Böhnlein E, Gruss P
Mol Cell Biol. 1986 May;6(5):1401-11. doi: 10.1128/mcb.6.5.1401-1411.1986.
The interaction between cellular factors and polyoma virus (Py) DNA was investigated by using a gel retention assay. Nuclear extracts from various cell lines (NIH 3T3, NIH 3T6, LTK-, F9) contained proteins that formed specific and distinct complexes with Py B enhancer fragments of either wild-type or F9-1 mutant origin. The presence of an excess amount of other well-characterized DNA sequences, including the Py A enhancer, the murine sarcoma virus enhancer, and the simian virus 40 enhancer-promoter region, did not interfere with this protein-DNA interaction. However, a fragment previously defined as containing the lymphotropic papovavirus enhancer shares the binding of some common factor. This observation, in combination with the results of retention gel assays at different Mg2+ concentrations, indicates the interaction of several nuclear factors and Py DNA. The assay systems that were used allowed a distinction between some factors on the basis of their different biochemical and sequence requirements. The contact sites of these complexes were mapped to the B enhancer region of Py with Bal 31-derived mutant restriction fragments and ExoIII nuclease and are compatible with the functional domains determined in vivo.
利用凝胶滞留试验研究了细胞因子与多瘤病毒（Py）DNA之间的相互作用。来自各种细胞系（NIH 3T3、NIH 3T6、LTK-、F9）的核提取物中含有与野生型或F9-1突变型来源的Py B增强子片段形成特异性且不同复合物的蛋白质。过量存在的其他特征明确的DNA序列，包括Py A增强子、鼠肉瘤病毒增强子和猿猴病毒40增强子-启动子区域，均不干扰这种蛋白质-DNA相互作用。然而，一个先前定义为含有嗜淋巴细胞乳头瘤病毒增强子的片段共享一些共同因子的结合。这一观察结果与在不同Mg2+浓度下的滞留凝胶试验结果相结合，表明了几种核因子与Py DNA之间的相互作用。所使用的检测系统能够根据一些因子不同的生化和序列要求对它们进行区分。利用Bal 31衍生的突变限制片段和ExoIII核酸酶将这些复合物的接触位点定位到Py的B增强子区域，并且与体内确定的功能域一致。
