Unk I, Kiss-Toth E, Boros I
Institute of Biochemistry, Hungarian Academy of Sciences, Szeged.
Nucleic Acids Res. 1994 Nov 25;22(23):4872-5. doi: 10.1093/nar/22.23.4872.
Three 21bp repeats can be found in the bovine leukemia virus long terminal repeat, which are crucial for the LTR directed gene expression by the trans activator protein Tax. Previous studies demonstrated that the major target of the Tax directed activation are the CRE-like elements in the center of these repeats. In this work we report that another motif of the 21bp repeats is also required for the Tax activation. Gel retardation--with the wild type or mutant 21bp repeats--revealed that cellular factors from HeLa cells were specifically bound to the center (CRE-like element) and the 3' region of the repeats, which contains a CAGCTG consensus AP-4 binding site. In vivo analysis using the synthetic 21bp repeats indicated that beyond the consensus CRE-like motif, the AP-4 site is also essential for Tax activation. To determine the role of AP-4 in BLV Tax trans activation, we used the AP-4 cDNA in antisense transient assays. In the in vivo experiments the antisense AP-4 RNA resulted in strongly decreased Tax activation. On the basis of these results we conclude that AP-4 is a good candidate of cellular factors involved in BLV Tax trans activation.
在牛白血病病毒长末端重复序列中可发现三个21bp的重复序列，它们对于反式激活蛋白Tax介导的LTR定向基因表达至关重要。先前的研究表明，Tax介导激活的主要靶点是这些重复序列中心的CRE样元件。在本研究中，我们报告21bp重复序列的另一个基序对于Tax激活也是必需的。凝胶阻滞实验（使用野生型或突变型21bp重复序列）表明，来自HeLa细胞的细胞因子特异性结合到重复序列的中心（CRE样元件）和3'区域，该区域包含一个CAGCTG共有AP-4结合位点。使用合成的21bp重复序列进行的体内分析表明，除了共有CRE样基序外，AP-4位点对于Tax激活也是必不可少的。为了确定AP-4在BLV Tax反式激活中的作用，我们在反义瞬时分析中使用了AP-4 cDNA。在体内实验中，反义AP-4 RNA导致Tax激活显著降低。基于这些结果，我们得出结论，AP-4是参与BLV Tax反式激活的细胞因子的一个良好候选者。
