A Novel Distal Enhancer Mediates Cytokine Induction of Mouse Rankl Gene Expression
A Novel Distal Enhancer Mediates Cytokine Induction of Mouse Rankl Gene Expression
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DOI：
10.1210/me.2009-0209
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发表时间：
2009-12-01
影响因子：
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通讯作者：
Pike, J. Wesley
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文献类型：
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作者：
Bishop, Kathleen A.;Meyer, Mark B.;Pike, J. Wesley
慢性炎症状态与骨丢失增加相关。这种增加通常与核因子-κ B配体（RANKL）受体激活剂（一种破骨细胞形成所必需的TNF α样因子）的升高有关。在这项研究中，我们记录了IL-6与IL-6可溶性受体（IL-6/IL-6sR）和制瘤素M组合通过信号转导子和转录激活子3（STAT 3）诱导基质细胞中Rankl表达的能力。我们使用染色质免疫沉淀-平铺DNA微阵列分析，以确定在Rankl位点的STAT 3的作用位点，并评估结合组蛋白H4乙酰化和RNA聚合酶II募集的后果。IL-6/IL-6可溶性受体和制瘤素M均刺激Rankl转录起始位点上游的STAT 3结合。尽管先前鉴定的增强子结合STAT 3，但称为mRLD 6的更远增强子是STAT 3结合的特定焦点。当与异源启动子融合时，该增强子具有高度活性，含有两个功能活性STAT应答元件。重要的是，Stat 3 mRNA和蛋白质的小干扰RNA敲低，而不是Stat 1或Stat 5a的小干扰RNA敲低，在限制Rankl mRNA上调中是有效的。有趣的是，尽管RNA聚合酶II和组蛋白H4乙酰化标记了基础条件下的许多增强子，但两者的水平在细胞因子处理后强烈增加，特别是在mRLD 6。最后，mRLD 6也是毛喉素诱导的细胞反应元件结合蛋白（CREB）募集的靶点，这增强了细胞因子活性。我们的研究为糖蛋白130激活细胞因子诱导RANKL表达的机制提供了新的见解。（分子内分泌学23：2095-2110，2009）
Chronic inflammatory states are associated with increased bone loss. This increase is often linked to an elevation in receptor activator of nuclear factor-kappa B ligand (RANKL), a TNF alpha-like factor essential to osteoclast formation. In this study, we document the ability of IL-6 in combination with IL-6 soluble receptor (IL-6/IL-6sR) and oncostatin M to induce Rankl expression in stromal cells via signal transducer and activator of transcription 3 (STAT3). We used chromatin immunoprecipitation-tiled DNA microarray analysis to determine sites of action of STAT3 at the Rankl locus and to assess the consequences of binding on histone H4 acetylation and RNA polymerase II recruitment. Both IL-6/IL-6 soluble receptor and oncostatin M stimulated STAT3 binding upstream of the Rankl transcriptional start site. Although previously identified enhancers bound STAT3, a more distal enhancer termed mRLD6 was a particular focus of STAT3 binding. When fused to a heterologous promoter, this enhancer was highly active, containing two functionally active STAT response elements. Importantly, small interfering RNA knockdown of Stat3 mRNA and protein, but not that of Stat1 or Stat5a, was effective in limiting Rankl mRNA up-regulation. Interestingly, although RNA polymerase II and histone H4 acetylation marked many of the enhancers under basal conditions, the levels of both were strongly increased after cytokine treatment, particularly at mRLD6. Finally, mRLD6 was also a target for forskolin-induced cellular response element-binding protein (CREB) recruitment, which potentiated cytokine activity. Our studies provide new insight into mechanisms by which glycoprotein 130 activating cytokines induce RANKL expression. (Molecular Endocrinology 23: 2095-2110, 2009)