Progressive recruitment of Runx2 to genomic targets despite decreasing expression during osteoblast differentiation.
Progressive recruitment of Runx2 to genomic targets despite decreasing expression during osteoblast differentiation.
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DOI：
10.1002/jcb.21900
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发表时间：
2008-11-01
影响因子：
4
通讯作者：
Frenkel, Baruch
中科院分区：
文献类型：
--
作者：
Pregizer, Steven;Baniwal, Sanjeev K.;Yan, Xiting;Brook, Zea;Frenkel, Baruch
编码成骨细胞主要转录因子Runx2及其靶基因骨钙素(OC)的mRNAs通常被用作成骨细胞分化的标志。我们发现，虽然在MC3T3-E1培养的成骨细胞表型发育过程中，OC的mRNA水平确实增加了，但Runx2的mRNA水平却意外地下降。无论是对Runx2的翻译控制(基于Western分析)，还是对其DNA结合能力的调节(通过凝胶迁移率改变分析)，都不能解释Runx2和OC表达的意外相反模式。相反，成骨细胞分化过程中的一系列芯片分析显示，在早期，当Runx2蛋白含量和DNA结合活性最大时，OC启动子几乎不存在。在后期，Runx2被招募到OC启动子，而Runx2的mRNA、蛋白质和体外DNA结合逐渐减少。我们还跟踪了Runx2在一个新的基因组靶点上的占据情况，这个新的基因组靶点是通过对细胞的芯片分析发现的，其中OC启动子占据了最大的位置。结果表明，Runx2被招募到该基因座和OC启动子上，具有非常相似的时间模式。这些观察结果突显了一种机制，即通过将Runx2对基因组靶标的访问限制在狭窄的时间窗口来抑制Runx2介导的转录控制。需要如此严格的控制与Runx2在体内过度表达的严重后果是一致的。
The mRNAs encoding Runx2, a master osteoblast transcription factor, and its target gene Osteocalcin (OC), are commonly used as markers of osteoblast differentiation. We found that while OC mRNA levels do indeed increase during development of the osteoblast phenotype in MC3T3-E1 cultures, Runx2 mRNA levels surprisingly decrease. Neither translational control of Runx2 (based on Western analysis) nor regulation of its DNA-binding ability (assessed by electrophoretic mobility shift assay) could explain the unexpected opposite patterns of Runx2 and OC expression. Instead, a series of ChIP assays during osteoblast differentiation revealed that early on, when Runx2 protein amount and DNA-binding activity are maximal, it is practically absent from the OC promoter. At later stages, Runx2 is recruited to the OC promoter while Runx2 mRNA, protein, and in vitro DNA binding progressively decrease. We also followed Runx2 occupancy at a novel genomic target discovered by ChIP-Chip analysis of cells in which the OC promoter is maximally occupied. The results revealed that Runx2 is recruited to this locus and to the OC promoter with a remarkably similar temporal pattern . These observations highlight a mechanism that restrains Runx2-mediated transcriptional control by confining its access to genomic targets to a narrow window of time. The need for such stringent control is consistent with the severe consequences of Runx2 over-expression in vivo.
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