Molecular Mechanisms of RANKL Activation in Osteoblasts

破骨细胞是一种大型多核细胞，在骨吸收过程中起重要作用。他们是 来源于造血前体，响应于许多调节因子，最 重要的是RANKL。RANKL是破骨细胞生成的必要和充分条件， 尽管诸如M-CSF、TGF β、炎性细胞因子和胰高血糖素等因子也 在与此过程相关的事件中非常重要。RANKL在许多细胞中合成， 类型，包括成骨细胞谱系。体外和体内研究均表明， RANKL mRNA表达在生理上受关键骨重塑激素的调节， PTH和1，25（OH）2D 3以及细胞因子如IL-1、TNF α、IL-6和前列腺素 前列腺素E2。这些调节剂中任何一种的异常产生和/或表达涉及 RANKL过表达，骨吸收增强和溶骨性或骨化性疾病。尽管 尽管有相当多的生理和病理学见解，但对 控制RANKL从支持细胞的转录输出。然而初步研究表明 其机制可能非常复杂。考虑到 RANKL对骨吸收的影响，我们建议定义RANKL基因的关键机制， 在分子水平上表达。目的1：定义与以下相关的分子事件： 在RANKL远端控制区（RL-1），VDR、CREB和STATS激活RANKL基因 DCR）位于小鼠RankL转录起始位点（TSS）上游76 kb处。的相关性 将使用基因改变的小鼠模型来证实该位点的活性。目标2：评估 位于小鼠RankL基因TSS上游的四个额外调控结构域的贡献 在-16、-22、-60和-69kb处。这些位点对RankL基因调控的贡献 将通过制备独特的小鼠模型来检查表达。目标3：评估 RankL基因上游区域内转录因子占据的结果是， 染色质组织、修饰和结构，并确定RNA的功能 聚合酶II，其响应于活化而特异性地出现在这些位点。这些研究将 提供了钙激素和细胞因子 调节RANKL的表达，RANKL是一种其产物对骨吸收至关重要的基因。

Osteoclasts are large, multinucleated cells that play a central role in bone resorption. They are derived from hematopoietic precursors in response to a number of regulatory factors, the most important of which is RANKL. RANKL is both necessary and sufficient for osteoclastogenesis, although factors such as M-CSF, TGFb, inflammatory cytokines and prostaglandins are also important in the events associated with this process. RANKL is synthesized in a number of cell types, including those of the osteoblast lineage. Studies both in vitro and in vivo indicate that RANKL mRNA expression is regulated physiologically by key bone remodeling hormones such as PTH and 1,25(OH)2D3 as well as by cytokines such as IL-1, TNFa, IL-6 and the prostaglandin PGE2. Aberrant production and/or expression of any one these modulators has been implicated in RANKL overexpression, enhanced bone resorption and osteolytic or osteoporotic disease. Despite considerable physiologic and pathologic insight, little is known of the molecular mechanisms that control the transcriptional output of RANKL from support cells. Initial studies, however, suggest that the mechanisms responsible are likely to be highly complex. In view of the critical impact of RANKL on bone resorption, we propose to define key mechanisms instrumental to RANKL gene expression at the molecular level. Aim 1: To define the molecular events associated with activation of the RANKL gene by VDR, CREB, and STATS at the RANKL distal control region (RL- DCR) located 76 kb upstream of the mouse RankL transcriptional start site (TSS). The relevance of this site will be confirmed using a genetically altered mouse model. Aim 2: To evaluate the contribution of four additional regulatory domains located upstream of the mouse RankL gene TSS at -16, -22, -60 and -69 kb. The contribution of these sites to the regulation of RankL gene expression will be examined through preparation of a unique mouse model. Aim 3: To assess the consequence of transcription factor occupancy within the RankL gene upstream region on chromatin organization, modification and structure, and to determine the function of RNA polymerase II which appears specifically at these sites in response to activation. These studies will provide important detail into the mechanism by which calciotropic hormones and cytokines regulate RANKL expression, a gene whose product is central to bone resorption.