Effects of interleukin-11 on the expression of human bone sialoprotein gene
Effects of interleukin-11 on the expression of human bone sialoprotein gene
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DOI：
10.1007/s00774-014-0576-8
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发表时间：
2015-03-01
影响因子：
3.3
通讯作者：
Ogata, Yorimasa
中科院分区：
文献类型：
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作者：
Matsumura, Hiroyoshi;Nakayama, Yohei;Ogata, Yorimasa
白细胞介素-11（IL-11）是一种来源于骨髓基质成纤维细胞的细胞因子，在不同的生物系统中具有广泛的活性。IL-11和IL-6是两种已知依赖于成骨细胞-破骨细胞通讯来影响破骨细胞分化的细胞因子。骨涎蛋白（BSP）是一种矿化的结缔组织特异性蛋白，在分化的成骨细胞、成牙本质细胞和成牙骨质细胞中表达。为了确定IL-11对人BSP基因转录调控的分子基础，我们进行了实时聚合酶链反应（PCR）、与荧光素酶报告基因连接的人BSP基因启动子嵌合构建体的瞬时转染分析、凝胶迁移率变动分析和使用人成骨细胞样Saos 2细胞的染色质免疫沉淀分析。IL-11（20 ng/ml）在成骨样Saos 2细胞中6 h时增加BSP、Runx 2和Osterix mRNA水平，12 h时增加碱性磷酸酶（ALP）mRNA水平。在瞬时转染试验中，IL-11（20 ng/ml，12 h）增加了包含人BSP基因启动子的构建体在-60 LUC和-868 LUC之间的荧光素酶活性。在cAMP反应元件1（CRE 1，-72至-79）和CRE 2（-667至-674）中包含2-bp突变的构建体中，IL-11的转录刺激被部分抑制。当在-868LUC中成对地进行CRE 1和CRE 2突变时，IL-11对荧光素酶活性的影响几乎完全消除。IL-11诱导的转录活性被蛋白激酶A、酪氨酸激酶、ERK 1/2和PI 3-激酶抑制剂抑制。凝胶迁移率变化分析表明，IL-11增加核蛋白结合CRE 1和CRE 2。CREB 1、磷酸化CREB 1、c-Fos和c-Jun抗体破坏CRE 1和CRE 2蛋白复合物的形成。这些数据表明IL-11通过人BSP基因启动子中的CRE 1和CRE 2元件刺激BSP基因转录。
Interleukin-11 (IL-11) is a bone marrow stromal fibroblast-derived cytokine with a wide spectrum of activities in different biological systems. IL-11 and IL-6 are two cytokines known to rely on osteoblast-osteoclast communication for their effects on osteoclast differentiation. Bone sialoprotein (BSP) is a mineralized connective tissue-specific protein expressed in differentiated osteoblasts, odontoblasts, and cementoblasts. To determine the molecular basis of the transcriptional regulation of the human BSP gene by IL-11, we conducted real-time polymerase chain reactions (PCR), transient transfection analyses with chimeric constructs of the human BSP gene promoter linked to a luciferase reporter gene, gel mobility shift assays, and a chromatin immunoprecipitation assay using human osteoblast-like Saos2 cells. IL-11 (20 ng/ml) increased BSP, Runx2, and Osterix mRNA levels at 6 h and the alkaline phosphatase (ALP) mRNA level at 12 h in osteoblast-like Saos2 cells. In a transient transfection assay, IL-11 (20 ng/ml, 12 h) increased luciferase activities of constructs between -60LUC and -868LUC including the human BSP gene promoter. Transcriptional stimulations by IL-11 were partially inhibited in the constructs that included 2-bp mutations in the cAMP response element 1 (CRE1, -72 to -79) and CRE2 (-667 to -674). When mutations were made in pairs of CRE1 and CRE2 in -868LUC, the effect of IL-11 on luciferase activity was almost totally abrogated. Transcriptional activities induced by IL-11 were inhibited by protein kinase A, tyrosine kinase, ERK1/2, and PI3-kinase inhibitors. Gel mobility shift analyses showed that IL-11 increased nuclear proteins binding to CRE1 and CRE2. CREB1, phospho-CREB1, c-Fos, and c-Jun antibodies disrupted the formation of CRE1 and CRE2 protein complexes. These data demonstrate that IL-11 stimulates BSP gene transcription via CRE1 and CRE2 elements in the human BSP gene promoter.