Shakoori A R, van Wijnen A J, Bortell R, Owen T A, Stein J L, Lian J B, Stein G S
Department of Cell Biology, University of Massachusetts Medical Center, Worcester 01655.
J Cell Biochem. 1994 Jun;55(2):218-29. doi: 10.1002/jcb.240550209.
Vitamin D responsive transcription of the bone-specific osteocalcin gene differs markedly in osteosarcoma cells and normal diploid osteoblasts. In osteoblasts the osteocalcin gene is transcribed, and upregulated by Vitamin D, only in post-proliferative cells, but in osteosarcoma cells expression is constitutive. This distinction in transcriptional regulation of the osteocalcin gene correlates with striking differences in the relative representation of two principal Vitamin D-dependent protein/DNA complexes designated V1 and V2 at the Vitamin D responsive element in the osteocalcin promoter. Formation of both complexes is Vitamin D dependent and they contain the Vitamin D receptor as well as an RXR related protein. Pore size exclusion and sedimentation velocity analyses suggest that the V1 and V2 complexes represent oligomeric protein assemblies (respectively, tetramers and trimers), and reflect primarily DNA-directed association of the monomeric protein components at the osteocalcin Vitamin D responsive element. UV crosslinking and methylation interference analyses of the V1 and V2 complexes at the osteocalcin Vitamin D responsive element indicate differences in protein/DNA recognition. For example, the V1 complex interacts with both steroid half-elements, whereas the V2 complex appears to recognize the proximal half-element. Our findings suggest variations in protein/protein and protein/DNA interactions of the VDR and RXR related complexes V1 and V2 at the osteocalcin Vitamin D responsive element that reflect unique properties of the osteosarcoma and normal diploid osteoblast phenotype.
骨特异性骨钙素基因的维生素D反应性转录在骨肉瘤细胞和正常二倍体成骨细胞中存在显著差异。在成骨细胞中，骨钙素基因仅在增殖后细胞中被转录并受维生素D上调，但在骨肉瘤细胞中表达是组成性的。骨钙素基因转录调控的这种差异与在骨钙素启动子的维生素D反应元件处两种主要的维生素D依赖性蛋白质/DNA复合物（称为V1和V2）的相对含量的显著差异相关。两种复合物的形成均依赖维生素D，且它们包含维生素D受体以及一种与RXR相关的蛋白质。孔径排阻和沉降速度分析表明，V1和V2复合物代表寡聚蛋白组装体（分别为四聚体和三聚体），并且主要反映了在骨钙素维生素D反应元件处单体蛋白成分的DNA导向结合。对骨钙素维生素D反应元件处的V1和V2复合物进行紫外线交联和甲基化干扰分析表明蛋白质/DNA识别存在差异。例如，V1复合物与两个类固醇半元件相互作用，而V2复合物似乎识别近端半元件。我们的研究结果表明，在骨钙素维生素D反应元件处，VDR和与RXR相关的复合物V1和V2的蛋白质/蛋白质和蛋白质/DNA相互作用存在差异，这反映了骨肉瘤和正常二倍体成骨细胞表型的独特性质。
