NUCLEAR EVENTS IN PTH ACTION ON BONE CELLS

甲状旁腺激素(PTH)是钙稳态的重要调节者，也具有 骨骼合成代谢激素。甲状旁腺素诱导大鼠成纤维细胞基质金属蛋白酶-13基因转录 成骨细胞通过cAMP依赖的途径需要从头合成蛋白质，即这是一种 次要事件。我们已经确定PTH反应元件为矮小结构域和激活子 基质金属蛋白酶-13启动子上的蛋白-1结合部位。我们已经证明了一种依赖于PTH的合作 这些位点与结合它们的蛋白质(Runx2和Fos/Jun)之间的相互作用。我们现在有了 证实了基质金属蛋白酶-13启动子中的这两个位置位于靠近TATA盒的单个核小体中 甲状旁腺素可以松弛和修饰核小体，但不会引起核小体解离。放松的是 作为早期事件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 (a组蛋白乙酰转移酶，HAT)活性，导致Runx2的磷酸化，p300被招募到 启动子导致组蛋白H4的早期位置特异性乙酰化；在晚期事件中，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，与整个近端启动子结合，导致组蛋白H3乙酰化 核小体的松动。然后是基质金属蛋白酶-13基因的最大转录。我们的假设是 该基因处于抑制状态，甲状旁腺素引起染色质的逐步修饰 组蛋白脱乙酰酶和共抑制蛋白的解离及HATS与激活剂的结合 蛋白质。这项工作的长期目标是描述将PTH行动传递到 成骨细胞中基质金属蛋白酶-13基因转录的调控。因此，这一行动的具体目标是 相互竞争的延续建议是：1)调查基质金属蛋白酶-13研发部位的早期事件 启动子，以及，2)研究AP-1位点的晚期事件以及Fos/Jun与其他 基质金属蛋白酶-13启动子上的蛋白质。这项工作的成果将对我们了解 甲状旁腺素如何在成骨细胞功能中发挥其核作用。在这样做的同时，数据还将提供新的 钙代谢紊乱的治疗展望。 这项研究将研究一种蛋白质激素(甲状旁腺激素，PTH)是如何与 并将信号传递到细胞核，使DNA和相关蛋白改变其 参与骨骼分解的一种酶的结构和原因表达。甲状旁腺素是调节 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 正在开发替代甲状旁腺素的治疗骨质疏松症的新药。

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.