CCAAT/enhancer-binding proteins (C/EBP) β and δ activate osteocalcin gene transcription and synergize with Runx2 at the C/EBP element to regulate bone-specific expression
CCAAT/enhancer-binding proteins (C/EBP) β and δ activate osteocalcin gene transcription and synergize with Runx2 at the C/EBP element to regulate bone-specific expression
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DOI：
10.1074/jbc.m106611200
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发表时间：
2002-01-11
影响因子：
4.8
通讯作者：
Lian, JB
中科院分区：
文献类型：
--
作者：
Gutierrez, S;Javed, A;Lian, JB
CCAAT/增强子结合蛋白 (C/EBP) 是细胞分化和细胞类型特异性基因表达的关键决定因素。它们在成骨细胞发育中的功能作用尚未确定。我们研究了 C/EBP 因子在成骨细胞分化过程中调节组织特异性基因转录的机制的关键组成部分。 C/EBPbeta 和 C/EBPdelta 的表达从生长到成熟发育阶段都会增加，并且与骨特异性骨钙素 (OC) 基因一样，也会受到成骨细胞分化调节剂维生素 D(3) 的刺激 3-6 倍。我们鉴定了大鼠骨钙素基因近端启动子中的 C/EBP 增强子元件，该元件与 Runx2 (Cbfa1) 元件非常接近，对于组织特异性激活至关重要。我们发现 C/EBP 和 Runx2 因子以协同方式相互作用，增强细胞培养系统中的 OC 转录（35-40 倍）。我们通过突变分析表明，这种协同作用是通过 OC 启动子中的 C/EBP 响应元件以及 Runx2 和 C/EBPbeta 之间的直接相互作用介导的。此外，我们通过免疫沉淀在 Runx2 中绘制了这种相互作用所需的结构域。缺乏此相互作用结构域的 Runx2 突变体不会表现出功能协同作用。我们得出的结论是，除了Runx2 DNA结合功能外，Runx2还可以在C/EBP位点形成蛋白质复合物来调节转录。综上所述，我们的研究结果表明，C/EBP 是 OC 基因的主要反式激活因子，并且与 Runx2 的协同作用表明这些因子的组合相互作用是在成骨细胞分化过程中调节组织特异性表达的主要机制。
CCAAT/enhancer-binding proteins (C/EBP) are critical determinants for cellular differentiation and cell type-specific gene expression. Their functional roles in osteoblast development have not been determined. We addressed a key component of the mechanisms by which C/EBP factors regulate transcription of a tissue-specific gene during osteoblast differentiation. Expression of both C/EBPbeta and C/EBPdelta increases from the growth to maturation developmental stages and, like the bone-specific osteocalcin (OC) gene, is also stimulated 3-6-fold by vitamin D(3), a regulator of osteoblast differentiation. We characterized a C/EBP enhancer element in the proximal promoter of the rat osteocalcin gene, which resides in close proximity to a Runx2 (Cbfa1) element, essential for tissue-specific activation. We find that C/EBP and Runx2 factors interact together in a synergistic manner to enhance OC transcription (35-40-fold) in cell culture systems. We show by mutational analysis that this synergism is mediated through the C/EBP-responsive element in the OC promoter and by a direct interaction between Runx2 and C/EBPbeta. Furthermore, we have mapped a domain in Runx2 necessary for this interaction by immunoprecipitation. A Runx2 mutant lacking this interaction domain does not exhibit functional synergism. We conclude that, in addition to Runx2 DNA binding functions, Runx2 can also form a protein complex at C/EBP sites to regulate transcription. Taken together, our findings indicate that C/EBP is a principal transactivator of the OC gene and the synergism with Runx2 suggests that a combinatorial interaction of these factors is a principal mechanism for regulating tissue-specific expression during osteoblast differentiation.