋白质，即，这是一个 次要事件。我们已将 PTH 响应元件识别为矮域和激活子 MMP-13 启动子中的 Protein-1 结合位点。我们已经展示了 PTH 依赖的合作 位点和与其结合的蛋白质（Runx2 和 Fos/Jun）之间的相互作用。我们现在有 确定 MMP-13 启动子中的这两个位点位于靠近 TATA 盒的单个核小体中 PTH 可以松弛并修饰该核小体，但不会导致其解离。放松是 作为早期和晚期事件发生的几个步骤的产物：在早期事件中，首先，PTH 刺激 p300 （一种组蛋白乙酰转移酶，HAT）活性，导致 Runx2 磷酸化，并且招募 p300 导致组蛋白 H4 早期位点特异性乙酰化的启动子；在后期事件中，新合成 Fos/Jun 与启动子结合，然后似乎与 Runx2 相互作用，随后招募 CBP （CREB 结合蛋白），另一种 HAT，作用于整个近端启动子和组蛋白 H3 的乙酰化 在核小体的松弛中。 MMP-13 基因的最大转录随之而来。我们的假设是 尽管该基因处于抑制状态，但 PTH 会导致染色质的逐步修饰 组蛋白脱乙酰酶和共阻遏蛋白的解离以及 HAT 和激活剂的关联 蛋白质。这项工作的长期目标是描述将 PTH 作用传递给 成骨细胞中 MMP-13 基因转录的调控。因此，本次会议的具体目标 竞争的继续提案是，1）调查 MMP-13 RD 站点的早期事件 启动子，并且，2) 研究 AP-1 位点的晚期事件以及 Fos/Jun 与其他基因的相互作用 MMP-13 启动子上的蛋白质。这项工作的成果将对我们的知识做出重大贡献 PTH 如何对成骨细胞功能发挥其核作用。这样做，数据还将提供新的 钙代谢紊乱治疗的观点。 这项研究将调查蛋白质激素（甲状旁腺激素，PTH）如何与 细胞表面并向细胞核传递信号，导致 DMA 和相关蛋白改变其 参与骨分解的酶的结构和引起的表达。 PTH 对于调节至关重要 血清钙水平，也被用于治疗骨质疏松症。我们的研究结果可能会导致 正在开发新药来代替 PTH，治疗骨质疏松症。

Parathyroid hormone (PTH) is an essential regulator of calcium homeostasis and also has a role as an anabolic hormone for bone. PTH induces matrix metalloproteinase-13 (MMP-13) gene transcription in osteoblastic cells through a cAMP-dependent pathway requiring de novo protein synthesis, i.e., this is a secondary event. We have identified the PTH-response elements as being the runt domain and the activator protein-1 binding sites in the MMP-13 promoter. We have demonstrated a PTH-dependent cooperative interaction between the sites and the proteins (Runx2 and Fos/Jun) binding to them. We have now established that these two sites in the MMP-13 promoter are in a single nucleosome close to the TATA box and PTH relaxes and modifies this nucleosome but does not cause its dissociation. The relaxation is the product of several steps which occur as early and late events: In the early events, first, PTH stimulates p300 (a histone acetyltransferase, HAT) activity, causes the phosphorylation of Runx2, and p300 is recruited to the promoter resulting in early site-specific acetylation of histone H4; in the late events, newly synthesized Fos/Jun associate with the promoter and then appear to interact with Runx2 followed by recruitment of CBP (CREB binding protein), another HAT, to the entire proximal promoter and acetylation of histone H3 resulting in loosening of the nucleosome. Maximal transcription of the MMP-13 gene then ensues. Our hypothesis is that the gene is in a repressed state and PTH causes the stepwise modification of the chromatin though dissociation of histone deacetylases and co-repressor proteins and association of HATs and activator proteins. The long-term goals of this work are to delineate the mechanisms conveying PTH action to regulation of transcription of the MMP-13 gene in osteoblasts. Consequently, the specific aims of this competing continuation proposal are to, 1) investigate the early events at the RD site of the MMP-13 promoter, and, 2) investigate the late events at the AP-1 site and the interaction of Fos/Jun with other proteins on the MMP-13 promoter. The results of this work will make major contributions to our knowledge of how PTH exerts its nuclear effects on osteoblast function. In so doing, the data will also provide new perspectives into treatment of disorders of calcium metabolism. This research will investigate how a protein hormone (parathyroid hormone, PTH) is able to interact with a cell's surface and transmit signals to the nucleus to cause the DMAand associated proteins to change their structure and cause expression of an enzyme involved in bone breakdown. PTH is essential for regulating serum calcium levels, and is also being used to treat osteoporosis. The results of our research could lead to new drugs being developed, in place of PTH, to treat osteoporosis.