Department of Orthopaedics, University of Maryland School of Medicine, Baltimore, MD, United States.
Biochem Biophys Res Commun. 2019 Feb 12;509(3):728-733. doi: 10.1016/j.bbrc.2018.12.173. Epub 2019 Jan 7.
In bone, connexin43 expression in cells of the osteoblast lineage plays an important role in restraining osteoclastogenesis and bone resorption. While there is a consensus around the notion that the anti-osteoclastogenic factor, osteoprotegerin, is a driver of this effect, how connexin43 regulates osteoprotegerin gene expression is unclear. Here, we show that loss of connexin43 decreased osteoprotegerin gene expression and reduced ERK1/2 activation. Conversely, overexpression of connexin43 increased osteoprotegerin expression and enhanced ERK1/2 activation. This increase in phospho-ERK1/2 is required for connexin43 to induce transcription from the osteoprotegerin proximal promoter. Connexin43 increased promoter activity via a specific 200 base pair region of the osteoprotegerin promoter located at -1486 to -1286 with respect to the transcriptional start site, a region which includes four Sp1 binding elements. Further, activation of this promoter region required an intact functional connexin43, as hypomorphic or dominant negative connexin43 mutant constructs, including one with increased hemichannel activity, were unable to stimulate osteoprotegerin expression as strongly as wild type connexin43. Using chromatin immunoprecipitations, we show that connexin43 expression enhanced the recruitment of Sp1, but not Runx2, to the osteoprotegerin proximal promoter. In total, these data show that connexin43-dependent gap junctional communication among osteoblast cells permits efficient ERK1/2 activation. ERK1/2 signaling promotes the recruitment of the potent transcriptional activator, Sp1, to the osteoprotegerin proximal promoter, resulting in robust transcription of anti-osteoclastogenic factor, osteoprotegerin.
在骨骼中，成骨细胞谱系细胞中的连接蛋白 43 的表达在抑制破骨细胞生成和骨吸收方面发挥着重要作用。虽然人们普遍认为抗破骨细胞因子骨保护素是产生这种作用的驱动因素，但连接蛋白 43 如何调节骨保护素基因的表达尚不清楚。在这里，我们发现缺失连接蛋白 43 会降低骨保护素基因的表达并减少 ERK1/2 的激活。相反，过表达连接蛋白 43 会增加骨保护素的表达并增强 ERK1/2 的激活。这种磷酸化 ERK1/2 的增加是连接蛋白 43 诱导骨保护素近端启动子转录所必需的。连接蛋白 43 通过骨保护素启动子的一个特定的 200 个碱基对区域增加启动子活性，该区域相对于转录起始位点位于-1486 到-1286 ，包含四个 Sp1 结合元件。此外，该启动子区域的激活需要一个完整功能的连接蛋白 43，因为低功能或显性负性连接蛋白 43 突变体构建体，包括一个具有增强的半通道活性的构建体，不能像野生型连接蛋白 43 那样强烈地刺激骨保护素的表达。通过染色质免疫沉淀，我们发现连接蛋白 43 的表达增强了 Sp1 的募集，但不是 Runx2，到骨保护素近端启动子。总的来说，这些数据表明连接蛋白 43 依赖性的成骨细胞细胞之间的缝隙连接通讯允许有效的 ERK1/2 激活。ERK1/2 信号促进了强有力的转录激活因子 Sp1 募集到骨保护素近端启动子，从而导致抗破骨细胞因子骨保护素的强大转录。
